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  • 1.
    Adler, Jeremy
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi.
    Parmryd, Ingela
    Univ Gothenburg, Sahlgrenska Acad, Inst Biomed, Ingela Parmryd, Box 440, S-40530 Gothenburg, Sweden.
    Quantifying colocalization: the MOC is a hybrid coefficient - an uninformative mix of co-occurrence and correlation2019Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 132, nr 1, artikel-id UNSP jcs222455Artikel i tidskrift (Övrigt vetenskapligt)
  • 2. Agostinho, A.
    et al.
    Kouznetsova, A.
    Hernández-Hernández, A.
    Bernhem, Kristoffer
    KTH, Skolan för teknikvetenskap (SCI), Tillämpad fysik. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Blom, Hans
    KTH, Skolan för teknikvetenskap (SCI), Tillämpad fysik, Biofysik. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Brismar, Hjalmar
    KTH, Skolan för teknikvetenskap (SCI), Tillämpad fysik, Biofysik. KTH, Centra, Science for Life Laboratory, SciLifeLab.
    Höög, C.
    Sexual dimorphism in the width of the mouse synaptonemal complex2018Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 131, nr 5, artikel-id jcs212548Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Sexual dimorphism has been used to describe morphological differences between the sexes, but can be extended to any biologically related process that varies between males and females. The synaptonemal complex (SC) is a tripartite structure that connects homologous chromosomes in meiosis. Here, aided by superresolution microscopy techniques, we show that the SC is subject to sexual dimorphism, in mouse germ cells. We have identified a significantly narrower SC in oocytes and have established that this difference does not arise from a different organization of the lateral elements nor from a different isoform of transverse filament protein SYCP1. Instead, we provide evidence for the existence of a narrower central element and a different integration site for the C-termini of SYCP1, in females. In addition to these female-specific features, we speculate that post-translation modifications affecting the SYCP1 coiled-coil region could render a more compact conformation, thus contributing to the narrower SC observed in females.

  • 3.
    Alfredsson-Timmins, Jenny
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Henningson, Frida
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Bjerling, Pernilla
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    The Clr4 methyltransferase determines the subnuclear localization of the mating-type region in fission yeast2007Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 120, nr 11, s. 1935-1943Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The genome has a non-random spatial distribution in the cell nucleus. In Schizosaccharomyces pombe, it has been shown that the centromeres, telomeres and the mating-type region localize to the nuclear membrane (NM), the former by attaching to the spindle pole body (SPB). In addition, reporter genes inserted into these areas are transcriptionally repressed due to the formation of specialized chromatin structures. Performing live cell analysis we found that in a wild-type strain the mating-type region was positioned in the proximity of the SPB, the location where the pericentromeric heterochromatin is also found. In a strain lacking the histone methyltransferase, Clr4, crucial for the formation of heterochromatin, the mating-type region had a random localization in the nucleus. Moreover, in a strain where the two boundary elements IR-L and IR-R had been deleted the mating-type region was displaced from its position at the proximity of the SPB, but remained in the vicinity of the NM. Moreover, in all investigated strains with silencing deficiencies the distance between the mating-type region and the SPB increased. This result indicates a correlation between transcriptional derepression and displacement of the region. Two different models of how the mating-type chromatin is organized in the nucleus are discussed.

  • 4.
    Appelgren, Henrik
    et al.
    Södertörns högskola, Avdelning Naturvetenskap. Karolinska Institute.
    Kniola, Barbara
    Södertörns högskola, Avdelning Naturvetenskap. Karolinska Institute.
    Ekwall, Karl
    Södertörns högskola, Avdelning Naturvetenskap. Karolinska Institute.
    Distinct centromere domain structures with separate functions demonstrated in live fission yeast cells2003Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 116, nr 19, s. 4035-4042Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Fission yeast (Saccharomyces pombe) centromere DNA is organized in a central core region flanked on either side by a region of outer repeat (otr) sequences. The otr region is known to be heterochromatic and bound by the Swi6 protein whereas the central core region contains an unusual chromatin structure involving the histone H3 variant Cnp1 (S. pombe CENP-A). The central core is the base for formation of the kinetochore structure whereas the flanking region is important for sister centromere cohesion. We have previously shown that the ultrastructural domain structure of S. pombe centromeres in interphase is similar to that of human centromeres. Here we demonstrate that S. pombe centromeres are organized in cytologically distinct domains even in mitosis. Fluorescence in situ hybridization of fixed metaphase cells revealed that the otr regions of the centromere were still held together by cohesion even after the sister kinetochores had separated. In live cells, the central cores and kinetochores of sister chromosomes could be distinguished from one Another when they were subjected to mitotic tension. The function of the different centromeric domains was addressed. Transacting mutations affecting the kinetochore (nuf2) central core domain (mis6) and the heterochromatin domain (rik1) were analyzed in live cells. In interphase, both nuf2 and mis6 caused declustering of centromeres from the spindle pole body whereas centromere clustering was normal in rik1 despite an apparent decondensation defect. The declustering of centromeres in mis6 cells correlated with loss the Ndc80 kinetochore marker protein from the centromeres. Interestingly the declustered centromeres were still restricted to the nuclear periphery thus revealing a kinetochore-independent peripheral localization mechanism for heterochromatin. Time-lapse microscopy of live mis6 and nuf2-1 mutant cells in mitosis showed similar severe misaggregation phenotypes whereas the rik1 mutants showed a mild cohesion defect. Thus, S. pombe centromeres have two distinguishable domains even during mitosis, and our functional analyses support the previous observations that the kinetochore/central core and the heterochromatin domains have distinct functions both in interphase and mitosis.

  • 5.
    Appelqvist, Hanna
    et al.
    Linköpings universitet, Institutionen för fysik, kemi och biologi, Kemi. Linköpings universitet, Tekniska högskolan.
    Wäster, Petra
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Eriksson, Ida
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet.
    Rosdahl, Inger
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för inflammationsmedicin. Linköpings universitet, Hälsouniversitetet. Östergötlands Läns Landsting, Hjärt- och Medicincentrum, Hudkliniken i Östergötland.
    Öllinger, Karin
    Linköpings universitet, Institutionen för klinisk och experimentell medicin, Avdelningen för cellbiologi. Linköpings universitet, Hälsouniversitetet. Östergötlands Läns Landsting, Diagnostikcentrum, Klinisk patologi och klinisk genetik.
    Lysosomal exocytosis and caspase-8-mediated apoptosis in UVA-irradiated keratinocytes2013Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 126, nr 24, s. 5578-5584Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Ultraviolet (UV) irradiation is a major environmental carcinogen involved in the development of skin cancer. To elucidate the initial signaling during UV-induced damage in human keratinocytes, we investigated lysosomal exocytosis and apoptosis induction. UVA, but not UVB, induced plasma membrane damage, which was repaired by Ca2+-dependent lysosomal exocytosis. The lysosomal exocytosis resulted in extracellular release of cathepsin D and acid sphingomyelinase (aSMase). Two hours after UVA irradiation, we detected activation of caspase-8, which was reduced by addition of anti-aSMAse. Furthermore, caspase-8 activation and apoptosis was reduced by prevention of endocytosis and by the use of cathepsin inhibitors. We conclude that lysosomal exocytosis is part of the keratinocyte response to UVA and is followed by cathepsin-dependent activation of caspase-8. The findings have implications for the understanding of UV-induced skin damage and emphasize that UVA and UVB initiate apoptosis through different signaling pathways in keratinocytes.

  • 6.
    Arabi, Azadeh
    et al.
    Södertörns högskola, Avdelning Naturvetenskap. Karolinska Institute.
    Rustum, Cecilia
    Södertörns högskola, Avdelning Naturvetenskap. Stockholm University.
    Hallberg, Einar
    Södertörns högskola, Avdelning Naturvetenskap.
    Wright, Anthony P H
    Södertörns högskola, Avdelning Naturvetenskap. Karolinska Institute.
    Accumulation of c-Myc and proteasomes at the nucleoli of cells containing elevated c-Myc protein levels2003Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 116, nr 9, s. 1707-1717Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    c-Myc is a predominately nuclear transcription factor that is a substrate for rapid turnover by the proteasome system. Cancer-related mutations in c-Myc lead to defects in its degradation and thereby contribute to the increase in its cellular level that is associated with the disease. Little is known about the mechanisms that target c-Myc to the proteasomes. By using a GFP fusion protein and live analysis we show that c-Myc shuttles between the nucleus and cytoplasm and thus it could be degraded in either compartment. Strikingly, at elevated levels of expression c-Myc accumulates at nucleoli in some cells, consistent with saturation of a nucleolus-associated degradation system in these cells. This idea is further supported by the observation that proteasome inhibitor treatment causes accumulation of c-Myc at the nucleoli of essentially all cells. Under these conditions c-Myc is relatively stably associated with the nucleolus, as would be expected if the nucleolus functions as a sequestration/degradation site for excess c-Myc. Furthermore, during elevated c-Myc expression or proteasome inhibition, nucleoli that are associated with c-Myc also accumulate proteasomes. c-Myc and proteasomes co-localise in intranucleolar regions distinct from the dense fibrillar component of the nucleolus. Based on these results we propose a model for c-Myc downregulation where c-Myc is sequestered at the nucleoli. Sequestration of c-Myc is accompanied by recruitment of proteasomes and may lead to subsequent degradation.

  • 7.
    Barg, Sebastian
    et al.
    Department of Physiological Sciences, Lund University, Lund.
    Huang, Ping
    University of Chicago, Department of Neurobiology, Pharmacology and Physiology, Chicago.
    Eliasson, Lena
    Department of Physiological Sciences, Lund University, Lund.
    Nelson, Deborah J
    University of Chicago, Department of Neurobiology, Pharmacology and Physiology, Chicago.
    Obermüller, Stefanie
    Department of Physiological Sciences, Lund University, Lund.
    Rorsman, Patrik
    Department of Physiological Sciences, Lund University, Lund.
    Thévenod, Frank
    Physiologisches Institut, Universität des Saarlandes, Homburg/Saar.
    Renström, Erik
    Department of Physiological Sciences, Lund University, Lund.
    Priming of insulin granules for exocytosis by granular Cl(-) uptake and acidification2001Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 114, nr Pt 11, s. 2145-54Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    ATP-dependent priming of the secretory granules precedes Ca(2+)-regulated neuroendocrine secretion, but the exact nature of this reaction is not fully established in all secretory cell types. We have further investigated this reaction in the insulin-secreting pancreatic B-cell and demonstrate that granular acidification driven by a V-type H(+)-ATPase in the granular membrane is a decisive step in priming. This requires simultaneous Cl(-) uptake through granular ClC-3 Cl(-) channels. Accordingly, granule acidification and priming are inhibited by agents that prevent transgranular Cl(-) fluxes, such as 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) and an antibody against the ClC-3 channels, but accelerated by increases in the intracellular ATP:ADP ratio or addition of hypoglycemic sulfonylureas. We suggest that this might represent an important mechanism for metabolic regulation of Ca(2+)-dependent exocytosis that is also likely to be operational in other secretory cell types.

  • 8.
    Behm, Mikaela
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Wahlstedt, Helene
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Widmark, Albin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Eriksson, Maria
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Öhman, Marie
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Accumulation of nuclear ADAR2 regulates A-to-I RNA editing during neuronal development2017Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 130, s. 745-753Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Adenosine to inosine (A-to-I) RNA editing is important for a functional brain, and most known sites that are subject to selective RNA editing have been found to result in diversified protein isoforms that are involved in neurotransmission. In the absence of the active editing enzymes ADAR1 or ADAR2 (also known as ADAR and ADARB1, respectively), mice fail to survive until adulthood. Nuclear A-to-I editing of neuronal transcripts is regulated during brain development, with low levels of editing in the embryo and a dramatic increase after birth. Yet, little is known about the mechanisms that regulate editing during development. Here, we demonstrate lower levels of ADAR2 in the nucleus of immature neurons than in mature neurons. We show that importin-a4 (encoded by Kpna3), which increases during neuronal maturation, interacts with ADAR2 and contributes to the editing efficiency by bringing it into the nucleus. Moreover, we detect an increased number of interactions between ADAR2 and the nuclear isomerase Pin1 as neurons mature, which contribute to ADAR2 protein stability. Together, these findings explain how the nuclear editing of substrates that are important for neuronal function can increase as the brain develops. 

  • 9. Bembenek, Joshua N.
    et al.
    Meshik, Xenia
    Tsarouhas, Vasilios
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Meeting report - Cellular dynamics: membrane-cytoskeleton interface2017Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 130, nr 17, s. 2775-2779Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The first ever 'Cellular Dynamics' meeting on the membrane-cytoskeleton interface took place in Southbridge, MA on May 21-24, 2017 and was co-organized by Michael Way, Elizabeth Chen, Margaret Gardel and Jennifer Lippincott-Schwarz. Investigators from around the world studying a broad range of related topics shared their insights into the function and regulation of the cytoskeleton and membrane compartments. This provided great opportunities to learn about key questions in various cellular processes, from the basic organization and operation of the cell to higher-order interactions in adhesion, migration, metastasis, division and immune cell interactions in different model organisms. This unique and diverse mix of research interests created a stimulating and educational meeting that will hopefully continue to be a successful meeting for years to come.

  • 10. Berger, Susanne
    et al.
    Schäfer, Gritt
    Kesper, Dörthe A
    Holz, Anne
    Eriksson, Therese
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Palmer, Ruth H
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Beck, Lothar
    Klämbt, Christian
    Renkawitz-Pohl, Renate
    Onel, Susanne-Filiz
    WASP and SCAR have distinct roles in activating the Arp2/3 complex during myoblast fusion2008Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 121, nr Pt 8, s. 1303-1313Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Myoblast fusion takes place in two steps in mammals and in Drosophila. First, founder cells (FCs) and fusion-competent myoblasts (FCMs) fuse to form a trinucleated precursor, which then recruits further FCMs. This process depends on the formation of the fusion-restricted myogenic-adhesive structure (FuRMAS), which contains filamentous actin (F-actin) plugs at the sites of cell contact. Fusion relies on the HEM2 (NAP1) homolog Kette, as well as Blow and WASP, a member of the Wiskott-Aldrich-syndrome protein family. Here, we show the identification and characterization of schwächling--a new Arp3-null allele. Ultrastructural analyses demonstrate that Arp3 schwächling mutants can form a fusion pore, but fail to integrate the fusing FCM. Double-mutant experiments revealed that fusion is blocked completely in Arp3 and wasp double mutants, suggesting the involvement of a further F-actin regulator. Indeed, double-mutant analyses with scar/WAVE and with the WASP-interacting partner vrp1 (sltr, wip)/WIP show that the F-actin regulator scar also controls F-actin formation during myoblast fusion. Furthermore, the synergistic phenotype observed in Arp3 wasp and in scar vrp1 double mutants suggests that WASP and SCAR have distinct roles in controlling F-actin formation. From these findings we derived a new model for actin regulation during myoblast fusion.

  • 11. Bhattacharya, Resham
    et al.
    Kwon, Junhye
    Li, Xiujuan
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för genetik och patologi.
    Wang, Enfeng
    Patra, Sujata
    Bida, John Paul
    Bajzer, Zeljko
    Claesson-Welsh, Lena
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för genetik och patologi.
    Mukhopadhyay, Debabrata
    Distinct role of PLC{beta}3 in VEGF-mediated directional migration and vascular sprouting2009Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 122, nr 7, s. 1025-1034Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Endothelial cell proliferation and migration is essential to angiogenesis. Typically, proliferation and chemotaxis of endothelial cells is driven by growth factors such as vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). VEGF activates phospholipases (PLCs) - specifically PLCgamma1 - that are important for tubulogenesis, differentiation and DNA synthesis. However, we show here that VEGF, specifically through VEGFR2, induces phosphorylation of two serine residues on PLCbeta3, and this was confirmed in an ex vivo embryoid body model. Knockdown of PLCbeta3 in HUVEC cells affects IP3 production, actin reorganization, migration and proliferation; whereas migration is inhibited, proliferation is enhanced. Our data suggest that enhanced proliferation is precipitated by an accelerated cell cycle, and decreased migration by an inability to activate CDC42. Given that PLCbeta3 is typically known as an effector of heterotrimeric G-proteins, our data demonstrate a unique crosstalk between the G-protein and receptor tyrosine kinase (RTK) axes and reveal a novel molecular mechanism of VEGF signaling and, thus, angiogenesis.

  • 12. Bidla, Gawa
    et al.
    Dushay, Mitchell S.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för fysiologi och utvecklingsbiologi, Jämförande fysiologi.
    Theopold, Ulrich
    Crystal cell rupture after injury in Drosophila requires the JNK pathway, small GTPases and the TNF homolog eiger2007Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 120, nr 7, s. 1209-1215Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The prophenoloxidase-activating cascade is a key component of arthropod immunity. Drosophila prophenoloxidase is stored in crystal cells, a specialized class of blood cells from which it is released through cell rupture. Within minutes after bleeding, prophenoloxidase is activated leading to visible melanization of the clot matrix. Using crystal cell rupture and melanization as readouts to screen mutants in signal transduction pathways, we show that prophenoloxidase release requires Jun N-terminal kinase, small Rho GTPases and Eiger, the Drosophila homolog of tumor necrosis factor. We also provide evidence that in addition to microbial products, endogenous signals from dying hemocytes contribute to triggering and/or assembly of the prophenoloxidase-activating cascade, and that this process can be inhibited in vitro and in vivo using the viral apoptotic inhibitor p35. Our results provide a more comprehensive view of immune signal transduction pathways, with implications for immune reactions where cell death is used as a terminal mode of cell activation.

  • 13.
    Bidla, Gawa
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Dushay, Mitchell S.
    Theopold, Ulrich
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Crystal cell rupture after injury in Drosophila requires the JNK pathway, small GTPases and the TNF homolog Eiger2007Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 120, nr 7, s. 1209-15Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The prophenoloxidase-activating cascade is a key component of arthropod immunity. Drosophila prophenoloxidase is stored in crystal cells, a specialized class of blood cells from which it is released through cell rupture. Within minutes after bleeding, prophenoloxidase is activated leading to visible melanization of the clot matrix. Using crystal cell rupture and melanization as readouts to screen mutants in signal transduction pathways, we show that prophenoloxidase release requires Jun N-terminal kinase, small Rho GTPases and Eiger, the Drosophila homolog of tumor necrosis factor. We also provide evidence that in addition to microbial products, endogenous signals from dying hemocytes contribute to triggering and/or assembly of the prophenoloxidase-activating cascade, and that this process can be inhibited in vitro and in vivo using the viral apoptotic inhibitor p35. Our results provide a more comprehensive view of immune signal transduction pathways, with implications for immune reactions where cell death is used as a terminal mode of cell activation.

  • 14. Boban, Mirta
    et al.
    Pantazopoulou, Marina
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Schick, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Ljungdahl, Per O.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Foisner, Roland
    A nuclear ubiquitin-proteasome pathway targets the inner nuclear membrane protein Asi2 for degradation2014Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 127, nr 16, s. 3603-3613Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The nuclear envelope consists of inner and outer nuclear membranes. Whereas the outer membrane is an extension of the endoplasmic reticulum, the inner nuclear membrane (INM) represents a unique membranous environment containing specific proteins. The mechanisms of integral INM protein degradation are unknown. Here, we investigated the turnover of Asi2, an integral INM protein in Saccharomyces cerevisiae. We report that Asi2 is degraded by the proteasome independently of the vacuole and that it exhibited a half-life of similar to 45 min. Asi2 exhibits enhanced stability in mutants lacking the E2 ubiquitin conjugating enzymes Ubc6 or Ubc7, or the E3 ubiquitin ligase Doa10. Consistent with these data, Asi2 is post-translationally modified by poly-ubiquitylation in a Ubc7- and Doa10-dependent manner. Importantly Asi2 degradation is significantly reduced in a sts1-2 mutant that fails to accumulate proteasomes in the nucleus, indicating that Asi2 is degraded in the nucleus. Our results reveal a molecular pathway that affects the stability of integral proteins of the inner nuclear membrane and indicate that Asi2 is subject to protein quality control in the nucleus.

  • 15. Buch, Charlotta
    et al.
    Lindberg, Robert
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Södertörn University, Sweden.
    Figueroa, Ricardo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Södertörn University, Sweden.
    Gudise, Santhosh
    Onischenko, Evgeny
    Hallberg, Einar
    An integral protein of the inner nuclear membrane localizes to the mitotic spindle in mammalian cells2009Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 122, nr 12, s. 2100-2107Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Here, we characterize a transmembrane protein of the nuclear envelope that we name spindle-associated membrane protein 1 (Samp1). The protein is conserved in metazoa and fission yeast and is homologous to Net5 in rat and Ima1 in Schizosaccharomyces pombe. We show that, in human cells, the protein is a membrane-spanning polypeptide with an apparent molecular mass of 43 kDa. This is consistent with a predicted polypeptide of 392 amino acids that has five transmembrane segments and its C-terminus exposed to the nucleoplasm. During interphase, Samp1 was specifically distributed in the inner nuclear membrane. Post-transcriptional silencing of Samp1 expression resulted in separation of centrosomes from the nuclear envelope, indicating that it is functionally connected to the cytoskeleton. At the onset of mitosis, most of the protein dispersed out into the ER, as expected. However, during mitosis, a significant fraction of the protein specifically localized to the polar regions of the mitotic spindle. We demonstrate for the first time, in human cells, the existence of a membranous structure overlapping with the mitotic spindle. Interestingly, another integral inner nuclear membrane protein, emerin, was absent from the spindle-associated membranes. Thus, Samp1 defines a specific membrane domain associated with the mitotic spindle.

  • 16.
    Buch, Charlotta
    et al.
    Södertörns högskola, Institutionen för livsvetenskaper.
    Lindberg, Robert
    Södertörns högskola, Institutionen för livsvetenskaper.
    Figueroa, Ricardo
    Södertörns högskola, Institutionen för livsvetenskaper.
    Gudise, Santhosh
    Södertörns högskola, Institutionen för livsvetenskaper.
    Onischenko, Evgeny
    Hallberg, Einar
    Södertörns högskola, Institutionen för livsvetenskaper.
    An integral protein of the inner nuclear membrane localizes to the mitotic spindle in mammalian cells2009Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 122, nr 12, s. 2100-2107Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Here, we characterize a transmembrane protein of the nuclear envelope that we name spindle-associated membrane protein 1 (Samp1). The protein is conserved in metazoa and fission yeast and is homologous to Net5 in rat and Ima1 in Schizosaccharomyces pombe. We show that, in human cells, the protein is a membrane-spanning polypeptide with an apparent molecular mass of 43 kDa. This is consistent with a predicted polypeptide of 392 amino acids that has five transmembrane segments and its C-terminus exposed to the nucleoplasm. During interphase, Samp1 was specifically distributed in the inner nuclear membrane. Post-transcriptional silencing of Samp1 expression resulted in separation of centrosomes from the nuclear envelope, indicating that it is functionally connected to the cytoskeleton. At the onset of mitosis, most of the protein dispersed out into the ER, as expected. However, during mitosis, a significant fraction of the protein specifically localized to the polar regions of the mitotic spindle. We demonstrate for the first time, in human cells, the existence of a membranous structure overlapping with the mitotic spindle. Interestingly, another integral inner nuclear membrane protein, emerin, was absent from the spindle-associated membranes. Thus, Samp1 defines a specific membrane domain associated with the mitotic spindle.

  • 17. Busayavalasa, Kiran
    et al.
    Chen, Xin
    Östlund Farrants, Ann-Kristin
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut, Avdelningen för cellbiologi.
    Wagner, Nicole
    Sabri, Nafiseh
    The nup155 mediated organisation of inner nuclear membrane proteins is independent of nup155 anchoring to the metazoan nuclear pore complex2012Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 125, nr 18, s. 4214-4218Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The nuclear envelope (NE), an important barrier between the nucleus and the cytoplasm, is composed of three structures: the outer nuclear membrane, which is continuous with the ER, the inner nuclear membrane (INM), which interfaces with chromatin, and nuclear pore complexes (NPCs), which are essential for the exchange of macromolecules between the two compartments. The NPC protein Nup155 has an evolutionarily conserved role in the metazoan NE formation; but the in vivo analysis of Nup155 has been severely hampered by the essential function of this protein in cell viability. Here, we take advantage of the hypomorphicity of RNAi systems and use a combination of protein binding and rescue assays to map the interaction sites of two neighbouring NPC proteins Nup93 and Nup53 on Nup155, and to define the requirements of these interactions in INM protein organization. We show that different parts of Drosophila Nup155 have distinct functions: the Nup155 beta-propeller anchors the protein to the NPC, whereas the alpha-solenoid part of Nup155 is essential for the correct localisation of INM proteins lamin-B receptor (LBR) and otefin. Using chromatin extracts from semisynchronized cells, we also provide evidence that the Nup155 alpha-solenoid has a chromatin-binding activity that is stronger at the end of mitosis. Our results argue that the role of Nup155 in INM protein localisation is not mediated through the NPC anchoring activity of the protein and suggest that regions other than Nup155 beta-propeller are necessary for the targeting of proteins to the INM.

  • 18.
    Carlsson, Sven R
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Simonsen, Anne
    Membrane dynamics in autophagosome biogenesis2015Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 128, nr 2, s. 193-205Artikel i tidskrift (Övrigt vetenskapligt)
    Abstract [en]

    Bilayered phospholipid membranes are vital to the organization of the living cell. Based on fundamental principles of polarity, membranes create borders allowing defined spaces to be encapsulated. This compartmentalization is a prerequisite for the complex functional design of the eukaryotic cell, yielding localities that can differ in composition and operation. During macroautophagy, cytoplasmic components become enclosed by a growing double bilayered membrane, which upon closure creates a separate compartment, the autophagosome. The autophagosome is then primed for fusion with endosomal and lysosomal compartments, leading to degradation of the captured material. A large number of proteins have been found to be essential for autophagy, but little is known about the specific lipids that constitute the autophagic membranes and the membrane modeling events that are responsible for regulation of autophagosome shape and size. In this Commentary, we review the recent progress in our understanding of the membrane shaping and remodeling events that are required at different steps of the autophagy pathway.

  • 19. Carolina Touz, Maria
    et al.
    Silvana Ropolo, Andrea
    Romina Rivero, Maria
    Veronica Vranych, Cecilia
    Conrad, John Thomas
    Svärd, Staffan Gunnar
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Nash, Theodore Elliott
    Arginine deiminase has multiple regulatory roles in the biology of Giardia lamblia2008Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 121, nr 17, s. 2930-2938Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The protozoan parasite Giardia lamblia uses arginine deiminase (ADI) to produce energy from free L-arginine under anaerobic conditions. In this work, we demonstrate that, in addition to its known role as a metabolic enzyme, it also functions as a peptidylarginine deiminase, converting protein-bound arginine into citrulline. G. lamblia ADI specifically binds to and citrullinates the arginine in the conserved CRGKA tail of variant-specific surface proteins (VSPs), affecting both antigenic switching and antibody-mediated cell death. During encystation, ADI translocates from the cytoplasm to the nuclei and appears to play a regulatory role in the expression of encystation-specific genes. ADI is also sumoylated, which might modulate its activity. Our findings reveal a dual role played by ADI and define novel regulatory pathways used by Giardia for survival.

  • 20.
    Dinic, Jelena
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut.
    Parmryd, Ingela
    Uppsala universitet, Institutionen för medicinsk cellbiologi.
    Actin filaments at the plasma membrane in live cells cause the formation of ordered lipid domains via phosphatidylinositol 4,5-bisphosphateIngår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The relationship between ordered plasma membrane nanodomains, known as lipid rafts, and actin filaments is the focus of this study. Plasma membrane order was followed in live cells at 37°C using laurdan and di-4-ANEPPDHQ to report on lipid packing. Disrupting actin polymerization decreased the fraction of ordered domains, which strongly argue that unstimulated cells have a basal level of ordered domains. Stabilising actin filaments had the opposite effect and increased the proportion of ordered domains. Decreasing the plasma membrane level of phosphatidylinositol 4,5-bisphosphate lowers the number of attachment points for actin filaments and reduced the proportion of ordered domains. Aggregation of plasma membrane molecules, both lipid raft and non-lipid raft markers, leads to the formation of ordered domains that is correlated with an increase in cell peripheral actin filaments. In membrane blebs, which are detached from the underlying actin filaments the fraction of ordered domains was low and GM1 could not be patched to form ordered domains. We conclude that ordered domains form where actin filaments attach to the plasma membrane via phosphatidylinositol 4,5-bisphosphate. This downplays lipid-lipid interactions as the main driving force behind the formation of ordered membrane domains in vivo, giving greater prominence to membrane-intracellular filament interactions.

  • 21.
    Dyachok, Oleg
    et al.
    Uppsala universitet, Medicinska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
    Gylfe, Erik
    Uppsala universitet, Medicinska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
    Store-operated influx of Ca2+ in the pancreatic β-cells exhibits graded dependence on the filling of the endoplasmic reticulum2001Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 114, nr Pt 11, s. 2179-2186Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The store-operated pathway for Ca2+ entry was studied in individual mouse pancreatic β-cells by measuring the cytoplasmic concentrations of Ca2+ ([Ca2+]i) and Mn2+ ([Mn2+]i) with the fluorescent indicator fura-2. Influx through the store-operated pathway was initially shut off by pre-exposure to 20 mM glucose, which maximally stimulates intracellular Ca2+ sequestration. To avoid interference with voltage-dependent Ca2+ entry the cells were hyperpolarized with diazoxide and the channel blocker methoxyverapamil was present. Activation of the store-operated pathway in response to Ca2+ depletion of the endoplasmic reticulum was estimated from the sustained elevation of [Ca2+]i or from the rate of increase in [Mn2+]i due to influx of these extracellular ions. Increasing concentrations of the inositol 1,4,5-trisphosphate-generating agonist carbachol or the sarco(endo)plasmatic reticulum Ca2+-ATPase inhibitor cyclopiazonic acid (CPA) cause gradual activation of the store-operated pathway. In addition, the carbachol- and CPA-induced influx of Mn2+ depended on store filling in a graded manner. The store-operated influx of Ca2+/Mn2+ was inhibited by Gd3+ and 2-aminoethoxydiphenyl borate but neither of these agents discriminated between store-operated and voltage-dependent entry. The finely tuned regulation of the store-operated mechanisms in the β-cell has direct implications for the control of membrane potential and insulin secretion.

  • 22.
    Edlund, Sofia
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwiginstitutet för cancerforskning.
    Landström, Maréne
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwiginstitutet för cancerforskning.
    Heldin, Carl-Henrik
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwiginstitutet för cancerforskning.
    Aspenström, Pontus
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwiginstitutet för cancerforskning.
    Smad7 is required for TGF-ß-induced activation of the small GTPase Cdc422004Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 117, nr Pt 9, s. 1835-1847Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Transforming growth factor beta (TGF-beta) is a potent regulator of cell growth and differentiation in many cell types. The Smad signaling pathway constitutes a main signal transduction route downstream of TGF-beta receptors. The inhibitory Smads, Smad6 and Smad7, are considered to function as negative regulators of the TGF-beta/Smad signaling cascade. In a previous study, we found that TGF-beta induces rearrangements of the actin filament system in human prostate carcinoma cells and that this response requires the small GTPases Cdc42 and RhoA. On the basis of the current view on the function of Smad7 in TGF-beta signaling, we hypothesized that Smad7 would function as a negative regulator of the TGF-beta-induced activation of Cdc42 and RhoA, but instead we found that the reverse is the case; Smad7 is required for the TGF-beta-induced activation of Cdc42 and the concomitant reorganization of the actin filament system. These observations propose a novel role for Smad7 in TGF-beta-dependent activation of Rho GTPases.

  • 23.
    Fauvarque, Marie-Odile
    et al.
    CEA, Institut de Recherches en Technologies et Sciences pour le Vivant, Laboratoire de Biologie à Grande Echelle, F-38054 Grenoble, France.
    Williams, Michael J
    Institute of Biological and Environmental Sciences, University of Aberdeen, Aberdeen AB24 2TZ, UK .
    Drosophila cellular immunity: a story of migration and adhesion2011Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 124, nr 9, s. 1373-1382Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Research during the past 15 years has led to significant breakthroughs, providing evidence of a high degree of similarity between insect and mammalian innate immune responses, both humoural and cellular, and highlighting Drosophila melanogaster as a model system for studying the evolution of innate immunity. In a manner similar to cells of the mammalian monocyte and macrophage lineage, Drosophila immunosurveillance cells (haemocytes) have a number of roles. For example, they respond to wound signals, are involved in wound healing and contribute to the coagulation response. Moreover, they participate in the phagocytosis and encapsulation of invading pathogens, are involved in the removal of apoptotic bodies and produce components of the extracellular matrix. There are several reasons for using the Drosophila cellular immune response as a model to understand cell signalling during adhesion and migration in vivo: many genes involved in the regulation of Drosophila haematopoiesis and cellular immunity have been maintained across taxonomic groups ranging from flies to humans, many aspects of Drosophila and mammalian innate immunity seem to be conserved, and Drosophila is a simplified and well-studied genetic model system. In the present Commentary, we will discuss what is known about cellular adhesion and migration in the Drosophila cellular immune response, during both embryonic and larval development, and where possible compare it with related mechanisms in vertebrates.

  • 24. Fotin-Mleczek, Mariola
    et al.
    Henkler, Frank
    Samel, Dierk
    Reichwein, Monica
    Hausser, Angelika
    Parmryd, Ingela
    Scheurich, Peter
    Schmid, Johannes A
    Wajant, Harald
    Apoptotic crosstalk of TNF receptors: TNF-R2-induces depletion of TRAF2 and IAP proteins and accelerates TNF-R1-dependent activation of caspase-82002Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 115, nr Pt 13, s. 2757-70Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    We have recently shown that stimulation of TNF-R2 selectively enhances apoptosis induction by the death receptor TNF-R1. Here, we demonstrate that stimulation of CD30 or CD40 also leads to selective enhancement of TNF-R1-induced cell death. Enhancement of apoptosis was correlated with the depletion of endogenous TRAF2 within 1 to 6 hours. Selective prestimulation of TNF-R2 for several hours inhibited TNF-R2-induced activation of the anti-apoptotic NF-kappaB pathway up to 90% and dramatically enhanced apoptosis induction by this receptor. When both TNF-receptors were stimulated simultaneously, TNF-R1-induced NF-kappaB activation remained unaffected but TNF-R1-induced apoptosis was still significantly enhanced. Compared with FasL-induced cell death TNF-R1-induced activation of caspase-8 was significantly weaker and delayed. Costimulation or prestimulation of TNF-R2 enhanced caspase-8 processing. Life cell imaging and confocal microscopy revealed that both TNF-R1 and TNF-R2 recruited the anti-apoptotic factor cIAP1 in a TRAF2-dependent manner. Thus, TNF-R2 may compete with TNF-R1 for the recruitment of newly synthesized TRAF2-bound anti-apoptotic factors, thereby promoting the formation of a caspase-8-activating TNF-R1 complex. Hence, TNF-R2 triggering can interfere with TNF-R1-induced apoptosis by inhibition of NF-kappaB-dependent production of anti-apoptotic factors and by blocking the action of anti-apoptotic factors at the post-transcriptional level.

  • 25.
    Francis, Monika K.
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Holst, Mikkel R.
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Vidal-Quadras, Maite
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Henriksson, Sara
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Santarella-Mellwig, Rachel
    Sandblad, Linda
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Lundmark, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Endocytic membrane turnover at the leading edge is driven by a transient interaction between Cdc42 and GRAF12015Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 128, nr 22, s. 4183-4195Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Changes in cell morphology require coordination of plasma membrane turnover and cytoskeleton dynamics, processes that are regulated by Rho GTPases. Here, we describe how a direct interaction between the Rho GTPase Cdc42 and the GTPase activating protein (GAP) GRAF1, facilitate rapid cell surface turnover at the leading edge. Both Cdc42 and GRAF1 were required for fluid phase uptake and regulated the generation of transient GRAF1-coated endocytic carriers, distinct from clathrin coated vesicles. GRAF1 was found to transiently assemble at discrete Cdc42-enriched punctae at the plasma membrane resulting in a corresponding decrease in Cdc42 microdomain association. However, Cdc42 captured in its active state was, via a GAP domain mediated interaction, localised together with GRAF1 on accumulated internal structures derived from the cell surface. Correlative fluorescence and electron tomography microscopy revealed that these structures were clusters of small membrane carriers affected in their endosomal processing. We conclude that a transient interaction between Cdc42 and GRAF1 drives endocytic turnover and controls the transition essential for endosomal maturation of plasma membrane internalised by this mechanism.

  • 26.
    Griesche, Nadine
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
    Sanchez, Gonzalo
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
    Hermans, Cedric
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
    Idevall Hagren, Olof
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
    Cortical mitochondria regulate insulin secretion by local Ca2+ buffering in rodent beta cells2019Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 132, nr 9, artikel-id jcs228544Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Mitochondria play an essential role in regulating insulin secretion from beta cells by providing the ATP needed for the membrane depolarization that results in voltage-dependent Ca2+ influx and subsequent insulin granule exocytosis. Ca2+, in turn, is also rapidly taken up by the mitochondria and exerts important feedback regulation of metabolism. The aim of this study was to determine whether the distribution of mitochondria within beta cells is important for the secretory capacity of these cells. We find that cortically localized mitochondria are abundant in rodent beta cells, and that these mitochondria redistribute towards the cell interior following depolarization. The redistribution requires Ca2+-induced remodeling of the cortical F-actin network. Using light-regulated motor proteins, we increased the cortical density of mitochondria twofold and found that this blunted the voltage-dependent increase in cytosolic Ca2+ concentration and suppressed insulin secretion. The activity-dependent changes in mitochondria distribution are likely to be important for the generation of Ca2+ microdomains required for efficient insulin granule release.

  • 27.
    Gudise, Santhosh
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. Karolinska Institute (NOVUM), Sweden.
    Figueroa, Ricardo A.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lindberg, Robert
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Larsson, Veronica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Hallberg, Einar
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Samp1 is functionally associated with the LINC complex and A-type lamina networks2011Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 124, s. 2077-2085Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The transmembrane inner nuclear membrane (INM) protein Samp1 is required for anchoring centrosomes near the nuclei. Using high-resolution fluorescence microscopy we show that Samp1 is distributed in a distinct and characteristic pattern in the nuclear envelope (NE), where it partially colocalizes with the LINC complex protein Sun1. By studying the localization of Samp1 deletion mutants and fusion proteins, we conclude that the cysteine-rich N-terminal half of Samp1 is nucleoplasmically exposed and is responsible for targeting to the INM. It contains four conserved CxxC motifs with the potential to form zinc fingers. The distribution of cysteine-to-alanine substitution mutants, designed to prevent zinc finger formation, showed that NE localization of Samp1 depends on intact CxxC motifs. Overexpression of Samp1 zinc finger mutants produced an abnormal dominant phenotype characterized by disrupted organization of a selective subset NE proteins, including emerin, Sun1, endogenous Samp1 and, in some cases, lamin A/C, but not lamin B, Sun2 or nucleoporins. Silencing of Samp1 expression showed that emerin depends on Samp1 for its correct localization in the NE. Our results demonstrate that Samp1 is functionally associated with the LINC complex protein Sun1 and proteins of the A-type lamina network.

  • 28. Guerra, Lina
    et al.
    Guidi, Riccardo
    Slot, Ilse
    Callegari, Simone
    Sompallae, Ramakrishna
    Pickett, Carol L.
    Åström, Stefan
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut.
    Eisele, Frederik
    Wolf, Dieter
    Sjögren, Camilla
    Masucci, Maria G.
    Frisan, Teresa
    Bacterial genotoxin triggers FEN1-dependent RhoA activation, cytoskeleton remodeling and cell survival2011Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 124, nr 16, s. 2735-2742Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The DNA damage response triggered by bacterial cytolethal distending toxins (CDTs) is associated with activation of the actin-regulating protein RhoA and phosphorylation of the downstream-regulated mitogen-activated protein kinase (MAPK) p38, which promotes the survival of intoxicated (i.e. cells exposed to a bacterial toxin) cells. To identify the effectors of this CDT-induced survival response, we screened a library of 4492 Saccharomyces cerevisiae mutants that carry deletions in nonessential genes for reduced growth following inducible expression of CdtB. We identified 78 genes whose deletion confers hypersensitivity to toxin. Bioinformatics analysis revealed that DNA repair and endocytosis were the two most overrepresented signaling pathways. Among the human orthologs present in our data set, FEN1 and TSG101 regulate DNA repair and endocytosis, respectively, and also share common interacting partners with RhoA. We further demonstrate that FEN1, but not TSG101, regulates cell survival, MAPK p38 phosphorylation, RhoA activation and actin cytoskeleton reorganization in response to DNA damage. Our data reveal a previously unrecognized crosstalk between DNA damage and cytoskeleton dynamics in the regulation of cell survival, and might provide new insights on the role of chronic bacteria infection in carcinogenesis.

  • 29. Guerra, Lina
    et al.
    Guidi, Riccardo
    Slot, Ilse
    Callegari, Simone
    Sompallae, Ramakrishna
    Pickett, Carol L
    Åström, Stefan
    Eisele, Frederik
    Wolf, Dieter
    Sjögren, Camilla
    Masucci, Maria G
    Frisan, Teresa
    Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm.
    Bacterial genotoxin triggers FEN1-dependent RhoA activation, cytoskeleton remodeling and cell survival2011Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 124, nr 16, s. 2735-2742Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The DNA damage response triggered by bacterial cytolethal distending toxins (CDTs) is associated with activation of the actin-regulating protein RhoA and phosphorylation of the downstream-regulated mitogen-activated protein kinase (MAPK) p38, which promotes the survival of intoxicated (i.e. cells exposed to a bacterial toxin) cells. To identify the effectors of this CDT-induced survival response, we screened a library of 4492 Saccharomyces cerevisiae mutants that carry deletions in nonessential genes for reduced growth following inducible expression of CdtB. We identified 78 genes whose deletion confers hypersensitivity to toxin. Bioinformatics analysis revealed that DNA repair and endocytosis were the two most overrepresented signaling pathways. Among the human orthologs present in our data set, FEN1 and TSG101 regulate DNA repair and endocytosis, respectively, and also share common interacting partners with RhoA. We further demonstrate that FEN1, but not TSG101, regulates cell survival, MAPK p38 phosphorylation, RhoA activation and actin cytoskeleton reorganization in response to DNA damage. Our data reveal a previously unrecognized crosstalk between DNA damage and cytoskeleton dynamics in the regulation of cell survival, and might provide new insights on the role of chronic bacteria infection in carcinogenesis.

  • 30.
    Gustavsson, Anna
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Armulik, Annika
    Brakebusch, Cord
    Fässler, Reinhard
    Johansson, Staffan
    Fällman, Maria
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Role of the β1-integrin cytoplasmic tail in mediating invasin-promoted internalization of Yersinia2002Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 115, nr 13, s. 2669-2678Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Invasin of Yersinia pseudotuberculosis binds to beta1-integrins on host cells and triggers internalization of the bacterium. To elucidate the mechanism behind the beta1-integrin-mediated internalization of Yersinia, a beta1-integrin-deficient cell line, GD25, transfected with wild-type beta1A, beta1B or different mutants of the beta1A subunit was used. Both beta1A and beta1B bound to invasin-expressing bacteria, but only beta1A was able to mediate internalization of the bacteria. The cytoplasmic region of beta1A, differing from beta1B, contains two NPXY motifs surrounding a double threonine site. Exchanging the tyrosines of the two NPXYs to phenylalanines did not inhibit the uptake, whereas a marked reduction was seen when the first tyrosine (Y783) was exchanged to alanine. A similar reduction was seen when the two nearby threonines (TT788-9) were exchanged with alanines. It was also noted that cells affected in bacterial internalization exhibited reduced spreading capability when seeded onto invasin, suggesting a correlation between the internalization of invasin-expressing bacteria and invasin-induced spreading. Likewise, integrins defective in forming peripheral focal complex structures was unable to mediate uptake of invasin-expressing bacteria.

  • 31. Höög, Johanna L
    et al.
    Huisman, Stephen M
    Sebö-Lemke, Zsofia
    Sandblad, Linda
    European Mol Biol Labs, Cell Biol & Biophys Program, D-69117 Heidelberg, Germany .
    McIntosh, J Richard
    Antony, Claude
    Brunner, Damian
    Electron tomography reveals a flared morphology on growing microtubule ends2011Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 124, nr Pt 5, s. 693-698Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Microtubules (MTs) exhibit dynamic instability, alternating between phases of growth and shortening, mostly at their uncapped plus ends. Based on results from cryo-electron microscopy it was proposed that growing MTs display mainly curved sheets and blunt ends; during depolymerisation curled 'ramshorns' predominate. Observations of MTs in mitotic cells have suggested that the situation in vivo differs from that in vitro, but so far, a clear comparison between in vivo and in vitro results has not been possible because MT end structures could not be correlated directly with the dynamic state of that particular MT. Here we combine light microscopy and electron tomography (ET) to show that growing MT plus ends in the fission yeast Schizosaccharomyces pombe display predominantly a flared morphology. This indicates that MT polymerisation in vivo and in vitro can follow different paths.

  • 32. Imreh, Gabriela
    et al.
    Norberg, Helin Vakifahmetoglu
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut.
    Imreh, Stefan
    Zhivotovsky, Boris
    Chromosomal breaks during mitotic catastrophe trigger gamma H2AX-ATM-p53-mediated apoptosis2011Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 124, nr 17, s. 2951-2963Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Although the cause and outcome of mitotic catastrophe (MC) has been thoroughly investigated, precisely how the ensuing lethality is regulated during or following this process and what signals are involved remain unknown. Moreover, the mechanism of the decision of cell death modalities following MC is still not well characterised. We demonstrate here a crucial role of the gamma H2AX-ATM-p53 pathway in the regulation of the apoptotic outcome of MC resulting from cells entering mitosis with damaged DNA. In addition to p53 deficiency, the depletion of ATM (ataxia telangiectasia mutated), but not ATR ( ataxia telangiectasia and Rad3-related protein), protected against apoptosis and shifted cell death towards necrosis. Activation of this pathway is triggered by the augmented chromosomal damage acquired during anaphase in doxorubicin-treated cells lacking 4-3-3 sigma (also known as epithelial cell marker protein-1 or stratifin). Moreover, cells that enter mitosis with damaged DNA encounter segregation problems because of their abnormal chromosomes, leading to defects in mitotic exit, and they therefore accumulate in G1 phase. These multi- or micronucleated cells are prevented from cycling again in a p53- and p21-dependent manner, and subsequently die. Because increased chromosomal damage resulting in extensive H2AX phosphorylation appears to be a direct cause of catastrophic mitosis, our results describe a mechanism that involves generation of additional DNA damage during MC to eliminate chromosomally unstable cells.

  • 33.
    Jin, Shao-Bo
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Zhao, Jian
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Wieslander, Lars
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    CRM1 and Ran are present but a NES-CRM1-RanGTP complex is not required in Balbiani ring mRNP particles from the gene to the cytoplasm2004Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 117, s. 1553-1566Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Messenger RNA is formed from precursors known as pre-mRNA. Theseprecursors associate with proteins to form pre-mRNA-protein(pre-mRNP) complexes. Processing machines cap, splice and polyadenylatethe pre-mRNP and in this way build the mRNP. These processingmachines also affect the export of the mRNP complexes from thenucleus to the cytoplasm. Export to the cytoplasm takes placethrough a structure in the nuclear membrane called the nuclearpore complex (NPC). Export involves adapter proteins in themRNP and receptor proteins that bind to the adapter proteinsand to components of the NPC. We show that the export receptorchromosomal region maintenance protein 1 (CRM1), belonging toa family of proteins known as importin-ß-like proteins,binds to gene-specific Balbiani ring (BR) pre-mRNP while transcriptiontakes place. We also show that the GTPase known as Ran bindsto BR pre-mRNP, and that it binds mainly in the interchromatin.However, we also show using leptomycin B treatment that a NES-CRM1-RanGTPcomplex is not essential for export, even though both CRM1 andRan accompany the BR mRNP through the NPC. Our results thereforesuggest that several export receptors associate with BR mRNPand that these receptors have redundant functions in the nuclearexport of BR mRNP.

  • 34.
    Johansson, M
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för evolutionsbiologi, Fysiologisk botanik.
    Patarroyo, M
    Öberg, Fredrik
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för genetik och patologi.
    Siegbahn, Agneta
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinska vetenskaper, Klinisk kemi.
    Nilsson, Kenneth
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för genetik och patologi.
    Myeloperoxidase mediates cell adhesion via the alpha M beta 2 integrin (Mac-1, CD11b/CD18)1997Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 110, nr 9, s. 1133-1139Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Myeloperoxidase is a leukocyte component able to generate potent microbicidal substances. A homologous invertebrate blood cell protein, peroxinectin, is not only a peroxidase but also a cell adhesion ligand. We demonstrate in this study that human myeloperoxidase also mediates cell adhesion. Both the human myeloid cell line HL-60, when differentiated by treatment with 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or retinoic acid, and human blood leukocytes, adhered to myeloperoxidase; however, undifferentiated HL-60 cells showed only minimal adhesion. No cells adhered to horseradish peroxidase, and cell adhesion to myeloperoxidase was not decreased by catalase, thus showing that peroxidase activity, per se, was neither sufficient nor necessary for the adhesion activity. Mannan, which has been reported to inhibit the binding of peroxidases to cells, did not affect adhesion to myeloperoxidase. However, adhesion to myeloperoxidase was inhibited by monoclonal antibodies to alpha M (CD11b) or to beta2 (CD18) integrin subunits, but not by antibodies to alpha L (CD11a), alpha M (CD11c), or to other integrins. Native myeloperoxidase mediated dose-dependent cell adhesion down to relatively low concentrations, and denaturation abolished the adhesion activity. It is evident that myeloperoxidase supports cell adhesion, a function which may be of considerable importance for leukocyte migration and infiltration in inflammatory reactions, that alpha M beta2 integrin (Mac-1 or CD11b/CD18) mediates this adhesion, and that the alphaM beta2 integrin-mediated adhesion to myeloperoxidase is distinct from the previously reported ability of this integrin to bind to certain denatured proteins at high concentrations.

  • 35. Jung, Sung-Jun
    et al.
    Kim, Ji Eun Hani
    Reithinger, Johannes H.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Kim, Hyun
    The Sec62-Sec63 translocon facilitates translocation of the C-terminus of membrane proteins2014Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 127, nr 19, s. 4270-4278Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The Sec62-Sec63 complex mediates post-translational translocation of a subset of primarily secretory proteins into the endoplasmic reticulum (ER) in yeast. Therefore, it has been thought that membrane proteins, which are mainly co-translationally targeted into the ER, are not handled by the Sec62-Sec63 translocon. By systematic analysis of single and multi-spanning membrane proteins with broad sequence context [with differing hydrophobicity, flanking charged residues and orientation of transmembrane (TM) segments], we show that mutations in the N-terminal cytosolic domain of yeast Sec62 impair its interaction with Sec63 and lead to defects in membrane insertion and translocation of the C-terminus of membrane proteins. These results suggest that there is an unappreciated function of the Sec62-Sec63 translocon in regulating topogenesis of membrane proteins in the eukaryotic cell.

  • 36.
    Kandasamy, Ganapathi
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Andréasson, Claes
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Hsp70-Hsp110 chaperones deliver ubiquitin-dependent and -independent substrates to the 26S proteasome for proteolysis in yeast2018Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 131, nr 6, artikel-id jcs210948Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    During protein quality control, proteotoxic misfolded proteins are recognized by molecular chaperones, ubiquitylated by dedicated quality control ligases and delivered to the 26S proteasome for degradation. Proteins belonging to the Hsp70 chaperone and Hsp110 (the Hsp70 nucleotide exchange factor) families function in the degradation of misfolded proteins by the ubiquitin-proteasome system via poorly understood mechanisms. Here, we report that the Saccharomyces cerevisiae Hsp110 proteins (Sse1 and Sse2) function in the degradation of Hsp70-associated ubiquitin conjugates at the post-ubiquitylation step and are also required for ubiquitin-independent proteasomal degradation. Hsp110 associates with the 19S regulatory particle of the 26S proteasome and interacts with Hsp70 to f acilitate the delivery of Hsp70 substrates for proteasomal degradation. By using a highly defined ubiquitin-independent proteasome substrate, we show that the mere introduction of a single Hsp70-binding site renders its degradation dependent on Hsp110. The findings define a dedicated and chaperone-dependent pathway for the efficient shuttling of cellular proteins to the proteasome with profound implications for understanding protein quality control and cellular stress management.

  • 37.
    Kihlmark, Madeleine
    et al.
    Södertörns högskola, Avdelning Naturvetenskap.
    Imreh, Gabriela
    Södertörns högskola, Avdelning Naturvetenskap.
    Hallberg, Einar
    Södertörns högskola, Avdelning Naturvetenskap.
    Sequential degradation of proteins from the nuclear envelope during apoptosis2001Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 114, nr 20, s. 3643-3653Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    We have produced new antibodies specific for the integral pore membrane protein POM121. Using these antibodies we show that during apoptosis POM121 becomes proteolytically degraded in a caspase-dependent manner. The POM121 antibodies and antibodies specific for other proteins of the nuclear envelope were used in a comparative study of nuclear apoptosis in staurosporine-treated buffalo rat liver cells. Nuclei from these cells were classified in three different stages of apoptotic progression: stage I, moderately condensed chromatin surrounded by a smooth nuclear periphery; stage II, compact patches of condensed chromatin collapsing against a smooth nuclear periphery; stage III, round compact chromatin bodies surrounded by grape-shaped nuclear periphery. We have performed double labeling immunofluorescence microscopy of individual apoptotic cells and quantitative immunoblotting analysis of total proteins from apoptotic cell cultures. The results showed that degradation of nuclear envelope marker proteins occurred in a specific order. POM121 degradation occurred surprisingly early and was initiated before nucleosomal DNA degradation could be detected using TUNEL assay and completed before clustering of the nuclear pores. POM121 was eliminated significantly more rapid compared with NUP153 (a peripheral protein located in the nucleoplasmic basket of the nuclear pore complex) and lamin B (a component of the nuclear lamina). Disappearance of NUP153 and lamin B was coincident with onset of DNA fragmentation and clustering of nuclear pores. By contrast, the peripheral NPC protein p62 was degraded much later. The results suggest that degradation of POM121 may be an important early step in propagation of nuclear apoptosis.

  • 38. Klinger, Stine C
    et al.
    Glerup, Simon
    Raarup, Merete K
    Mari, Muriel C
    Nyegaard, Mette
    Koster, Gerbrand
    Prabakaran, Thaneas
    Nilsson, Stefan K
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Fysiologisk kemi.
    Kjaergaard, Maj M
    Bakke, Oddmund
    Nykjær, Anders
    Olivecrona, Gunilla
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Fysiologisk kemi.
    Petersen, Claus Munck
    Nielsen, Morten S
    SorLA regulates the activity of lipoprotein lipase by intracellular trafficking2011Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 124, s. 1095-1105Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Many different tissues and cell types exhibit regulated secretion of lipoprotein lipase (LPL). However, the sorting of LPL in the trans Golgi network has not, hitherto, been understood in detail. Here, we characterize the role of SorLA (officially known as SorLA-1 or sortilin-related receptor) in the intracellular trafficking of LPL. We found that LPL bound to SorLA under neutral and acidic conditions, and in cells this binding mainly occurred in vesicular structures. SorLA expression changed the subcellular distribution of LPL so it became more concentrated in endosomes. From the endosomes, LPL was further routed to the lysosomes, which resulted in a degradation of newly synthesized LPL. Consequently, an 80% reduction of LPL activity was observed in cells that expressed SorLA. By analogy, SorLA regulated the vesicle-like localization of LPL in primary neuronal cells. Thus, LPL binds to SorLA in the biosynthetic pathway and is subsequently transported to endosomes. As a result of this SorLA mediated-transport, newly synthesized LPL can be routed into specialized vesicles and eventually sent to degradation, and its activity thereby regulated.

  • 39. Kourmouli, Niki
    et al.
    Jeppesen, Peter
    Mahadevhaiah, Shantha
    Burgoyne, Paul
    Wu, Rong
    Gilbert, David M.
    Bongiorni, Silvia
    Prantera, Giorgio
    Fanti, Laura
    Pimpinelli, Sergio
    Shi, Wei
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för fysiologi och utvecklingsbiologi, Zoologisk utvecklingsbiologi.
    Fundele, Reinald
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för fysiologi och utvecklingsbiologi, Zoologisk utvecklingsbiologi.
    Singh, Prim B.
    Heterochromatin and tri-methylated lysine 20 of histone H4 in animals2004Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 117, nr 14, s. 2491-2501Artikel i tidskrift (Refereegranskat)
  • 40. Lambaerts, Kathleen
    et al.
    Van Dyck, Stijn
    Mortier, Eva
    Ivarsson, Ylva
    KU Leuven.
    Degeest, Gisèle
    Luyten, Annouck
    Vermeiren, Elke
    Peers, Bernard
    David, Guido
    Zimmermann, Pascale
    Syntenin, a syndecan adaptor and an Arf6 phosphatidylinositol 4,5-bisphosphate effector, is essential for epiboly and gastrulation cell movements in zebrafish2012Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 125, nr Pt 5, s. 1129-1140Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Epiboly, the spreading and the thinning of the blastoderm to cover the yolk cell and close the blastopore in fish embryos, is central to the process of gastrulation. Despite its fundamental importance, little is known about the molecular mechanisms that control this coordinated cell movement. By a combination of knockdown studies and rescue experiments in zebrafish (Danio rerio), we show that epiboly relies on the molecular networking of syntenin with syndecan heparan sulphate proteoglycans, which act as co-receptors for adhesion molecules and growth factors. Furthermore, we show that the interaction of syntenin with phosphatidylinositol 4,5-bisphosphate (PIP2) and with the small GTPase ADP-ribosylation factor 6 (Arf6), which regulate the endocytic recycling of syndecan, is necessary for epiboly progression. Analysis of the earliest cellular defects suggests a role for syntenin in the autonomous vegetal expansion of the yolk syncytial layer and the rearrangement of the actin cytoskeleton in extra-embryonic tissues, but not in embryonic cell fate determination. This study identifies the importance of the syntenin-syndecan-PIP2-Arf6 complex for the progression of fish epiboly and establishes its key role in directional cell movements during early development.

  • 41.
    Larsson, Veronica J.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Jafferali, Mohammed Hakim
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Vijayaraghavan, Balaje
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Figueroa, Ricardo A.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Hallberg, Einar
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Mitotic spindle assembly and γ-tubulin localisation depend on the integral nuclear membrane protein, Samp12018Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 131, nr 8, artikel-id jcs211664Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    We have investigated a possible role of the inner nuclear membrane protein, Samp1, in the mitotic machinery. Live cell imaging showed that Samp1aYFP distributed as filamentous structures in the mitotic spindle, partially co-localising with ß-tubulin. Samp1 depletion resulted in an increased frequency of cells with signs of chromosomal mis-segregation and prolonged metaphase, indicating problems with spindle assembly and/or chromosomal alignment. Consistently, mitotic spindles in Samp1 depleted cells contained significantly lower levels of ß-tubulin and γ-tubulin, phenotypes which were rescued by overexpression of Samp1aYFP. We found that Samp1 can bind directly to γ-tubulin and that Samp1 co-precipitated with γ-tubulin and HAUS6 of the Augmin complex in live cells. The levels of Haus6, in the mitotic spindle also decreased after Samp1 depletion. We show that Samp1 is involved in the recruitment of Haus6 and γ-tubulin to the mitotic spindle. Samp1 is the first inner nuclear membrane protein shown to have a function in mitotic spindle assembly.

  • 42.
    Lundmark, Richard
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Carlsson, Sven R
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    SNX9 - a prelude to vesicle release2009Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 122, nr 1, s. 5-11Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The sorting nexin SNX9 has, in the past few years, been singled out as an important protein that participates in fundamental cellular activities. SNX9 binds strongly to dynamin and is partly responsible for the recruitment of this GTPase to sites of endocytosis. SNX9 also has a high capacity for modulation of the membrane and might therefore participate in the formation of the narrow neck of endocytic vesicles before scission occurs. Once assembled on the membrane, SNX9 stimulates the GTPase activity of dynamin to facilitate the scission reaction. It has also become clear that SNX9 has the ability to activate the actin regulator N-WASP in a membrane-dependent manner to coordinate actin polymerization with vesicle release. In this Commentary, we summarize several aspects of SNX9 structure and function in the context of membrane remodeling, discuss its interplay with various interaction partners and present a model of how SNX9 might work in endocytosis.

  • 43.
    Maddika, Subbareddy
    et al.
    Manitoba Institute of Cell Biology, CancerCare Manitoba, University of Manitoba, Winnipeg, MB, R3E 0V9, Canada; Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, MB, Canada.
    Ande, Sudharsana Rao
    Manitoba Institute of Cell Biology, CancerCare Manitoba, University of Manitoba, Winnipeg, MB, R3E 0V9, Canada.
    Wiechec, Emilia
    Manitoba Institute of Cell Biology, CancerCare Manitoba; Department of Human Genetics, University of Aarhus, Aarhus, Denmark; Department of Experimental and Clinical Radiobiology, Oncology Center, Maria Sklodowka-Curie Memorial Institute, Wybrzeze Armii Krajowej 15, PL-44100 Gliwice, Poland .
    Hansen, Lise Lotte
    Department of Experimental and Clinical Radiobiology, Oncology Center, Maria Sklodowka-Curie Memorial Institute, Wybrzeze Armii Krajowej 15, PL-44100 Gliwice, Poland .
    Wesselborg, Sebastian
    Department of Internal Medicine I, University of Tübingen, Tübingen, Germany.
    Los, Marek Jan
    BioApplications Enterprises, Winnipeg, MB, Canada; Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, MB, R3E 0V9, Canada; Manitoba Institute of Cell Biology, CancerCare Manitoba, University of Manitoba, Winnipeg, MB, R3E 0V9, Canada .
    Akt-mediated phosphorylation of CDK2 regulates its dual role in cell cycle progression and apoptosis2008Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 121, s. 979-988Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Here, we show that CDK2, an S-phase cyclin-dependent kinase, is a novel target for Akt during cell cycle progression and apoptosis. Akt phosphorylates CDK2 at threonine 39 residue both in vitro and in vivo. Although CDK2 threonine 39 phosphorylation mediated by Akt enhances cyclin-A binding, it is dispensable for its basal binding and the kinase activity. In addition, for the first time, we report a transient nucleo-cytoplasmic shuttling of Akt during specific stages of the cell cycle, in particular during the late S and G2 phases. The Akt that is re-localized to the nucleus phosphorylates CDK2 and causes the temporary cytoplasmic localization of the CDK2–cyclin-A complex. The CDK2 cytoplasmic redistribution is required for cell progression from S to G2-M phase, because the CDK2 T39A mutant, which lacks the phosphorylation site and is defective in cytoplasmic localization, severely affects cell cycle progression at the transition from S to G2-M. Interestingly, we also show that the Akt/CDK2 pathway is constitutively activated by some anticancer drugs, such as methotrexate and docetaxel, and under these conditions it promotes, rather than represses, cell death. Thus, the constitutive activation of the Akt/CDK2 pathway and changed subcellular localization promotes apoptosis. By contrast, the transient, physiological Akt/CDK2 activation is necessary for cell cycle progression.

  • 44.
    Maddika, Subbareddy
    et al.
    University of Manitoba, Winnipeg, Canada .
    Booy, Evan P.
    University of Manitoba, Winnipeg, Canada .
    Johar, Dina
    University of Manitoba, Winnipeg, Canada .
    Gibson, Spencer B.
    University of Manitoba, Winnipeg, Canada .
    Ghavami, Saeid
    University of Manitoba, Winnipeg, Canada .
    Los, Marek Jan
    University of Manitoba, Winnipeg, Canada .
    Cancer-specific toxicity of apoptin is independent of death receptors but involves the loss of mitochondrial membrane potential and the release of mitochondrial cell-death mediators by a Nur77-dependent pathway2005Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 118, s. 4485-4493Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Apoptin, a small proline-rich protein derived from the chicken anaemia virus, induces cell death selectively in cancer cells. The signalling pathways of apoptin-induced, cancer cell-selective apoptosis are not well understood. Here, we demonstrate that apoptin triggers apoptosis by activating the mitochondrial/intrinsic pathway, and that it acts independently of the death receptor/extrinsic pathway. Jurkat cells deficient in either FADD or caspase-8 (which are both necessary for the extrinsic pathway) were equally as sensitive to apoptin as their parental clones. This demonstrates that apoptin is likely to act through the mitochondrial death pathway. Apoptin treatment causes a loss of mitochondrial membrane potential, and release of the mitochondrial proteins cytochrome c and apoptosis-inducing factor. Apoptin-induced cell death is counteracted by the anti-apoptotic Bcl-2 family members, Bcl-2 itself and Bcl-XL, as shown in Jurkat leukaemia cells. In addition, we describe the processing and activation of caspase-3. By contrast, cleavage of caspase-8, which is predominantly triggered by the death receptor pathway, is not observed. Furthermore, apoptin triggers the cytoplasmic translocation of Nur77, and the inhibition of Nur77 expression by siRNA significantly protects MCF7 cells from apoptin-triggered cell death. Thus, our data indicate that the apoptin death signal(s) ultimately converges at the mitochondria, and that it acts independently of the death receptor pathway.                               

  • 45. Magee, Anthony I
    et al.
    Adler, Jeremy
    Parmryd, Ingela
    Cold-induced coalescence of T-cell plasma membrane microdomains activates signalling pathways2005Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 118, nr Pt 14, s. 3141-51Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The plasma membranes of eukaryotic cells are hypothesised to contain microdomains with distinct lipid and protein composition known as lipid rafts. In T cells, cross-linking of lipid raft components triggers signalling cascades. We show that the T-cell antigen receptor (TCR) and a protein tyrosine kinase, Lck, have a patchy plasma membrane distribution in Jurkat T cells at reduced temperatures, although they have a continuous distribution at physiological temperature (37 degrees C). GM1 displays a patchy distribution at reduced temperature after Triton X-100 extraction. The archetypal non-lipid raft marker, the transferrin receptor, displays a more continuous plasma membrane distribution uncorrelated with that of Lck at 0 degrees C. Cold-induced aggregation of the lipid raft-partitioning proteins is accompanied by increased tyrosine phosphorylation and ERK activation, peaking at 10-20 degrees C. Tyrosine phosphorylation is further greatly increased by ligating the TCR with anti-CD3 at 10-20 degrees C. The tyrosine phosphorylation mainly occurred at the plasma membrane, was dependent on Lck and on the surface expression of the TCR. The activation of tyrosine phosphorylation and ERK by TCR ligation at reduced temperature also occurred in human primary T cells. These results support the concept that lipid rafts can form in membranes of live cells and that their coalescence stimulates signalling.

  • 46.
    Magnusson, Peetra
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Klinisk immunologi.
    Rolny, Charlotte
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Cancer och vaskulärbiologi.
    Jakobsson, Lars
    Wikner, Charlotte
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Cancer och vaskulärbiologi.
    Wu, Y
    Hicklin, DJ
    Claesson-Welsh, Lena
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Cancer och vaskulärbiologi.
    Deregulation of Flk-1/vascular endothelial growth factor receptor-2 in fibroblast growth factor receptor-1-deficient vascular stem cell development2004Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 117, nr Pt 8, s. 1513-1523Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    We have employed embryoid bodies derived from murine embryonal stem cells to study effects on vascular development induced by fibroblast growth factor (FGF)-2 and FGF receptor-1, in comparison to the established angiogenic factor vascular endothelial growth factor (VEGF)-A and its receptor VEGF receptor-2. Exogenous FGF-2 promoted formation of morphologically distinct, long slender vessels in the embryoid bodies, whereas VEGF-A-treated bodies displayed a compact plexus of capillaries. FGF-2 stimulation of embryonal stem cells under conditions where VEGF-A/VEGFR-2 function was blocked, led to formation of endothelial cell clusters, which failed to develop into vessels. FGFR-1(-/-) embryoid bodies responded to VEGF-A by establishment of the characteristic vascular plexus, but FGF-2 had no effect on vascular development in the absence of FGFR-1. The FGFR-1(-/-) embryoid bodies displayed considerably increased basal level of vessel formation, detected by immunohistochemical staining for platelet-endothelial cell adhesion molecule (PECAM)/CD31. This basal vascularization was blocked by neutralizing antibodies against VEGFR-2 or VEGF-A and biochemical analyses indicated changes in regulation of VEGFR-2 in the absence of FGFR-1 expression. We conclude that VEGF-A/VEGFR-2-dependent vessel formation occurs in the absence of FGF-2/FGFR-1, which, however, serve to modulate vascular development.

  • 47.
    Majeed, Meytham
    et al.
    Linköpings universitet, Hälsouniversitetet. Linköpings universitet, Institutionen för molekylär och klinisk medicin, Medicinsk mikrobiologi.
    Krause, K-H
    Clark, RA
    Kihlström, Erik
    Linköpings universitet, Hälsouniversitetet. Linköpings universitet, Institutionen för hälsa och miljö. Östergötlands Läns Landsting, Laboratoriemedicinskt centrum, Klinisk mikrobiologi.
    Stendahl, Olle
    Linköpings universitet, Hälsouniversitetet. Linköpings universitet, Institutionen för molekylär och klinisk medicin, Medicinsk mikrobiologi.
    Localization of intracellular Ca2+ stores in HeLa cells during infection with Chlamydia trachomatis. 1999Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 112, s. 35-44Artikel i tidskrift (Refereegranskat)
  • 48.
    Marin-Vicente, Consuelo
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Domingo-Prim, Judit
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Eberle, Andrea B.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Visa, Neus
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    RRP6/EXOSC10 is required for the repair of DNA double-strand breaks by homologous recombination2015Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 128, nr 6, s. 1097-1107Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The exosome acts on different RNA substrates and plays important roles in RNA metabolism. The fact that short non-coding RNAs are involved in the DNA damage response led us to investigate whether the exosome factor RRP6 of Drosophila melanogaster and its human ortholog EXOSC10 play a role in DNA repair. Here, we show that RRP6 and EXOSC10 are recruited to DNA double-strand breaks (DSBs) in S2 cells and HeLa cells, respectively. Depletion of RRP6/ EXOSC10 does not interfere with the phosphorylation of the histone variant H2Av (Drosophila) or H2AX (humans), but impairs the recruitment of the homologous recombination factor RAD51 to the damaged sites, without affecting RAD51 levels. The recruitment of RAD51 to DSBs in S2 cells is also inhibited by overexpression of RRP6-Y361A-V5, a catalytically inactive RRP6 mutant. Furthermore, cells depleted of RRP6 or EXOSC10 are more sensitive to radiation, which is consistent with RRP6/EXOSC10 playing a role in DNA repair. RRP6/EXOSC10 can be co-immunoprecipitated with RAD51, which links RRP6/EXOSC10 to the homologous recombination pathway. Taken together, our results suggest that the ribonucleolytic activity of RRP6/EXOSC10 is required for the recruitment of RAD51 to DSBs.

  • 49.
    Mohan, Jagan
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Morén, Björn
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Larsson, Elin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Holst, Mikkel
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Lundmark, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Cavin3 interacts with cavin1 and caveolin1 to increase surface dynamics of caveolae2015Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 128, nr 5, s. 979-991Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Caveolae are invaginations of the cell surface thought to regulate membrane tension, signalling, adhesion and lipid homeostasis due to their dynamic behaviour ranging from stable surface association to dynamic rounds of fission and fusion with the plasma membrane. The caveolae coat is generated by oligomerisation of the membrane protein caveolin and the family of cavin proteins. Here, we show that cavin3 is targeted to caveolae by cavin1 where it interacts with the scaffolding domain of caveolin1 and promote caveolae dynamics. We found that the N-terminal region of cavin3 binds a trimer of the cavin1 N-terminus in competition with a homologous cavin2 region, showing that the cavins form distinct subcomplexes via their N-terminal regions. Our data shows that cavin3 is enriched at deeply invaginated caveolae and that loss of cavin3 in cells results in an increase of stable caveolae and a decrease of caveolae with short duration time at the membrane. We propose that cavin3 is recruited to the caveolae coat by cavin1 to interact with caveolin1 and regulate the duration time of caveolae at the plasma membrane.

  • 50.
    Moustakas, Aristidis
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwiginstitutet för cancerforskning.
    Smad signalling network2002Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 115, nr Pt 17, s. 3355-3356Artikel, forskningsöversikt (Övrigt vetenskapligt)
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