Background: Divergence in gene regulation has emerged as a key mechanism underlying species differentiation. Comparative analysis of co-expression networks across species can reveal conservation and divergence in the regulation of genes. Results: We inferred co-expression networks of A. thaliana, Populus spp. and O. sativa using state-of-the-art methods based on mutual information and context likelihood of relatedness, and conducted a comprehensive comparison of these networks across a range of co-expression thresholds. In addition to quantifying gene-gene link and network neighbourhood conservation, we also applied recent advancements in network analysis to do cross-species comparisons of network properties such as scale free characteristics and gene centrality as well as network motifs. We found that in all species the networks emerged as scale free only above a certain co-expression threshold, and that the high-centrality genes upholding this organization tended to be conserved. Network motifs, in particular the feed-forward loop, were found to be significantly enriched in specific functional subnetworks but where much less conserved across species than gene centrality. Although individual gene-gene co-expression had massively diverged, up to similar to 80% of the genes still had a significantly conserved network neighbourhood. For genes with multiple predicted orthologs, about half had one ortholog with conserved regulation and another ortholog with diverged or non-conserved regulation. Furthermore, the most sequence similar ortholog was not the one with the most conserved gene regulation in over half of the cases. Conclusions: We have provided a comprehensive analysis of gene regulation evolution in plants and built a web tool for Comparative analysis of Plant co-Expression networks (ComPlEx, http:// complex. plantgenie. org/). The tool can be particularly useful for identifying the ortholog with the most conserved regulation among several sequence-similar alternatives and can thus be of practical importance in e. g. finding candidate genes for perturbation experiments.

Accessing and exploring large-scale genomics data sets remains a significant challenge to researchers without specialist bioinformatics training. We present the integrated PlantGenIE.org platform for exploration of Populus, conifer and Arabidopsis genomics data, which includes expression networks and associated visualization tools. Standard features of a model organism database are provided, including genome browsers, gene list annotation, BLAST homology searches and gene information pages. Community annotation updating is supported via integration of WebApollo. We have produced an RNA-sequencing (RNA-Seq) expression atlas for Populus tremula and have integrated these data within the expression tools. An updated version of the COMPLEX resource for performing comparative plant expression analyses of gene coexpression network conservation between species has also been integrated. The PlantGenIE.org platform provides intuitive access to large-scale and genome-wide genomics data from model forest tree species, facilitating both community contributions to annotation improvement and tools supporting use of the included data resources to inform biological insight.

The secondary xylem of conifers is composed mainly of tracheids that differ anatomically and chemically from angiosperm xylem cells. There is currently no high-spatial-resolution data available profiling gene expression during wood formation for any coniferous species, which limits insight into tracheid development.

RNA-sequencing data from replicated, high-spatial-resolution section series throughout the cambial and woody tissues of Picea abies were used to generate the NorWood.conGenIE.org web resource, which facilitates exploration of the associated gene expression profiles and co-expression networks.

Integration within PlantGenIE.org enabled a comparative regulomics analysis, revealing divergent co-expression networks between P. abies and the two angiosperm species Arabidopsis thaliana and Populus tremula for the secondary cell wall (SCW) master regulator NAC Class IIB transcription factors. The SCW cellulose synthase genes (CesAs) were located in the neighbourhoods of the NAC factors in A. thaliana and P. tremula, but not in P. abies. The NorWood co-expression network enabled identification of potential SCW CesA regulators in P. abies.

The NorWood web resource represents a powerful community tool for generating evo-devo insights into the divergence of wood formation between angiosperms and gymnosperms and for advancing understanding of the regulation of wood development in P. abies.

Trees represent the largest terrestrial carbon sink and a renewable source of ligno-cellulose. There is significant scope for yield and quality improvement in these largely undomesticated species, however, efforts to engineer new, elite varieties are constrained by the lack of a comprehensive understanding of the transcriptional network underlying cambial growth and wood formation. We generated RNA Sequencing transcriptome data for four mature, wild-growing aspens (Populus tremula) from high-spatial-resolution tangential cryosection series spanning the secondary phloem, vascular cambium, expanding and secondary cell wall forming xylem cells, cell death zone and the previous years annual ring. The transcriptome comprised 28,294 expressed, previously annotated protein-coding genes, 78 novel protein-coding genes and 567 long intergenic non-coding RNAs. Most paralogs originating from the Salicaceae whole genome duplication had diverged expression, with the notable exception of those with high expression during secondary cell wall deposition. We performed co-expression network analysis to identify central transcriptional modules and associated several of these with known biological processes. This revealed previously uncharacterized complexity underlying the regulation of cambial growth and wood formation, with modules forming a continuum of activated processes across the tissues. The high spatial resolution suggested novel roles for known genes involved in xylan and cellulose biosynthesis, regulators of xylem vessel and fiber differentiation and components of lignification. The associated web resource (AspWood, http://aspwood.popgenie.org) integrates the data within a set of interactive tools for exploring the co-expression network of cambial growth and wood formation.