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Amino acids evolve at different speeds within protein sequences, because their functional and structural roles are different. Notably, amino acids located at the surface of proteins are known to evolve more rapidly than those in the core. In particular, amino acids at the N- and C-termini of protein sequences are likely to be more exposed than those at the core of the folded protein due to their location in the peptidic chain, and they are known to be less structured. Because of these reasons, we would expect that amino acids located at protein termini would evolve faster than residues located inside the chain. Here we test this hypothesis and found that amino acids evolve almost twice as fast at protein termini compared with those in the center, hinting at a strong topological bias along the sequence length. We further show that the distribution of solvent-accessible residues and functional domains in proteins readily explain how structural and functional constraints are weaker at their termini, leading to the observed excess of amino acid substitutions. Finally, we show that the specific evolutionary rates at protein termini may have direct consequences, notably misleading in silico methods used to infer sites under positive selection within genes. These results suggest that accounting for positional information should improve evolutionary models., Competing Interests: Conflict of interest statement. No conflict of interest from any of the authors., (© The Author(s) 2023. Published by Oxford University Press on behalf of Society for Molecular Biology and Evolution.)

Ancestral sequence reconstruction is a fundamental aspect of molecular evolution studies and can trace small-scale sequence modifications through the evolution of genomes and species. In contrast, fine-grained reconstructions of ancestral genome organizations are still in their infancy, limiting our ability to draw comprehensive views of genome and karyotype evolution. Here we reconstruct the detailed gene contents and organizations of 624 ancestral vertebrate, plant, fungi, metazoan and protist genomes, 183 of which are near-complete chromosomal gene order reconstructions. Reconstructed ancestral genomes are similar to their descendants in terms of gene content as expected and agree precisely with reference cytogenetic and in silico reconstructions when available. By comparing successive ancestral genomes along the phylogenetic tree, we estimate the intra- and interchromosomal rearrangement history of all major vertebrate clades at high resolution. This freely available resource introduces the possibility to follow evolutionary processes at genomic scales in chronological order, across multiple clades and without relying on a single extant species as reference., (© 2023. The Author(s).)

Accurate species phylogenies are a prerequisite for all evolutionary research. Teleosts are the largest and most diversified group of extant vertebrates, but relationships among their three oldest extant lineages remain unresolved. On the basis of seven high-quality new genome assemblies in Elopomorpha (tarpons, eels), we revisited the topology of the deepest branches of the teleost phylogeny using independent gene sequence and chromosomal rearrangement phylogenomic approaches. These analyses converged to a single scenario that unambiguously places the Elopomorpha and Osteoglossomorpha (arapaima, elephantnose fish) in a monophyletic sister group to all other teleosts, i.e., the Clupeocephala lineage (zebrafish, medaka). This finding resolves more than 50 years of controversy on the evolutionary relationships of these lineages and highlights the power of combining different levels of genome-wide information to solve complex phylogenies.

Phylogenetic gene trees recapitulate the evolutionary history of genes across species, forming an essential framework for comparative genomic studies. In particular, within the context of whole-genome duplications (WGDs), they serve as a basis to investigate patterns of duplicate gene retention and loss, timing of genome rediploidization, and, more generally, to explore the functional consequences of the duplication in descending species. Yet, despite ever more sophisticated models to describe the evolution of gene sequences, building accurate gene trees remains a challenge in ancient polyploid taxons. WGDs generate complex gene families with many duplicated copies and recurrent gene losses, which complicate this task even more. Here, we describe how to use SCORPiOs, a novel method that leverages synteny conservation to provide more accurate phylogenies in the presence of a known WGD event., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Motivation: FASTA files are the de facto standard for sharing, manipulating and storing biological sequences, while concatenated in multiFASTA they tend to be unwieldy for two main reasons: (i) they can become big enough that their manipulation with standard text-editing tools is unpractical, either due to slowness or memory consumption; (ii) by mixing metadata (headers) and data (sequences), bulk operations using standard text streaming tools (such as sed or awk) are impossible without including a parsing step, which may be error-prone and introduce friction in the development process., Results: Here, we present FUSTA (FUse for faSTA), a software utility which makes use of the FUSE technology to expose a multiFASTA file as a hierarchy of virtual files, letting users operate directly on the sequences as independent virtual files through classical file manipulation methods., Availability and Implementation: FUSTA is freely available under the CeCILL-C (LGPLv3-compatible) license at https://github.com/delehef/fusta., Supplementary Information: Supplementary data are available at Bioinformatics Advances online., (© The Author(s) 2022. Published by Oxford University Press.)

Whole genome sequencing is increasingly used to diagnose medical conditions of genetic origin. While both coding and non-coding DNA variants contribute to a wide range of diseases, most patients who receive a WGS-based diagnosis today harbour a protein-coding mutation. Functional interpretation and prioritization of non-coding variants represents a persistent challenge, and disease-causing non-coding variants remain largely unidentified. Depending on the disease, WGS fails to identify a candidate variant in 20-80% of patients, severely limiting the usefulness of sequencing for personalised medicine. Here we present FINSURF, a machine-learning approach to predict the functional impact of non-coding variants in regulatory regions. FINSURF outperforms state-of-the-art methods, owing in particular to optimized control variants selection during training. In addition to ranking candidate variants, FINSURF breaks down the score for each variant into contributions from individual annotations, facilitating the evaluation of their functional relevance. We applied FINSURF to a diverse set of 30 diseases with described causative non-coding mutations, and correctly identified the disease-causative non-coding variant within the ten top hits in 22 cases. FINSURF is implemented as an online server to as well as custom browser tracks, and provides a quick and efficient solution to prioritize candidate non-coding variants in realistic clinical settings., Competing Interests: The authors have declared that no competing interests exist.

Genomicus is a database and web-server dedicated to comparative genomics in eukaryotes. Its main functionality is to graphically represent the conservation of genomic blocks between multiple genomes, locally around a specific gene of interest or genome-wide through karyotype comparisons. Since 2010 and its first release, Genomicus has synchronized with 60 Ensembl releases and seen the addition of functions that have expanded the type of analyses that users can perform. Today, five public instances of Genomicus are supporting a total number of 1029 extant genomes and 621 ancestral reconstructions from all eukaryotes kingdoms available in Ensembl and Ensembl Genomes databases complemented with four additional instances specific to taxonomic groups of interest. New visualization and query tools are described in this manuscript. Genomicus is freely available at http://www.genomicus.bio.ens.psl.eu/genomicus., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The bowfin (Amia calva) is a ray-finned fish that possesses a unique suite of ancestral and derived phenotypes, which are key to understanding vertebrate evolution. The phylogenetic position of bowfin as a representative of neopterygian fishes, its archetypical body plan and its unduplicated and slowly evolving genome make bowfin a central species for the genomic exploration of ray-finned fishes. Here we present a chromosome-level genome assembly for bowfin that enables gene-order analyses, settling long-debated neopterygian phylogenetic relationships. We examine chromatin accessibility and gene expression through bowfin development to investigate the evolution of immune, scale, respiratory and fin skeletal systems and identify hundreds of gene-regulatory loci conserved across vertebrates. These resources connect developmental evolution among bony fishes, further highlighting the bowfin's importance for illuminating vertebrate biology and diversity in the genomic era., (© 2021. The Author(s).)

Identifying causative variants in cis-regulatory elements (CRE) in neurodevelopmental disorders has proven challenging. We have used in vivo functional analyses to categorize rigorously filtered CRE variants in a clinical cohort that is plausibly enriched for causative CRE mutations: 48 unrelated males with a family history consistent with X-linked intellectual disability (XLID) in whom no detectable cause could be identified in the coding regions of the X chromosome (chrX). Targeted sequencing of all chrX CRE identified six rare variants in five affected individuals that altered conserved bases in CRE targeting known XLID genes and segregated appropriately in families. Two of these variants, FMR1CRE and TENM1CRE, showed consistent site- and stage-specific differences of enhancer function in the developing zebrafish brain using dual-color fluorescent reporter assay. Mouse models were created for both variants. In male mice Fmr1CRE induced alterations in neurodevelopmental Fmr1 expression, olfactory behavior and neurophysiological indicators of FMRP function. The absence of another likely causative variant on whole genome sequencing further supported FMR1CRE as the likely basis of the XLID in this family. Tenm1CRE mice showed no phenotypic anomalies. Following the release of gnomAD 2.1, reanalysis showed that TENM1CRE exceeded the maximum plausible population frequency of a XLID causative allele. Assigning causative status to any ultra-rare CRE variant remains problematic and requires disease-relevant in vivo functional data from multiple sources. The sequential and bespoke nature of such analyses renders them time-consuming and challenging to scale for routine clinical use., Competing Interests: No authors have competing interests

Regulation of gene expression relies on the activity of specialized genomic elements, enhancers or silencers, distributed over sometimes large distance from their target gene promoters. A significant part of vertebrate genomes consists in such regulatory elements, but their identification and that of their target genes remains challenging, due to the lack of clear signature at the nucleotide level. For many years the main hallmark used for identifying functional elements has been their sequence conservation between genomes of distant species, indicative of purifying selection. More recently, genome-wide biochemical assays have opened new avenues for detecting regulatory regions, shifting attention away from evolutionary constraints. Here, we review the respective contributions of comparative genomics and biochemical assays for the definition of regulatory elements and their targets and advocate that both sequence conservation and preserved synteny, taken as signature of functional constraint, remain essential tools in this task., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Whole-genome duplications (WGDs) have major impacts on the evolution of species, as they produce new gene copies contributing substantially to adaptation, isolation, phenotypic robustness, and evolvability. They result in large, complex gene families with recurrent gene losses in descendant species that sequence-based phylogenetic methods fail to reconstruct accurately. As a result, orthologs and paralogs are difficult to identify reliably in WGD-descended species, which hinders the exploration of functional consequences of WGDs. Here, we present Synteny-guided CORrection of Paralogies and Orthologies (SCORPiOs), a novel method to reconstruct gene phylogenies in the context of a known WGD event. WGDs generate large duplicated syntenic regions, which SCORPiOs systematically leverages as a complement to sequence evolution to infer the evolutionary history of genes. We applied SCORPiOs to the 320-My-old WGD at the origin of teleost fish. We find that almost one in four teleost gene phylogenies in the Ensembl database (3,394) are inconsistent with their syntenic contexts. For 70% of these gene families (2,387), we were able to propose an improved phylogenetic tree consistent with both the molecular substitution distances and the local syntenic information. We show that these synteny-guided phylogenies are more congruent with the species tree, with sequence evolution and with expected expression conservation patterns than those produced by state-of-the-art methods. Finally, we show that synteny-guided gene trees emphasize contributions of WGD paralogs to evolutionary innovations in the teleost clade., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

PIWI-interacting RNAs (piRNAs) target transcripts by sequence complementarity serving as guides for RNA slicing in animal germ cells. The piRNA pathway is increasingly recognized as critical for essential cellular functions such as germline development and reproduction. In the Anopheles gambiae ovary, as much as 11% of piRNAs map to protein-coding genes. Here, we show that ovarian mRNAs and long non-coding RNAs (lncRNAs) are processed into piRNAs that can direct other transcripts into the piRNA biogenesis pathway. Targeting piRNAs fuel transcripts either into the ping-pong cycle of piRNA amplification or into the machinery of phased piRNA biogenesis, thereby creating networks of inter-regulating transcripts. RNAs of the same network share related genomic repeats. These repeats give rise to piRNAs, which target other transcripts and lead to a cascade of concerted RNA slicing. While ping-pong networks are based on repeats of several hundred nucleotides, networks that rely on phased piRNA biogenesis operate through short ~40-nucleotides long repeats, which we named snetDNAs. Interestingly, snetDNAs are recurring in evolution from insects to mammals. Our study brings to light a new type of conserved regulatory pathway, the snetDNA-pathway, by which short sequences can include independent genes and lncRNAs in the same biological pathway.

The spatiotemporal expression of genes is controlled by enhancer sequences that bind transcription factors. Identifying the target genes of enhancers remains difficult because enhancers regulate gene expression over long genomic distances. To address this, we used an evolutionary approach to build two genome-wide maps of predicted enhancer-gene associations in the human and zebrafish genomes. Evolutionary conserved sequences were linked to their predicted target genes using PEGASUS, a bioinformatics method that relies on evolutionary conservation of synteny. The analysis of these maps revealed that the number of predicted enhancers linked to a gene correlate with its expression breadth. Comparison of both maps identified hundreds of putative vertebrate ancestral regulatory relationships from which we could determine that predicted enhancer-gene distances scale with genome size despite strong positional conservation. The two maps represent a resource for further studies, including the prioritization of sequence variants in whole genome sequence of patients affected by genetic diseases., (© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.)

ANISEED (https://www.aniseed.cnrs.fr) is the main model organism database for the worldwide community of scientists working on tunicates, the vertebrate sister-group. Information provided for each species includes functionally-annotated gene and transcript models with orthology relationships within tunicates, and with echinoderms, cephalochordates and vertebrates. Beyond genes the system describes other genetic elements, including repeated elements and cis-regulatory modules. Gene expression profiles for several thousand genes are formalized in both wild-type and experimentally-manipulated conditions, using formal anatomical ontologies. These data can be explored through three complementary types of browsers, each offering a different view-point. A developmental browser summarizes the information in a gene- or territory-centric manner. Advanced genomic browsers integrate the genetic features surrounding genes or gene sets within a species. A Genomicus synteny browser explores the conservation of local gene order across deuterostome. This new release covers an extended taxonomic range of 14 species, including for the first time a non-ascidian species, the appendicularian Oikopleura dioica. Functional annotations, provided for each species, were enhanced through a combination of manual curation of gene models and the development of an improved orthology detection pipeline. Finally, gene expression profiles and anatomical territories can be explored in 4D online through the newly developed Morphonet morphogenetic browser., (© The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.)

mRNA translation and decay appear often intimately linked although the rules of this interplay are poorly understood. In this study, we combined our recent P-body transcriptome with transcriptomes obtained following silencing of broadly acting mRNA decay and repression factors, and with available CLIP and related data. This revealed the central role of GC content in mRNA fate, in terms of P-body localization, mRNA translation and mRNA stability: P-bodies contain mostly AU-rich mRNAs, which have a particular codon usage associated with a low protein yield; AU-rich and GC-rich transcripts tend to follow distinct decay pathways; and the targets of sequence-specific RBPs and miRNAs are also biased in terms of GC content. Altogether, these results suggest an integrated view of post-transcriptional control in human cells where most translation regulation is dedicated to inefficiently translated AU-rich mRNAs, whereas control at the level of 5' decay applies to optimally translated GC-rich mRNAs., Competing Interests: MC, YC, CB, DF, OV, ZY, MB, MB, MK, CV, ME, CA, AM, PB, AH, HR, NS, DW No competing interests declared, (© 2019, Courel et al.)

Background: It has been proposed that more than 450 million years ago, two successive whole genome duplications took place in a marine chordate lineage before leading to the common ancestor of vertebrates. A precise reconstruction of these founding events would provide a framework to better understand the impact of these early whole genome duplications on extant vertebrates., Results: We reconstruct the evolution of chromosomes at the beginning of vertebrate evolution. We first compare 61 extant animal genomes to reconstruct the highly contiguous order of genes in a 326-million-year-old ancestral Amniota genome. In this genome, we establish a well-supported list of duplicated genes originating from the two whole genome duplications to identify tetrads of duplicated chromosomes. From this, we reconstruct a chronology in which a pre-vertebrate genome composed of 17 chromosomes duplicated to 34 chromosomes and was subject to seven chromosome fusions before duplicating again into 54 chromosomes. After the separation of the lineage of Gnathostomata (jawed vertebrates) from Cyclostomata (extant jawless fish), four more fusions took place to form the ancestral Euteleostomi (bony vertebrates) genome of 50 chromosomes., Conclusions: These results firmly establish the occurrence of two whole genome duplications in the lineage that precedes the ancestor of vertebrates, resolving in particular the ambiguity raised by the analysis of the lamprey genome. This work provides a foundation for studying the evolution of vertebrate chromosomes from the standpoint of a common ancestor and particularly the pattern of duplicate gene retention and loss that resulted in the gene composition of extant vertebrate genomes.

Since 2010, the Genomicus web server is available online at http://genomicus.biologie.ens.fr/genomicus. This graphical browser provides access to comparative genomic analyses in four different phyla (Vertebrate, Plants, Fungi, and non vertebrate Metazoans). Users can analyse genomic information from extant species, as well as ancestral gene content and gene order for vertebrates and flowering plants, in an integrated evolutionary context. New analyses and visualization tools have recently been implemented in Genomicus Vertebrate. Karyotype structures from several genomes can now be compared along an evolutionary pathway (Multi-KaryotypeView), and synteny blocks can be computed and visualized between any two genomes (PhylDiagView)., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: Polycomb Repressive Complexes 2 (PRC2) are multi-protein chromatin modifiers that are evolutionarily conserved among eukaryotes and play key roles in the regulation of gene expression, notably through the trimethylation of lysine 27 of histone H3 (H3K27me3). Although PRC2-mediated gene regulation has been studied in many organisms, few studies have explored in depth the evolutionary conservation of PRC2 targets., Results: Here, we compare the H3K27me3 epigenomic profiles for the two closely related species Arabidopsis thaliana and Arabidopsis lyrata and the more distant species Arabis alpina, three Brassicaceae that diverged from each other within the past 24 million years. Using a robust set of gene orthologs present in the three species, we identify two classes of evolutionarily conserved PRC2 targets, which are characterized by either developmentally plastic or developmentally constrained H3K27me3 marking across species. Constrained H3K27me3 marking is associated with higher conservation of promoter sequence information content and higher nucleosome occupancy compared to plastic H3K27me3 marking. Moreover, gene orthologs with constrained H3K27me3 marking exhibit a higher degree of tissue specificity and tend to be involved in developmental functions, whereas gene orthologs with plastic H3K27me3 marking preferentially encode proteins associated with metabolism and stress responses. In addition, gene orthologs with constrained H3K27me3 marking are the predominant contributors to higher-order chromosome organization., Conclusions: Our findings indicate that developmentally plastic and constrained H3K27me3 marking define two evolutionarily conserved modes of PRC2-mediated gene regulation that are associated with distinct selective pressures operating at multiple scales, from DNA sequence to gene function and chromosome architecture.

A conserved segment, i.e. a segment of chromosome unbroken during evolution, is an important operational concept in comparative genomics. Until now, algorithms that are designed to identify conserved segments often return synteny blocks that overlap, synteny blocks that include micro-rearrangements or synteny blocks erroneously short. Here we present definitions of conserved segments and synteny blocks independent of any heuristic method and we describe four new post-processing strategies to refine synteny blocks into accurate conserved segments. The first strategy identifies micro-rearrangements, the second strategy identifies mono-genic conserved segments, the third returns non-overlapping segments and the fourth repairs incorrect ruptures of synteny. All these refinements are implemented in a new version of PhylDiag that has been benchmarked against i-ADHoRe 3.0 and Cyntenator, based on a realistic simulated evolution and true simulated conserved segments.

Human doublecortin (DCX) mutations are associated with severe brain malformations leading to aberrant neuron positioning (heterotopia), intellectual disability and epilepsy. The Dcx protein plays a key role in neuronal migration, and hippocampal pyramidal neurons in Dcx knockout (KO) mice are disorganized. The single CA3 pyramidal cell layer observed in wild type (WT) is present as two abnormal layers in the KO, and CA3 KO pyramidal neurons are more excitable than WT. Dcx KO mice also exhibit spontaneous epileptic activity originating in the hippocampus. It is unknown, however, how hyperexcitability arises and why two CA3 layers are observed.Transcriptome analyses were performed to search for perturbed postnatal gene expression, comparing Dcx KO CA3 pyramidal cell layers with WT. Gene expression changes common to both KO layers indicated mitochondria and Golgi apparatus anomalies, as well as increased cell stress. Intriguingly, gene expression analyses also suggested that the KO layers differ significantly from each other, particularly in terms of maturity. Layer-specific molecular markers and BrdU birthdating to mark the final positions of neurons born at distinct timepoints revealed inverted layering of the CA3 region in Dcx KO animals. Notably, many early-born 'outer boundary' neurons are located in an inner position in the Dcx KO CA3, superficial to other pyramidal neurons. This abnormal positioning likely affects cell morphology and connectivity, influencing network function. Dissecting this Dcx KO phenotype sheds light on coordinated developmental mechanisms of neuronal subpopulations, as well as gene expression patterns contributing to a bi-layered malformation associated with epilepsy., (© The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Fibroblasts are important players in regulating tissue homeostasis. In the dermis, they are involved in wound healing where they differentiate into contractile myofibroblasts leading to wound closure. In nonhealing chronic wounds, fibroblasts fail to undertake differentiation. We established and used a human ex vivo model of chronic wounds where fibroblasts can undergo normal myofibroblast differentiation, or take on a nondifferentiable pathological state. At the whole genome scale, we identified the genes that are differentially regulated in these two cell fates. By coupling the search of evolutionary conserved regulatory elements with global gene network expression changes, we identified transcription factors (TF) potentially involved in myofibroblast differentiation, and constructed a network of relationship between these key factors. Among these, we found that TCF4, SOX9, EGR2, and FOXS1 are major regulators of fibroblast to myofibroblast differentiation. Conversely, down-regulation of MEOX2, SIX2, and MAF causes reprogramming of fibroblasts to myofibroblasts even in absence of TGF-β, the natural inducer of myofibroblast differentiation. These results provide insight into the fibroblast differentiation program and reveal a TF network essential for cellular reprogramming. They could lead to the development of new therapeutics to treat fibroblast-related human pathologies., (© 2015 by the Wound Healing Society.)

Background: Brassicaceae is a family of green plants of high scientific and economic interest, including thale cress (Arabidopsis thaliana), cruciferous vegetables (cabbages) and rapeseed., Results: We reconstruct an evolutionary framework of Brassicaceae composed of high-resolution ancestral karyotypes using the genomes of modern A. thaliana, Arabidopsis lyrata, Capsella rubella, Brassica rapa and Thellungiella parvula. The ancestral Brassicaceae karyotype (Brassicaceae lineages I and II) is composed of eight protochromosomes and 20,037 ordered and oriented protogenes. After speciation, it evolved into the ancestral Camelineae karyotype (eight protochromosomes and 22,085 ordered protogenes) and the proto-Calepineae karyotype (seven protochromosomes and 21,035 ordered protogenes) genomes., Conclusions: The three inferred ancestral karyotype genomes are shown here to be powerful tools to unravel the reticulated evolutionary history of extant Brassicaceae genomes regarding the fate of ancestral genes and genomic compartments, particularly centromeres and evolutionary breakpoints. This new resource should accelerate research in comparative genomics and translational research by facilitating the transfer of genomic information from model systems to species of agronomic interest.

Enhancers can regulate the transcription of genes over long genomic distances. This is thought to lead to selection against genomic rearrangements within such regions that may disrupt this functional linkage. Here we test this concept experimentally using the human X chromosome. We describe a scoring method to identify evolutionary maintenance of linkage between conserved noncoding elements and neighbouring genes. Chromatin marks associated with enhancer function are strongly correlated with this linkage score. We test >1,000 putative enhancers by transgenesis assays in zebrafish to ascertain the identity of the target gene. The majority of active enhancers drive a transgenic expression in a pattern consistent with the known expression of a linked gene. These results show that evolutionary maintenance of linkage is a reliable predictor of an enhancer's function, and provide new information to discover the genetic basis of diseases caused by the mis-regulation of gene expression.

Genomic rearrangements are a major source of evolutionary divergence in eukaryotic genomes, a cause of genetic diseases and a hallmark of tumor cell progression, yet the mechanisms underlying their occurrence and evolutionary fixation are poorly understood. Statistical associations between breakpoints and specific genomic features suggest that genomes may contain elusive “fragile regions” with a higher propensity for breakage. Here, we use ancestral genome reconstructions to demonstrate a near-perfect correlation between gene density and evolutionary rearrangement breakpoints. Simulations based on functional features in the human genome show that this pattern is best explained as the outcome of DNA breaks that occur in open chromatin regions coming into 3D contact in the nucleus. Our model explains how rearrangements reorganize the order of genes in an evolutionary neutral fashion and provides a basis for understanding the susceptibility of “fragile regions” to breakage.

The Genomicus web server (http://www.genomicus.biologie.ens.fr/genomicus) is a visualization tool allowing comparative genomics in four different phyla (Vertebrate, Fungi, Metazoan and Plants). It provides access to genomic information from extant species, as well as ancestral gene content and gene order for vertebrates and flowering plants. Here we present the new features available for vertebrate genome with a focus on new graphical tools. The interface to enter the database has been improved, two pairwise genome comparison tools are now available (KaryoView and MatrixView) and the multiple genome comparison tools (PhyloView and AlignView) propose three new kinds of representation and a more intuitive menu. These new developments have been implemented for Genomicus portal dedicated to vertebrates. This allows the analysis of 68 extant animal genomes, as well as 58 ancestral reconstructed genomes. The Genomicus server also provides access to ancestral gene orders, to facilitate evolutionary and comparative genomics studies, as well as computationally predicted regulatory interactions, thanks to the representation of conserved non-coding elements with their putative gene targets., (© The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Development of neuronal circuits is controlled by evolutionarily conserved axon guidance molecules, including Slits, the repulsive ligands for roundabout (Robo) receptors, and Netrin-1, which mediates attraction through the DCC receptor. We discovered that the Robo3 receptor fundamentally changed its mechanism of action during mammalian evolution. Unlike other Robo receptors, mammalian Robo3 is not a high-affinity receptor for Slits because of specific substitutions in the first immunoglobulin domain. Instead, Netrin-1 selectively triggers phosphorylation of mammalian Robo3 via Src kinases. Robo3 does not bind Netrin-1 directly but interacts with DCC. Netrin-1 fails to attract pontine neurons lacking Robo3, and attraction can be restored in Robo3(-/-) mice by expression of mammalian, but not nonmammalian, Robo3. We propose that Robo3 evolution was key to sculpting the mammalian brain by converting a receptor for Slit repulsion into one that both silences Slit repulsion and potentiates Netrin attraction., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Pufferfish species of the Tetraodontidae family carry the smallest genomes among vertebrates. Their compressed genomes are thought to be enriched for functional DNA compared to larger vertebrate genomes, and they are important models for comparative genomics. The significance of pufferfish as model organisms in comparative genomics is due to the availability of two sequenced genomes, that of spotted green pufferfish (Tetraodon nigroviridis) and fugu (Takifugu rubripes). However, there is only a very limited utilization of pufferfish as an experimental model organism, due to the lack of established husbandry and developmental genetics protocols. In this study, we provide the first description of the normal embryonic development of Tetraodon nigroviridis. Embryos were obtained by in vitro fertilization of eggs, and subsequent development was monitored by brightfield microscopy at constant temperature. Tetraodon development was divided into distinct stages based on diagnostic morphological features, which were adopted from published literature on normal development of other fish species like medaka (Oryzias latipes), zebrafish (Danio rerio), and fugu. Tetraodon embryos show more similar morphologies to medaka than to zebrafish, reflecting its phylogenetic position. The early developmental stage series described in this study forms the foundation for the utilization of tetraodon as an experimental model organism for comparative developmental studies.

Gene loss is one of the main drivers in the evolution of genomes and species. The demonstration that a gene has been lost by pseudogenization is truly complete when one finds the pseudogene in the orthologous genomic region with respect to active genes in other species. In some cases, the identification of such orthologous loci is not possible because of chromosomal rearrangements or if the gene of interest has not yet been sequenced. This question is particularly important in the case of birds because the genomes of avian species possess only about 15,000 predicted genes, in comparison with 20,000 in mammals. Yet, gene loss raises the question of which functions are affected by the changes in gene counts. We describe a systematic approach that makes it possible to demonstrate gene loss in the chicken genome even if a pseudogene has not been found. By using phylogenetic and synteny analysis in vertebrates, genome-wide comparisons between the chicken genome and expressed sequence tags, RNAseq data analysis, statistical analysis of the chicken genome, and radiation hybrid mapping, we show that resistin, TNFα, and PAI-1 (SERPINE1), three genes encoding adipokines inhibiting insulin sensitivity, have been lost in chicken and zebra finch genomes. Moreover, omentin, a gene encoding an adipokine that enhances insulin sensitivity, has also been lost in the chicken genome. Overall, only one adipokine inhibiting insulin sensitivity and five adipokines enhancing insulin sensitivity are still present in the chicken genome. These genetic differences between mammals and chicken, given the functions of the genes in mammals, would have dramatic consequences on chicken endocrinology, leading to novel equilibriums especially in the regulation of energy metabolism, insulin sensitivity, as well as appetite and reproduction., (© The Author 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Extant genomes share regions where genes have the same order and orientation, which are thought to arise from the conservation of an ancestral order of genes during evolution. Such regions of so-called conserved synteny, or synteny blocks, must be precisely identified and quantified, as a prerequisite to better understand the evolutionary history of genomes., Results: Here we describe PhylDiag, a software that identifies statistically significant synteny blocks in pairwise comparisons of eukaryote genomes. Compared to previous methods, PhylDiag uses gene trees to define gene homologies, thus allowing gene deletions to be considered as events that may break the synteny. PhylDiag also accounts for gene orientations, blocks of tandem duplicates and lineage specific de novo gene births. Starting from two genomes and the corresponding gene trees, PhylDiag returns synteny blocks with gaps less than or equal to the maximum gap parameter gap(max). This parameter is theoretically estimated, and together with a utility to graphically display results, contributes to making PhylDiag a user friendly method. In addition, putative synteny blocks are subject to a statistical validation to verify that they are unlikely to be due to a random combination of genes., Conclusions: We benchmark several known metrics to measure 2D-distances in a matrix of homologies and we compare PhylDiag to i-ADHoRe 3.0 on real and simulated data. We show that PhylDiag correctly identifies small synteny blocks even with insertions, deletions, incorrect annotations or micro-inversions. Finally, PhylDiag allowed us to identify the most relevant distance metric for 2D-distance calculation between homologies.

Vertebrate evolution has been shaped by several rounds of whole-genome duplications (WGDs) that are often suggested to be associated with adaptive radiations and evolutionary innovations. Due to an additional round of WGD, the rainbow trout genome offers a unique opportunity to investigate the early evolutionary fate of a duplicated vertebrate genome. Here we show that after 100 million years of evolution the two ancestral subgenomes have remained extremely collinear, despite the loss of half of the duplicated protein-coding genes, mostly through pseudogenization. In striking contrast is the fate of miRNA genes that have almost all been retained as duplicated copies. The slow and stepwise rediploidization process characterized here challenges the current hypothesis that WGD is followed by massive and rapid genomic reorganizations and gene deletions.