The differentiation of plant vascular tissue is regulated by plant hormones and transcription factors. One of the key plant hormones involved in this process is auxin. Auxin signals are mediated by auxin response factor transcription factors (ARFs). These transcription factors are involved in the perception of auxin signals and the subsequent activation or deactivation of suites of downstream genes. Based on its mutant phenotype, one of the most interesting members of this family is the ARF MONOPTEROS (MP). This thesis investigates the role played by MP in secondary vascular differentiation, as well as taking a look at other molecular aspects of secondary vascular differentiation, with a focus on the model plants Arabidopsis thaliana and poplar (Populus trichocarpa and hybrid poplar). A dexamethasone inducible RNAi silencing strategy was developed, and transgenic Arabidopsis lines produced. When silencing was induced in these lines from germination, a phenotype closely resembling the mp mutant was observed. When MP silencing was induced in bolting stems, early senescence, as well as a dramatic reduction in interfascicular fibre production was observed, and these stems were thinner and less rigid than empty vector controls. RNA from these stems was isolated and used in a global transcript profiling microarray experiment. This experiment showed that several auxin-related genes, as well as several transcription factors, were differentially regulated in response to MP silencing. Because Arabidopsis is not a typical woody plant, further investigation into the role played by MP in wood formation was done using the model tree poplar. A BLAST search of a poplar xylem EST database identified a single promising partial sequence. Based on this sequence information, a poplar MP homolog was isolated and named PopMP1. The full-length sequence of this gene demonstrated remarkable structural conservation when compared with that of Arabidopsis. Subsequent complete sequencing of the poplar genome revealed a second copy of the MP gene in poplar and named PopMP2. Expression profiling across a range of tissues suggests that subfunctionalization has occurred between the two copies. Overexpression transgenic lines for PoptrMP1 were developed. AtHB8 is known to be regulated by MP in Arabidopsis, and a poplar HB8 homolog was upregulated in the transgenic lines. However, no obvious physical phenotype in these lines was apparent. To investigate the transcriptome-wide changes associated with initiation of cambium formation in poplar stems, a global transcript profiling experiment was performed. Out of 15400 genes tested, 2320 met an arbitrary cutoff of >1.3 fold and p-value <0.05 and were labeled differentially expressed (DE). These included several transcription factors and showed remarkable similarity to analogous data from Arabidopsis. The conclusions drawn from this thesis support the hypothesis that MP plays roles in later development, and do not rule out the possibility that MP is directly involved in wood development. The data reported also offer a large number of candidate for further investigation into the genetic control of wood development.

Self-incompatibility (SI) is an important genetic mechanism to prevent the inbreeding of flowering plants and also an excellent system for studying cell-cell recognition and signal transduction. During evolution, several SI systems have been evolved. A unique SI system widely spreads in the grasses. In the grasses, two unlinked, multi-allelic loci (S and Z) determine SI specificity. A putative self-incompatibility gene (Bm2) was previously cloned. In this study, the role of Bm2 in self-incompatibility was investigated first. The cDNA homologues of Bm2 were sequenced from two pollen-only mutants. The results indicated that Bm2 is not the one of SI genes in Phalaris, but represents a subclass of thioredoxin h. Thus a map-based cloning strategy was then adopted to clone the SI genes from Phalaris. Fine linkage maps of the S and Z regions were constructed. RFLP probes from wheat, barley, oat and rye were screened and the S locus was delimited to 0.26 cM and the Z locus to 1.0 cM from one side using specially designed segregating populations. The S locus was located to the sub-centromere region of triticeae chromosome group 1 and the Z locus to the middle of the long arm of group 2. Finally, barley and rice bacterial artificial chromosome (BAC) clones corresponding to the S and Z region were identified to analyse the chromosome structures and to seek candidate SI genes. The abundant repetitive sequences in the identified barley BAC clones limit their usefulness. Identification of Rice BAC clones orthologous to the S and Z regions open the gate to use rice genome information to clone SI genes from the grasses. A positive rice clone (139.9 kb) orthologous to the S region contained 19 predicted genes. Several of these genes might be involved in pollen tube germination and pollen-stigma interaction, which are the major parts of SI reaction. A positive clone (118.9 kb) orthologous to the Z region gave 16 predicted genes. The predicted genes on the outmost ends of these clones could be used to construct contigs to cover the S and Z regions and delimit the S and Z loci in the grasses. / Thesis (Ph.D.)--Department of Plant Science, 2001.

Under certain environmental conditions, some varieties of wheat synthesize the enzyme alpha amylase late in grain ripening, even in the absence of rain or sprouting. The resulting grain has a sound appearance but can be unsuitable for end-product applications due to the presence of late maturity alpha amylase (LMA) activity. Reduction of LMA and the development of cultivars tolerant to boron toxic soils are high priority traits in the WA wheat breeding program and the use of molecular markers closely linked to these traits for marker assisted selection (MAS) is highly desirable. The aims of this study were to take a genomics approach to provide detailed structural information for the region on wheat chromosome 7BL in which quantitative trait loci (QTLs) for LMA and boron tolerance (Bo1) have been mapped. Once the structure had been determined, this then laid the foundation for further studies to investigate the function of putative candidate genes identified within this region. The research involved the use of bioinformatic tools and rice/wheat synteny to investigate the structure of this chromosome region, followed by the use of molecular probes to isolate genomic DNA clones (BAC clones) corresponding to this region. A two-step bioinformatics strategy was used, involving (1) alignment of portions of the wheat and rice genomes, to identify rice genomic regions syntenic to wheat group 7L and (2) selection of candidate genes from those regions of the rice genome. The selected candidate genes included an anion transporter, as a candidate gene for boron tolerance, and GAMYB-like genes, as candidate genes for LMA. The GAMYB class of transcription factors identified were of particular interest because of published literature indicating its importance in controlling ƒÑ-amylase levels in cereal grains. The key phenotype of interest in this thesis is LMA and different levels of expression of ƒÑ-amylase are a key feature of this phenotype. Molecular markers and candidate genes were then used to screen two BAC libraries, one derived from the French cultivar, ¡¥Renan¡¦ and the other derived from Aegilops tauschii (the source of the D genome of wheat). About 300 BAC clones corresponding to the chromosome region of interest were obtained. Of these, 8 BAC clones (6 chosen through hybridization to a GAMYB-like probe, and 2 from wheat ESTs anchored to the rice genome) were selected for sequencing, allowing for the development of new microsatellite and single-nucleotide polymorphism (SNP) markers and for the discovery of novel transposable elements that provide a rich source of polymorphism for the development of additional markers. Novel microsatellite and SNP markers that were identified from the BAC clone sequence were mapped on the Cranbrook/Halberd doubled haploid (DH) mapping population. Markers were located to chromosomes 7AL, 7BL and 7DL. New markers derived from the BAC sequence information were used to anchor the BAC clones to the genetic map and develop a framework physical-genetic map. An automated annotation pipeline has been established and was used to annotate selected contigs of the sequenced BAC clones. A new marker assisted selection strategy, termed Multiplex Trait Signature (MuTs) analysis, was developed and tested on 39 wheat cultivars of known LMA phenotype. MuTs provides a graphical genotype of individuals for a particular chromosomal region and is a convenient tool for interrogating genetic similarity in the individuals surveyed. Based on assays of 22 markers (12 spanning the LMA QTL on chromosome 7BL and 10 spanning the LMA QTL on chromosome 3BS) on these 39 wheat cultivars, it was found that the varieties can be grouped according to pedigree and provides a tool for interpreting LMA status for a variety. Validation of the 7BL LMA and boron tolerance (Bo1) QTL regions was achieved using a targeted mapping approach using the doubled haploid population Pastor/RAC891 using published molecular markers and markers developed in this thesis. The main outcome of this study is that the genomic organisation of this region on chromosome 7BL is complex, and that the identification of candidate genes in wheat controlling 1) tolerance of cultivars to boron toxic soils and 2) pathways regulating the expression of LMA, is likely to involve the interplay of a network of regulatory genes.

As described over 20 years ago with the discovery of RNA interference (RNAi), double-stranded RNAs occupied key roles in regulation and as defense-line in animal cells. This thesis focuses on metazoan microRNAs (miRNAs). These small non-coding RNAs are distinguished from their small-interfering RNA (siRNA) relatives by their tightly controlled, efficient and flexible biogenesis, together with a broader flexibility to target multiple mRNAs by a seed imperfect base-pairing. As potent regulators, miRNAs are involved in mRNA stability and post-transcriptional regulation tasks, being a conserved mechanism used repetitively by the evolution, not only in metazoans, but plants and unicellular organisms. Through a comprehensive revision of the current animal miRNA model, the canonical pathway dominates the extensive literature about miRNAs, and served as a scaffold to understand the scenes behind the regulatory landscape performed by the cell. The characterization of a diverse set of non-canonical pathways has expanded this view, suggesting a diverse, rich and flexible regulatory landscape to generate mature miRNAs. The production of miRNAs, derived from isolated or clustered transcripts, is an efficient and highly conserved mechanism traced back to animals with high fidelity at family level. In evolutionary terms, expansions of miRNA families have been associated with an increasing morphological and developmental complexity. In particular, the Chordata clade (the ancient cephalochordates, highly derived and secondary simplified tunicates, and the well-known vertebrates) represents an interesting scenario to study miRNA evolution. Despite clearly conserved miRNAs along these clades, tunicates display massive restructuring events, including emergence of highly derived miRNAs. As shown in this thesis, model organisms or vertebrate-specific bias exist in current animal miRNA annotations, misrepresenting more diverse groups, such as marine invertebrates. Current miRNA databases, such as miRBase and Rfam, classified miRNAs under different definitions and possessed annotations that are not simple to be linked. As an alternative, this thesis proposes a method to curate and merge those annotations, making use of miRBase precursor/mature annotations and genomes together with Rfam predicted sequences. This approach generated structural models for shared miRNA families, based on the alignment of their correct-positioned mature sequences as anchors. In this process, the developed structural curation steps flagged 33 miRNA families from the Rfam as questionable. Curated Rfam and miRBase anchored-structural alignments provided a rich resource for constructing predictive miRNA profiles, using correspondent hidden Markov (HMMs) and covariance models (CMs). As a direct application, the use of those models is time-consuming, and the user has to deal with multiple iterations to achieve a genome-wide non-overlapping annotation. To resolve this, the proposed miRNAture pipeline provides an automatic and flexible solution to annotate miRNAs. It combines multiple homology approaches to generate the best candidates validated at sequence and structural levels. This increases the achievable sensitivity to annotate canonical miRNAs, and the evaluation against human annotation shows that clear false positive calls are rare and additional counterparts lie in retained-introns, transcribed lncRNAs or repeat families. Further development of miRNAture suggests an inclusion of multiple rules to distinguish non-canonical miRNA families. This thesis describes multiple homology approaches to annotate the genomic information from a non-model chordate: the colonial tunicate Didemnum vexillum. Detected high levels of genetic variance and unexpected levels of DNA degradation were evidenced through a comprehensive analysis of genome-assembly methods and gene annotation. Despite those challenges, it was possible to find candidate homeobox and skeletogenesis- related genes. On its own, the ncRNA annotation included expected conserved families, and an extensive search of the Rhabdomyosarcoma 2-associated transcript (RMST) lncRNA family traced-back at the divergence of deuterostomes. In addition, a complete study of the annotation thresholds suggested variations to detect miRNAs, later implemented on the miRNAture tool. This chapter is a showcase of the usual workflow that should follow comprehensive sequencing, assembly and annotation project, in the light of the increasing research approaching DNA sequencing. In the last 10 years, the remarkable increment in tunicate sequencing projects boosted the access to an expanded miRNA annotation landscape. In this way, a comprehensive homology approach annotated the miRNA complement of 28 deuterostome genomes (including current 16 reported tunicates) using miRNAture. To get proper structural models as input, corrected miRBase structural alignments served as a scaffold for building correspondent CMs, based on a developed genetic algorithm. By this means, this automatic approach selected the set of sequences that composed the alignments, generating 2492 miRNA CMs. Despite the multiple sources and associated heterogeneity of the studied genomes, a clustering approach successfully gathered five groups of similar assemblies and highlighted low quality assemblies. The overall family and loci reduction on tunicates is notorious, showing on average 374 microRNA (miRNA) loci, in comparison to other clades: Cephalochordata (2119), Vertebrata (3638), Hemichordata (1092) and Echinodermata (2737). Detection of 533 miRNA families on the divergence of tunicates shows an expanded landscape regarding currently miRNA annotated families. Shared sets of ancestral, chordates, Olfactores, and specific clade-specific miRNAs were uncovered using a phyloge- netic conservation criteria. Compared to current annotations, the family repertories were expanded in all cases. Finally, relying on the adjacent elements from annotated miRNAs, this thesis proposes an additional syntenic support to cluster miRNA loci. In this way, the structural alignment of miR-1497, originally annotated in three model tunicates, was expanded with a clear syntenic support on tunicates.

Les cultivars modernes de canne à sont de hauts polyploïdes, aneuploïdes issus de croisements interspécifiques entre deux espèces polyploïdes, une espèce sucrée domestiquée Saccharum officinarum et une espèce sauvage Saccharum spontaneum. Le gène majeur de résistance durable à la rouille brune, Bru1 a été identifié chez le cultivar de canne à sucre R570. Une approche de clonage positionnel de ce gène a été entreprise et a permis de construire une première carte physique. Elle comprend sept haplotype hom(é)ologues dont un correspond à l'haplotype cible porteur du gène Bru1 qui comprend sept clones BAC qui ne se chevauchent que partiellement laissant deux espaces non couverts. Il a été montré que cette situation résulte de la présence d'une insertion dans l'haplotype porteur de Bru1. Pour combler les deux espaces présents sur l'haplotype cible, deux stratégies utilisant l'annotation des BAC ont été employées : (i) une se basant sur la conservation des gènes existante entre les différents haplotypes hom(é)ologues de la région de Bru1 et (ii) une en utilisant des marqueurs flanquants les deux espaces. Ces stratégies no us ont permis de combler un des deux espaces, de couvrir partiellement le deuxième espace et de montrer que le gène de résistance se situerait dans l'insertion. Nous avons identifié un gène candidat correspondant à une Sérine/Thréonine kinase située dans l'insertion. Des tests d'expression ont été effectués en condition normale afin de vérifier si ce gène est exprimé mais aucune amplification n'a été obtenue. Parallèlement, la recherche de l'origine de l'insertion présente sur l'haplotype cible a été entreprise en retraçant son origine dans la généalogie de notre cultivar d'étude R570 et en criblant une banque de clones de Saccharum contenant différentes espèces de canne à sucre. Les résultats sur la généalogie tendent à nous dire que cette insertion est ancienne et aurait été transmise à R570 via S. barberi. / Modern sugarcane cultivars are high polyploids, aneuploids derived interspecific crosses between two polyploid species, domesticated sugar species Saccharum officinarum and a wild species Saccharum spontaneum. The major gene for sustainable resistance to brown rust, Bru1 was identified in the modern cultivar R570. An map-based approach has been undertaken and has built a first physical map. It includes seven haplotype hom(e)ologous which one corresponds to the haplotype carrying Bru1 which includes seven BACs that overlap only partially, including two gaps. It was shown that this situation results from the presence of an insertion in the target haplotype. To fill the two gaps, two strategies using the annotation of BACs were used: (i) Based on the good genes conservation between haplotypes hom(e)ologous and (ii) by using markers flanking the two gaps. These strategies have enabled us to fill one of two gaps, partially cover the second and show that the resistance gene would be in the insertion. We identified a candidate gene corresponding to a serine/threonine kinase located in the insertion. Expression tests were performed in normal condition to see if this gene is expressed but no amplification was obtained. Meanwhile, the search for the origin of the insertion present on the target haplotype was undertaken by tracing its origin in the genealogy of R570 and analysing a library of clones of Saccharum spp containing different types of cane sugar. Results on the genealogy we tend to say that this insertion is old and were sent to R570 via S. barberi.

Sorgo sacarino e cana-de-açúcar são duas importantes gramíneas com fins potencialmente bioenergéticos. No entanto, apesar do conhecido relacionamento evolutivo, os genomas dessas espécies diferem em complexidade e tamanho. O sorgo, Sorghum bicolor, é diploide, com número básico de cromossomos igual a dez, os quais totalizam ~ 726 Mb já sequenciadas. Já a cana cultivada, Saccharum × officinarum, é um autopoliploide com frequente aneuploidia, e apresenta genoma monoploide estimado em ~ 1 Gb. Provavelmente, decorre deste fato, e dos cruzamentos interespecíficos que originaram as variedades atuais, a relativa dificuldade em se realizar estudos genéticos em cana, e, como consequência, em se incrementar os trabalhos de melhoramento na espécie. Nesse contexto, a possibilidade de integrar estudos de mapeamento entre sorgo e cana torna-se viável dado o emprego de metodologias apropriadas. O presente trabalho objetivou mapear e comparar QTLs para caracteres agro-industriais nos genomas de ambas as espécies, baseando-se no relacionamento evolutivo existente entre elas. Para tanto, foram utilizadas duas populações de mapeamento. A população de sorgo sacarino foi constituída por 223 RILs genotipadas por mais de cem mil marcadores baseados em GBS fisicamente mapeados contra o genoma da espécie. A população de cana-de-açúcar constituiu-se de uma progênie F1 segregante com 153 indivíduos genotipados por 500 marcadores baseados em géis (SSR e TRAP) e 7.049 marcadores baseados em GBS, segregando em dose única. Esses marcadores possibilitaram a construção de um mapa genético informativo e saturado, com 993 marcadores distribuídos ao longo de 223 grupos de ligação, totalizando 3.682,05 cM. Ambas as populações foram avaliadas para quatro caracteres de interesse bioenergético: altura de colmos, toneladas de colmos ou de massa verde por hectare, e porcentagens de pol de caldo e de fibra. Modelos mistos foram utilizados para a análise dos dados fenotípicos, evidenciando a existência de interação genótipo-ambiente a partir da estruturação de matrizes de variâncias-covariâncias genéticas. As médias ajustadas conjunta e marginalmente foram utilizadas na descoberta de QTLs. Para este fim, modelos de mapeamento de múltiplos intervalos uni- e multivariados foram utilizados e determinaram a descoberta de 53 e 36 regiões contendo QTLs para as populações de sorgo e cana, respectivamente, para o conjunto dos quatro caracteres. Os genomas foram comparados utilizando os marcadores baseados em GBS de cana com informação posicional em relação ao genoma do sorgo. Um total de 16 regiões sintênicas identificadas entre as espécies possibilitaram inferências a respeito do controle evolutivamente conservado dos caracteres relacionados. Mais oito regiões foram adicionadas a estas após análise de marcadores individualmente para a população de cana. A descoberta dessas regiões subjacente à variação de caracteres bioenergéticos sugere aplicações na clonagem de genes e na seleção assistida por marcadores, beneficiando os programas de melhoramento de ambas as espécies. / Sweet sorghum and sugarcane are two important grasses for bioenergy purposes. However, despite their known evolutionary relationship, the genomes of these species differ in complexity and size. Sorghum bicolor is a diploid species, with basic chromosome number of ten and ~ 726 Mb completely sequenced, whereas Saccharum × officinarum has a autopolyploid genome with frequent aneuploidy and monoploid size estimated at ~ 1 Gb. Therefore, genetic studies and breeding in sugarcane is challenging. In this context, the possibility of integrating mapping studies between sorghum and sugarcane becomes feasible given the recent development of appropriate methodologies. In this work, we aimed to map and compare QTLs for bioenergy traits in both species. To do this, two mapping populations were used. The population of sorghum consisted of 223 RILs genotyped by more than one hundred thousand GBS-based markers, which were physically mapped against the species genome. The population of sugarcane is an F1 segregating progeny with 153 individuals genotyped by 500 gel-based (SSR and TRAP) and 7,049 GBS-based single-dose markers. These markers allowed the construction of an informative and dense genetic map with 993 markers belonging to 223 linkage groups and spanning 3,682.05 cM. Both populations were evaluated for four bioenergy traits: stalk height, prodution in tons per hectare, and percentages of pol and fiber. Mixed models were used to analyze phenotypic data and showed genotype-by-environment interaction on their genetic variance-covariance structures. Joint and marginal adjusted means were used for QTL discovery. Toward this end, univariate and multivariate multiple interval mapping models were used, and a total of 53 and 36 QTLs were found for sorghum and sugarcane, respectively. Comparison of the genomes were based on GBS markers in sugarcane with relative sorghum chromosome information. A total of 16 syntenic regions were identified between the species, allowing inferences in relation to evolutionary conserved control of the related traits. In addition, eight regions were also identified by considering single marker analyses. The discovery of QTLs underlying such bioenergy traits may suggest further applications in gene cloning and marker assisted selection for both sweet sorghum and sugarcane species.

Les Poaceae aussi appelées Graminées forment une importante famille botanique regroupant près de 12 000 espèces en plus de 700 genres dont les céréales. Cette famille présente un intérêt économique majeur car elle est importante dans la nutrition humaine et animale. De ce fait, cette famille a été très étudiée en génomique comparée depuis les années 1990 révélant une grande conservation de la structure de leur génome depuis leur divergence d’un ancêtre commun. Avec le séquençage de Brachypodium distachyon en 2009, nous avons réalisé l’analyse de son génome par l’identification de douze blocs de synténie avec les génomes séquencés du riz, du sorgho et du maïs ainsi que sept blocs de duplications partagées entre ces graminées. Ces données nous ont permis de suggérer que les cinq chromosomes modernes de Brachypodium sont issus de l’ancêtre commun des graminées constitué de douze chromosomes et ayant subi sept fusions au cours de l’évolution. Ces travaux nous ont permis de confirmer un possible génome ancêtre des graminées constitué de cinq chromosomes porteurs de près de 10 000 gènes et d’une taille minimale de près de 35Mb. Ensuite, sur la base des résultats de génomique comparée, nous nous sommes intéressés à l’évolution des différentes familles de micro-ARN (miARN). La comparaison de ces ARN non-codants réalisée pour le riz, le sorgho, le maïs et Brachypodium montre une conservation de cette famille chez les graminées avec 50% d’orthologues et 20% de paralogues. Sur la base des résultats de paléogénomique, nous avons proposé une modélisation de l’évolution des miARN qui corrobore l’hypothèse d’une origine très ancienne de ce mécanisme de « gene silencing ». Au-delà des nouvelles connaissances fondamentales générées au cours de ce travail de thèse sur l’évolution des génomes de graminées, les résultats que nous avons obtenus ont des applications potentielles dans le domaine de l’amélioration variétale, comme avec par exemple la possibilité de définir des marqueurs moléculaires de type COS (Conserved Orthologous Set). Ces marqueurs COS ont été mis en oeuvre pour l’étude de caractères agronomiques d’intérêt dans des espèces dont le génome n’est pas encore complètement séquencé comme le blé. / Poaceae also called Grasses are an important botanical family consisting in nearly 12,000 species in over 700 genres including cereals. This family is of major economic interest because it comprises cereals that are among the most important crops for human and animal nutrition. This family has been extensively studied in comparative genomics since the 1990s and showed a high degree of gene conservation among species since they diverged from a common ancestor. With the sequencing of Brachypodium distachyon in 2009, we performed an analysis of its genome by the identification of twelve synteny blocks with the sequenced genomes of rice, sorghum and maize and seven duplications blocks shared with these last grass species. These data allowed us to suggest the five chromosomes of Brachypodium are from the common ancestor composed of twelve chromosomes and having undergone seven fusions during the evolution. This work allowed us to confirm a possible grass ancestor with five chromosomes carrying almost 10,000 genes with a size of 35Mb. Then, based on these comparative genomics results, we studied more particularly the evolution of different families of microRNAs (miRNAs). The comparison of non-coding RNA from rice, sorghum, maize and Brachypodium showed conservation into this family for the grass species with 50% of orthologs and 20% of paralogs. Based on the paleogenomics results, we proposed an evolutionary scenario of miRNA genes, which supports the hypothesis of an ancient origin of this gene silencing mechanism in plants. Beyond the fundamental knowledge generated on the evolution of grass genomes during this PhD, these results have potential applications in breeding, for example with the possibility to identify COS (Conserved Orthologous Set) molecular markers. Such COS markers have been used for the study of agronomic traits in species not completely sequenced as wheat.

<p>Paralogs arise by duplications and belong to families. Ten paralogs (insulin; <i>IGF-1</i> and <i>-2</i>; <i>INSL3-6</i> and 3-relaxins) constitute the human insulin-relaxin family. The aim of this study was to outline the duplications that gave rise to the vertebrate insulin-relaxin genes and the chromosomal regions in which they reside. Neurotrophin and Trk-receptor families with more than 300, otherwise unrelated, families had paralogs in the regions hosting insulin/relaxin genes, defining two quadruplicate paralogy-regions, namely: insulin/IGF and INSL/relaxin paralogons. Thereby, the localization of insulin/relaxins in human shows that these regions were formed during two genome duplications at the stem of the vertebrates.</p><p>We characterized insulin-like genes (<i>INS-L1</i>, <i>-L2</i> and <i>-L3</i>) in the <i>Ciona intestinalis</i> genome, a species that split from the chordate lineage before the genome duplications. Conserved synteny between the Ciona region hosting the <i>INS-Ls</i> and two human paralogons as well as linkage of the actual paralogons, suggest that a segmental duplication gave rise to the entire region prior to the genome duplications. Synteny together with gene and protein structures demonstrate that <i>INS-L1</i> is orthologous to the vertebrate <i>INSLs</i>/relaxins, <i>INS-L2</i> to insulins and <i>INS-L3</i> to <i>IGFs</i>. This indicates that pro-orthologs of the insulin-relaxin family were formed before Ciona. Our analysis also implies that the INSL/relaxin ancestor switched receptor from tyrosine kinase- to GPCR-type. This probably occurred after the Ciona-stage, but before the genome duplications.</p><p>Using genes residing within the analyzed human paralogons that were present in a chromosomal region in the Ciona-human ancestor, we identified 37 segments with conserved synteny between the <i>Drosophila melanogaster</i> and human genomes. Orthologs residing in Ciona-, sea urchin- and the fly syntenic segments imply that such segments approximate an ancestral region from which the human paralogons originated.</p><p>To conclude, the human paralogons are remnants of genome duplications that in addition to segmental- and single duplications, shaped the extant vertebrate genomes. Using the quadruplicate paralogy-regions we were able to deduce duplication events of the insulin-relaxin genes and their chromosomal regions.</p>

Paralogs arise by duplications and belong to families. Ten paralogs (insulin; IGF-1 and -2; INSL3-6 and 3-relaxins) constitute the human insulin-relaxin family. The aim of this study was to outline the duplications that gave rise to the vertebrate insulin-relaxin genes and the chromosomal regions in which they reside. Neurotrophin and Trk-receptor families with more than 300, otherwise unrelated, families had paralogs in the regions hosting insulin/relaxin genes, defining two quadruplicate paralogy-regions, namely: insulin/IGF and INSL/relaxin paralogons. Thereby, the localization of insulin/relaxins in human shows that these regions were formed during two genome duplications at the stem of the vertebrates. We characterized insulin-like genes (INS-L1, -L2 and -L3) in the Ciona intestinalis genome, a species that split from the chordate lineage before the genome duplications. Conserved synteny between the Ciona region hosting the INS-Ls and two human paralogons as well as linkage of the actual paralogons, suggest that a segmental duplication gave rise to the entire region prior to the genome duplications. Synteny together with gene and protein structures demonstrate that INS-L1 is orthologous to the vertebrate INSLs/relaxins, INS-L2 to insulins and INS-L3 to IGFs. This indicates that pro-orthologs of the insulin-relaxin family were formed before Ciona. Our analysis also implies that the INSL/relaxin ancestor switched receptor from tyrosine kinase- to GPCR-type. This probably occurred after the Ciona-stage, but before the genome duplications. Using genes residing within the analyzed human paralogons that were present in a chromosomal region in the Ciona-human ancestor, we identified 37 segments with conserved synteny between the Drosophila melanogaster and human genomes. Orthologs residing in Ciona-, sea urchin- and the fly syntenic segments imply that such segments approximate an ancestral region from which the human paralogons originated. To conclude, the human paralogons are remnants of genome duplications that in addition to segmental- and single duplications, shaped the extant vertebrate genomes. Using the quadruplicate paralogy-regions we were able to deduce duplication events of the insulin-relaxin genes and their chromosomal regions.

No description available.