Background: The Atlantic salmon, Salmo salar L., is a prominent member of the Salmonidae family, and has been the focus of intense research because of its environmental and economic significance as an iconic sporting species and its global importance as an aquaculture species. Furthermore, salmonids constitute ideal organisms for the study of evolution by gene duplication as they are pseudotetraploid descendants of a common ancestor whose genome was duplicated some 25 to 100 million years ago. Whole-genome duplication is considered a major evolutionary force capable of creating vast amounts of new genetic material for evolution to act upon, promoting speciation by acquisition of new traits. Recently, large-scale comparison of paralogous genes in Atlantic salmon suggested that asymmetrical selection was acting on a significant proportion of them. However, to elucidate the physiological consequences of gene and genome duplications, studies integrating molecular evolution and functional biology are crucial. To this end, sequence and molecular analyses were performed on duplicated Elovl5 fatty-acyl elongases of Atlantic salmon, as they are responsible for a rate-limiting reaction in the elongation process of long-chain polyunsaturated fatty acids (LC-PUFA), critical components of all vertebrates. The aim of the research presented here was to investigate the role of gene duplication as an evolutionary process capable of creating genetic novelty, and to identify the potential ecological and physiological implications. Results: Linkage analyses indicated that both fatty-acyl elongases segregated independently and located elovl5 duplicates on different linkage groups. Genetic mapping using microsatellites identified in each elovl5 locus assigned elovl5a and elovl5b to chromosomes ssa28 and ssa13, respectively. In silico sequence analysis and selection tests indicated that both salmon Elovl5 proteins were subject to purifying selection, in agreement with previous results showing indistinguishable substrate specificities. Gene expression and promoter analysis indicated that Elovl5 duplicates differed in response to dietary lipids and tissue expression profile. Lipid biosynthesis and metabolic gene expression profiling performed in Atlantic salmon SHK-1 cells, suggested that the control of lipid homeostasis in fish is similar to that described in higher vertebrates, and revealed the particular importance of Lxr and Srebp transcription factors (TFs) in the regulation of LC-PUFA biosynthetic enzymes. Sequence comparison of upstream promoter regions of elovl5 genes showed intense differences between duplicates. Promoter functional analysis by co-transfection and transcription factor transactivation showed that both elovl5 duplicates were upregulated by Srebp overexpression. However, elovl5b exhibited a higher response and its promoter contained a duplication of a region containing response elements for Srebp and NF-Y cofactors. Furthermore, these studies indicated an Lxr/Rxr dependant response of elovl5a, which was not observed in elovl5b. Analysis of the genomic sequences of elovl5 duplicates by comparison to various sequence databases showed an asymmetrical distribution of transposable elements (TEs) in both introns and promoter regions. Further comparison to introns of the single elovl5 gene in pike indicated much higher TE distribution in salmon genes compared to the pike. Conclusions: Although not conclusive, the most parsimonious origin for the salmon elovl5 duplicates is that they are derived from a WGD event. This conclusion is also supported by the close similarity of two elovl5 paralogs in the recently available rainbow trout genome. Regardless of their origin, Atlantic salmon elovl5 genes have been efficiently retained in the genome under strong functional constraints indicating a physiological requirement for both enzymes to be functionally active. In contrast, upstream promoter regions have strongly diverged from one another, indicating a relaxation of purifying selection following the duplication event. This divergence of cis-regulatory regions has resulted in regulatory diversification of the elovl5 duplicates and regulatory neofunctionalisation of elovl5a, which displayed a novel Lxr/Rxr-dependant response not described in sister or other vertebrate lineages. Promoter analysis indicated that the observed elovl5 differential response to dietary variation could be partly attributed to varying transcriptional regulation driven by lipid-modulated TFs. The distribution of TEs in elvol5 genes of Atlantic salmon shows a clear increase in TE mobilisation after the divergence of esocids and salmonids. This must have occurred after the elongase duplication and thus the salmonid WGD event and contributes to the observed regulatory divergence of elovl5 paralogs.

Regulation of expression DLX1 gene, whose elevated levels are detected in patients with acute myeloid leukemia with FLT3-ITD mutations, is not still completely explored topic. The first aim of this study was to determine which selected signaling pathways regulate gene expression of DLX1. ERK a JNK pathways were selected by using qRT-PCR and western blot. These pathways cause activation of the transcription factor AP-1 subunits, the AP-1 putative promoter binding site was identified also in the promoter of the DLX1 gene. The second aim of this study was to test the hypothesis on the regulation of gene expression of DLX1 (via ERK/JNK pathway) through AP-1 binding site on the promoter. Dual luciferase assay using luminescent luciferase activity was performed to test this hypothesis. Gene of the luciferase is contained in the used luciferase vector. The short and the long part of the DLX1 promoter (around AP-1 site) were inserted before the gene of the luciferase in the constructs used in this method. The results of this study indicate that the regulation of gene expression through AP-1 promoter binding site is important but not sufficient part of the regulatory cascade running through ERK and JNK pathway. There must be another transcription factors activated by ERK1/2 kinase which are probably also involved in...

The ParaHox cluster is the evolutionary sister of the Hox cluster. Like the Hox cluster, the ParaHox cluster is subject to complex regulatory phenomena such as collinearity. Despite the breakup of the ParaHox cluster within many animals, intact and collinear clusters have now been discovered within the chordate phyla in amphioxus and the vertebrates, and more recently within the hemichordates and echinoderms. The archetypal ParaHox cluster of amphioxus places it in a unique position in which to examine the regulatory mechanisms controlling ParaHox gene expression within the last common ancestor of chordates, and perhaps even the wider Deuterostomia. In this thesis, the genomic and regulatory landscape of the amphioxus ParaHox cluster is characterised in detail. New genomic and transcriptomic resources are used to better characterise the B.floridae ParaHox cluster and surrounding genomic region, and conserved non-coding regions and regulatory motifs are identified across the ParaHox cluster of three species of amphioxus. In conjunction with this, the impact of retrotransposition upon the ParaHox cluster is examined and analyses of transposable elements and the AmphiSCP1 retrogene reveal that the ParaHox cluster may be more insulated from outside influence than previously thought. Finally, the detailed analyses of a regulatory element upstream of AmphiGsx reveals conserved mechanisms regulating Gsx CNS expression within the chordates, and TCF/Lef is likely a direct regulator of AmphiGsx within the CNS. The work in this thesis makes use of new genomic and transcriptomic resources available for amphioxus to better characterise the genomic and regulatory landscape of the amphioxus ParaHox cluster, serving as a basis for the improved identification and characterisation of functional regulatory elements and conserved regulatory mechanisms. This work also highlights the potential of Ciona intestinalis as a ‘living test tube' to allow the detailed characterisation of amphioxus ParaHox regulatory elements.

Licenciatavhandling Uppsala : Sveriges lantbruksuniversitet, 2009. / Härtill 2 uppsatser.

Licenciatavhandling Uppsala : Sveriges lantbruksuniversitet, 2009.

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv., 2008. / Härtill 4 uppsatser.

En production animale, notamment dans les filières bovines, il est dun intérêt économique daugmenter la quantité dhormone de croissance dans la circulation sanguine. La ghréline est un peptide principalement produit au niveau de la paroi stomacale. Ce ligand endogène au GHSR stimule la sécrétion hypophysaire de lhormone de croissance. Peptide orexigène, la ghréline est impliquée dans les mécanismes relatifs au maintien de lhoméostasie énergétique. Dans loptique daméliorer les performances animales, deux approches de la ghréline ont été effectuées. La première approche consiste en létude des gènes bovins codant pour la ghréline (bGHRL) et son récepteur (bGHSR). Ces deux gènes ont été respectivement localisés sur BTA 22 et BTA 1. Quatorze polymorphismes ont été détectés sur ces deux gènes et trois dentre eux affectent la structure primaire du GHSR bovin. Des associations, à différents niveaux de signification, entre certains de ces 14 sites polymorphiques et des traits de production et de conformation ont été mis en évidence au sein dun groupe de 127 taureaux Holstein sur base de leurs descendances directes présentes en Région Wallonne. La seconde approche aborde les effets dune immunisation passive contre la ghréline sur des rats mâles en croissance en comparaison à celles contre la leptine et la cholécystokinine. Lors dune alimentation équilibrée, le traitement envers la ghréline sur ces rats na pas influencé la croissance et lingestion par rapport aux animaux témoins. Des effets ont été observés entre les différentes immunomodulations au niveau des paramètres de croissance, dingestion et endocrinologiques. Les présents résultats invitent à de nouvelles investigations des gènes bGHRL et bGHSR sur des données relatives à dautres populations/races bovines et de limmunomodulation de la ghréline lors de conditions dexpérimentation différentes (alimentation déséquilibrée, stade physiologique, espèce, etc.). Ces investigations pourraient être valorisées en sélection et production animale mais également en médecine tant humaine que vétérinaire.