Glycerol metabolism in smelt (Osmerus mordax)

Rainbow smelt (Osmerus mordax) is a small fish that accumulates high amounts of glycerol in winter as a cryoprotectant. The principal objective of my thesis was to investigate the metabolic control of the synthesis and clearance (loss/catabolism) of glycerol in this species. As rainbow smelt lose 4-13% of their glycerol content daily, they have to vigorously synthesize this metabolite at temperatures below 0ﾟC to reach the exceptional levels reported (> 200 mM). In this context, the two main hypotheses of my thesis were: 1) glycerol loss/catabolism might be turned-off during the accumulation period and be turned on during the decrease period; and 2) the last step of glycerol synthesis, leading from glycerol-3-phosphate (G3P) to glycerol might be direct, involving a single reaction catalyzed by glycerol 3-phosphatase (G3Pase), an enzyme not yet convincingly described in any animal species. -- Activity and mRNA levels of glycerol kinase (GK), the first enzyme involved in glycerol breakdown, were measured in liver, the tissue displaying the highest activity in vertebrates. mRNA levels did not change over the cycle but GK activity did, suggesting a regulation at a post transcriptional level. Overall, GK activity was so low in liver, and in all other tissues assayed, that it likely has a minor quantitative role in glycerol clearance. Daily loss toward water was also determined over the cycle but was not lower over the accumulation period, ruling out the presence of any mechanisms to retain glycerol during this period. Taken together, these results suggest that glycerol levels are dictated primarily by the rate of glycerol synthesis. -- To produce glycerol, G3P can be directly dephosphorylated by a G3Pase, as in yeast (Saccharomyces cerevisiae) or can enter the triacylglycerol pool and then be released as glycerol via reactions well recognized in mammals. Lipid content and activities of G3Pase and key enzymes involved in lipid metabolism were measured over the glycerol cycle in liver, the site of glycerol synthesis. Lipid content did not change over the cycle and was too low to account for a significant amount of glycerol, but activities of G3Pase and of enzymes involved in lipid degradation were up-regulated early in the cycle, at the onset of glycerol accumulation. A second set of experiments was performed using isolated smelt hepatic cells incubated in optimal conditions to produce glycerol over 72h in the presence or absence of a potent inhibitor of lipid synthesis. It was shown that neither on-board lipids nor newly synthesized lipids could be the source of the glycerol produced, pointing out for the first time a direct dephosphorylation by G3Pase as the main source of glycerol in an animal species. -- The last objective of my thesis was to partially purify and characterize G3Pase from smelt liver. The enzyme is most likely a cytosolic, acidic, low molecular weight phosphatase, in contrast to other species where the enzyme was previously purified and that has an optimal activity at neutral pH. -- These results are crucial in terms of understanding glycerol metabolism in rainbow smelt but also for the elucidation of pathways involved in glycerol synthesis in vertebrates as they clearly showed that glycerol is not necessarily only a by-product of lipid degradation

Unusual duplication of the insulin-like receptor in the crustacean Daphnia pulex

Background: The insulin signaling pathway (ISP) has a key role in major physiological events like carbohydrate metabolism and growth regulation. The ISP has been well described in vertebrates and in a few invertebrate model organisms but remains largely unexplored in non-model invertebrates. This study is the first detailed genomic study of this pathway in a crustacean species, Daphnia pulex. Results: The Daphnia pulex draft genome sequence assembly was scanned for major components of the ISP with a special attention to the insulin-like receptor. Twenty three putative genes are reported. The pathway appears to be generally well conserved as genes found in other invertebrates are present. Major findings include a lower number of insulin-like peptides in Daphnia as compared to other invertebrates and the presence of multiple insulin-like receptors (InR), with four genes as opposed to a single one in other invertebrates. Genes encoding for the Dappu_InR are likely the result of three duplication events and bear some unusual features. Dappu_InR-4 has undergone extensive evolutionary divergence and lacks the conserved site of the catalytic domain of the receptor tyrosine kinase. Dappu_InR-1 has a large insert and lacks the transmembranal domain in the b-subunit. This domain is also absent in Dappu_InR-3. Dappu_InR-2 is characterized by the absence of the cystein-rich region. Real-time q-PCR confirmed the expression of all four receptors. EST analyses of cDNA libraries revealed that the four receptors were differently expressed under various conditions. Conclusions: Duplications of the insulin receptor genes might represent an important evolutionary innovation in Daphnia as they are known to exhibit extensive phenotypic plasticity in body size and in the size of defensive structures in response to predation

Variabilité du taux de croissance chez l'omble chevalier (Salvelinus alpinus) effets génétiques et physiologiques

Le potentiel de deux souches d'omble chevalier disponibles au Québec a été évalué dans le cadre de la mise en place éventuelle d'un programme de sélection visant à améliorer le taux de croissance. La variabilité génétique a été estimée pour chacune des souches et s' est avérée non seulement inférieure à celle de populations sauvages mais aussi à celles d'autres populations aquacoles de la même espèce. Puisqu'une trop faible variabilité génétique peut rapidement amener à l'apparition de consanguinité il a été suggéré d' augmenter la variabilité de ces souches avant la mise en place de tout programme de sélection . Les marqueurs microsatellites se sont révélés être des outils efficaces pour estimer le degré de parenté des géniteurs quand leur pedigree n'est pas connu et leur utilisation devrait être considérée afin de minimiser les croisements consanguins et la diminution rapide de la variabilité génétique. Des familles à fort et à faible potentiel de croissance ont été identifiées pour chacune des souches évaluées et une corrélation négative a été observée entre les performances de croissance des familles et le degré de parenté des géniteurs (estimé à l' aide de marqueurs microsatellites), les familles les plus proches génétiquement présentant des performances de croissance diminuées. Il a donc été suggéré d'utiliser les informations de parentés dans un programme de sélection puisqu' elles peuvent favoriser la sélection de familles performantes tout en évitant les effets de la consanguinité. Finalement les voies physiologiques impliquées dans le processus de croissance ont été investi guées. Les poissons présentant les taux de croissance les plus élevés avaient des capacités digestives et métaboliques plus importantes au niveau des caeca pyloriques et du muscle blanc respectivement. La sélection des individus les plus performants en ce qui a t~ait à la croissance favoriserait donc la sélection des individus présentant les meilleures capacités digestives et métaboliques

Sexing a sex-role-reversed species based on plumage: potential challenges in the red phalarope

Sex-role reversal, in which males care for offspring, can occur when mate competition is stronger between females than males. Secondary sex traits and mate attracting displays in sex-role-reversed species are usually more pronounced in females than in males. The red phalarope (Phalaropus fulicarius) is a textbook example of a sex-role-reversed species. It is generally agreed that males are responsible for all incubation and parental care duties, whereas females typically desert males after having completed a clutch and may pair with new males to lay additional clutches. The breeding plumage of female red phalaropes is usually more brightly colored than male plumage, a reversed sexual dichromatism usually associated with sex-role reversal. Here, we confirm with PCR-based sexing that male red phalaropes can exhibit both the red body plumage typical of a female and the incubation behavior typical of a male. Our result, combined with previous observations of brightly colored red phalaropes incubating nests at the same arctic location (Igloolik Island, Nunavut, Canada), suggests that plumage dichromatism alone may not be sufficient to distinguish males from females in this breeding population of red phalaropes. This stresses the need for more systematic genetic sexing combined with standardized description of intersexual differences in red phalarope plumages. Determining whether such female-like plumage on males is a result of phenotypic plasticity or genetic variation could contribute to further understanding sex-role reversal strategies in the short Arctic summer

Characterization of the early-stages of the wolffish hybrid

This is the first report of early-life developmental characteristics (coloration pattern, growth and survival) and genetic identification of the wolffish interspecific hybrid between Anarhichas minor and A. lupus, both endangered species in Canadian coastal water and of interest for cold-water aquaculture diversification. A first growth trial at 8 °C featuring pure strains A. minor and the hybrid A. minor × A. lupus in triplicates was conducted in 2006 during the period from 0 to 140 days post-hatch. A second growth trial was runned in 2007 featuring A. minor, A. lupus and the reciprocal hybrids A. minor × A. lupus and A. lupus × A. minor. Egg development indicators and early-hatching characteristics are reported