Caenorhabditis elegans possui quatro genes de kpcs (kex2/subtilisin-like proprotein convertases): kpc-1, kpc-2/egl-3, kpc-3/aex-5, kpc-4/bli-4. Em C. elegans, dois dos quatro polipeptídeos de vitelogenina encontrados dentro dos ovócitos, YP115 e YP88, se originam a partir de um precursor polipeptídico (VIT-6) clivado pós-traducionalmente após o motivo RGKR. Nematóides transgênicos foram produzidos com construções repórteres transcricionais de GFP. Foi verificada expressão de kpc-1 tanto em neurônios quanto em células musculares e intestinais. Esses dados, aliados aos dados da literatura para os outros genes kpc de C. elegans, sugerem o envolvimento de KPC-1 no processamento de VIT-6, que é secretada por células intestinais. Ensaios de Western-blot compararam o processamento de VIT-6 em nematóides selvagens, mutantes e knock-down por RNAi para os diferentes genes kpc. A análise de nematóides mutantes e knock-down por RNAi combinado para os outros três genes de convertase de C. elegans confirmou a redundância da atividade dessas enzimas no processamento de VIT-6. / Four kpc genes are found in the Caenorhabditis elegans genome: (kex2/subtilisin-like proprotein convertases): kpc-1, kpc-2/egl-3, kpc-3/aex-5, kpc-4/bli-4. Two of the four vitellogenin polypeptides, YP115 and YP88, originate from a precursor, VIT-6. VIT-6 is cleaved post-translationally after the RGKR motif. Transgenic worms carrying GFP transcription reporter constructs were produced. Expression of kpc-1 has been localized to neurons as well as muscular and intestinal cells. These data, together with the ones available from the literature for the other kpc genes, suggest the involvement of KPC-1 in the processing of VIT-6, which is secreted from intestinal cells. Western-blot analysis compared the pattern of VIT-6 processing in wild-type, mutants and RNAi-treated worms for the other kpcs. Analysis of worms treated by combined RNAi confirmed the redundancy of KPCs in VIT-6 processing.

Tesis por compendio / European eel (Anguilla anguilla, L., 1758) is suffering a strong population decrease and at the same time it is a very appreciated species and by now it has not been possible closing its cycle life. In fact, this species does not mature in captivity unless hormonally induced. So all the production is up to the natural population. All these factors together make urgent achieving the closing of the productive cycle and for this aim it is important to understand the reproductive physiology and the reasons of this development blockage. The present thesis wants to be a new contribution to the knowledge of reproductive physiology in female European eel submitted at hormonal treatment. To achieve this goal, expression of genes not previously studied in this species (cyp19a1, ara, arb, gnrhr1a, gnrhr1b, gnrhr2, zpb and zpc) was analyzed in eels reared under a constant thermal regime, accordingly to the usual rearing conditions. Also, the effect of rearing temperature on gene expression and steroid profile (T, 11-KT and E2) was studied. In fact, eels migrate to Sargasso Sea to reproduce and during the travel experiment temperature changes, while traditionally they are reared at a constant high temperature which could affect vitellogenesis progression and final oocyte quality. For the study it was necessary cloning and characterizing some genes which have not still been sequenced in European eel. Gene expression was studied by qPCR after designing primer and optimizing the qPCR race. Steroid profiles were analyzed by immunoassays and the gonadal development stages were established by histology. The first result obtained at the end of the study were six new genes characterized in European eel. The analysis of gene expression allowed to understand the involvement of specific genes during vitellogenesis (arb, gnrhr1b and gnrhr2) in different brain regions. The temperature was conformed as a crucial environmental factor affecting vitellogenesis. On one hand, eels matured at lower starting temperatures showed better reproductive parameters which could have an influence in the final oocyte quality. On the other hand higher temperatures are necessary to achieve further vitellogenetic stages / Mazzeo, I. (2014). Effect of thermal regime on the expression of key reproductive genes during hormonally-induced vitellogenesis in female European eels [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/48490 / Compendio