| Popis: |
As a strategy to survive to the upcoming winter, many insects enter diapause (a typical overwintering response that results on their developmental arrest). Drosophila melanogaster undergoes an adult or reproductive diapause that can be easily spotted by looking at the stage of development of the females’ ovaries. The possibility of the circadian clock influencing this phenotype was proposed to explain photoperiodic differences in induction levels. Nevertheless, to the date the debate is still on. In this thesis, I looked at several canonical clock mutants and assessed their impact on diapause, finding that 1) depending on the temperature in which they were reared the effects on the adult flies varied enormously 2) most of the clock mutants gave a strong effect in one or other growing conditions. In particular, Pdf0 and ClkJrk mutants behave in completely opposite ways. A second part of the project consisted on looking at the effects of period temperature-sensitive splicing in diapause. Using splicing locked transgenic flies provided by Isaac Edery, I found that expression of the summer isoform impaired the ability of the flies to undergo diapause. Hence, I cloned the different splicing variants into a pUAST vector and generated UAS lines to perform a neuroanatomical dissection of the phenotype. Also, related with the previous project, I decided to look if any miRNA could be regulating diapause by affecting any of the splicing variants. I found several possible miRNAs that could target the summer (intron-containing) non-splicing isoform. I found that one particular, miRNA-276b, was having a huge effect on diapause. Using a sponge particularly against this miRNA (which would result in its downregulation) diapause levels halved compared to all the controls that were performed in parallel. |
