Activated Ras expression in eye discs with altered levels of hsrω lncRNA causes JNK-induced Dilp8 secretion and reduces post-pupal ecdysone leading to early pupal death in Drosophila

We examined reasons for pupal death following expression of certain transgenes with predominantly eye-disc expressing GMR-GAL4 or sev-GAL4 drivers in Drosophila. GMR-GAL4 or sev-GAL4 driven expression of UAS-Ras1 transgene, producing activated Ras, resulted in early (~25-30 Hr pupation) and late pupal death, respectively. Co-expression of UAS-hsrω-RNAi transgene or EP3037 to down or up-regulate, respectively, hsrω lncRNAs with sev-GAL4>UASRas1 advanced death to 25-30 Hr after pupation. The normal post-pupation ecdysone surge was absent in 24 Hr old GMR-GAL4>UAS-Ras1 pupae or those co-expressing sevGAL4>UAS-Ras1 with hsrω-RNAi or EP3037. Interestingly, exogenous ecdysone substantially suppressed their early death when provided for 12 Hr, beginning at 8-9 Hr after pupation. Microarray, qRT-PCR and immunostaining data revealed significantly elevated levels of Dilp8 and several members of JNK pathway genes but down-regulation of some of the ecdysone biosynthesis pathway genes in early dying pupae but not so much at their late third instar stage. Altered hsrω transcript levels in activated Ras expression background triggered greater Dilp8 secretion in early pupae. The consequent reduction in post-pupal ecdysone leads to early pupal death. This study explains for the first time the cause of early pupal death following expression of certain transgenes in developing eyes, which are otherwise dispensable for survival, and highlights the global consequences of deregulated signalling in one tissue through downstream events in other tissues.