Figure 1.
Identification of a panel of v-ATPase subunits that are essential for the UPR
mt
activation in C. elegans. (A) RNAi of multiple v-ATPase subunits attenuated UPRmt activation induced by cco-1 RNAi in hsp-6p::gfp worms. For RNAi treatment, RNAi targeting v-ATPase subunits or atfs-1 occupied 60%, cco-1 RNAi occupied 40%, control RNAi (empty vector [ev]) was used to supply to a final 100% of RNAi for all conditions. DIC, differential interference contrast image. (B) The two functional domains of v-ATPase: the V1 domain, which hydrolyzes ATP to generate the energy required for pumping protons, and the membrane-anchoring V0 domain, which transports H+ across the lipid bilayer. Subunits whose RNAi suppressed the UPRmt were highlighted in red, and the best four of them in red bold. (C) RNAi of vha-1, vha-4, vha-16, and vha-19 (25%) attenuated mrps-5 RNAi-induced UPRmt activation in hsp-6p::gfp worms. (D)vha-1 RNAi inhibited UPRmt activation induced by cco-1 or mrps-5 (40%) RNAi in a dose-dependent manner. (E)vha-1 RNAi attenuated UPRmt activation induced by spg-7, cts-1 or dlst-1 RNAi, and by doxycycline (Dox; 30 μg/ml) or antimycin A (2.5 μM). (F) Schematic diagram showing the regions on mRNA targeted by the two different vha-1 RNAi obtained from either the Vidal (vha-1_RNAi_1) or Ahringer library (vha-1_RNAi_2). (G) Both vha-1 RNAi as indicated in F disrupted the UPRmt induced by either cco-1 or mrps-5 RNAi. Note the more robust effect of vha-1_RNAi_2 in UPRmt suppression as compared to that of vha-1_RNAi_1. (H) The lysosomal acidification inhibitor chloroquine (CQ) suppressed UPRmt activation in a dose-dependent manner. hsp-6p::gfp worms were fed with control or cco-1 (40%) RNAi, in combination with 2–8 mM CQ. Scale bars, 0.3 mm.