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Figure 2


Fig. 2. Inactivation of the autophagy genes bec-1, unc-51 and lgg-1 suppresses ion-channel-dependent neuronal degeneration in C. elegans. (A) Number of vacuolated (dying) touch receptor neurons at the late L1 larval stage per 100 mec-4(u231) mutant animals with reduced autophagy gene activity. bec-1(–); Ex[bec-1(+)]refers to bec-1 mutant genotype bearing a non-integrated, extrachromosomal array (see Materials and Methods). For homozygous bec-1 mutants, n=50, P<0.001; for lgg-1(RNAi) animals, n=200, P<0.001; for other genotypes, n=200; P<0.0001 (unpaired t-tests). (B) Percentage of pmec-4::GFP-positive touch receptor neurons at young adulthood in mec-4(u231) mutants with reduced autophagy gene activity. In contrast to panel A, this panel indicates survival rate (%) of cells. For bec-1 mutants, n=50, P<0.001; for lgg-1(RNAi) animals, n=285, P<0.001; for other genotypes, n=200, P<0.0001 (unpaired t-tests). As a control, the expression of mec-4::GFP was assayed in bec-1(ok691) and unc-51(e369) single mutants as well as in lgg-1(RNAi) animals. Number of GFP-positive cells: unc-51(e369) mutants, 5.38 (n=263); bec-1(ok691) mutants, 5.47 (n=37); lgg-1(RNAi) animals, 5.67 (n=168); wild-type animals, 5.6 (n=493). These data show that inactivation of bec-1 and unc-51 only slightly influenced the development of touch receptor neurons, whereas lgg-1 RNAi treatment did not affect it at all (it is notable that lgg-1 RNAi completely abolished the expression of an lgg-1::gfp reporter). (C) Expression of a translational fusion pmec-4::MEC-4::GFP reporter in a PLM cell in a wild-type (left) and in an affected arrested bec-1(RNAi) (right) larva. In the latter, the expression intensity and number of GFP punctae were comparable with the wild type. (D) Number of vacuolated PVC interneurons at the L1 stage per 100 deg-1(u506) animals with reduced autophagy gene activity. For homozygous bec-1 mutants, n=50, P<0.001; for lgg-1(RNAi) animals, n=300, P<0.01; for other genotypes, n=300, P<0.0001 (unpaired t-tests). (E) Number of vacuolated IL1 sensory neurons and PVC interneurons at the L1 stage per 100 deg-3(u662) mutants with reduced autophagy gene activity. For lgg-1(RNAi) animals, n=300, P<0.018; for other genotypes, n=300, P<0.0001; (unpaired t-tests). In RNAi experiments, control animals were fed with E. coli HT115 expressing the empty vector under RNAi-inducing conditions. Data are mean ± s.e.m.





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