Figure 3From: Myotubularin family phosphatase ceMTM3 is required for muscle maintenance by preventing excessive autophagy in Caenorhabditis elegansKnockdown of ceMTM3 causes autophagy in multiples tissues of C. elegans worms at different ages. GFP: LGG-1 worms were cultured on NGM plates containing E. coli cells carrying vector control or ceMTM3 RNAi from the time they were hatched from eggs. GFP-positive puncta (indicated by arrows) represent autophagosomes which are abundant in ceMTM3 RNAi-treated worms. A. Autophagosomes in seam cells of day 2 worm larvae. B. Autophagosomes in muscle cells of day 3 young adult worms. Note that muscle fiber bands are visible in the background. C. Autophagosomes in intestine on day 5 adult worms. D. Average numbers of GFP puncta in 4 seam cells, 60 micrometer-long muscles, and entire intestine. Error bars denote standard deviation, n≥20, p<0.001). E. Western blotting analyses of lipid-bound GFP-LGG-1. Control and ceMTM3 RNAi-treated worms (mixed ages) were extracted in phosphate-buffer saline by sonication. After centrifugation at 800 g for 10 min, the supernatants were collected and subjected to further centrifugation at 100,000g for 1 h. The pellets were extracted in SDS gel sample buffer and directly used for western blotting analysis with an anti-GFP antibody. The position of GFP-LGG-1 is indicated. Equal protein loading is demonstrated by Coomassie blue staining.Back to article page