Figure 7. Atg9 interacts with Patj to regulate midgut cell growth.

(A) Lysates of cells expressing Myc-Patj were incubated with GST or GST–Atg9-C (amino acids 668–845) in GST pull-down assays. The pull-down products and input Myc-Patj were analyzed by Western blots with the anti-Myc antibody. (B–C) HEK293T cells transfected with Flag-Atg9 and Myc-Patj were subjected to immunoprecipitation with anti-Flag (B) or anti-Myc (C) antibody. The immunoprecipitates and total cell lysates (TCL) were analyzed by Western blot with antibodies as indicated. (D) S2 cells transfected with pWA-Gal4, pUAS-Flag-TSC2 and pUAS-Myc-Patj were subjected to immunoprecipitation with anti-Flag or anti-Myc antibody. The immunoprecipitates and total cell lysates were analyzed by Western blot with antibodies as indicated. (E) Depletion of Patj with NP1-Gal4 resulted in aberrant midgut epithelium and increased EC cell size. n ≥ 34, data are mean ±s.e.m. ***p<0.001. (F) Patj genetically interacts with Atg9. Overexpression of Patj suppressed the Atg9 depletion-induced midgut defects (60% penetrance, n = 24). The cell size of posterior midgut ECs of each genotype was quantified. n ≥ 20, data are mean ±s.e.m. ***p<0.001. Scale bar: 20 μm.

Figure 7—figure supplement 1. Atg9 genetically interacts with Patj.

(A) Overexpression Atg9 or Patj could not suppress the Atg1^RNAi-induced midgut defects. (B) Overexpression of Patj could not suppress the Atg17^RNAi-induced midgut defects. The cell size of posterior midgut ECs of each genotype was quantified. n ≥ 12, data are mean ±s.e.m. ns, not statistically significant. Scale bar: 20 µm. (C) Overexpression of Patj rescued the paraquat-induced lethality of Atg9 knockdown animals, compared with controls. (Long-rank test, p<0.001).