Starvation-sensitivity assays define the range of autophagy defects in vap flies.

The survival rate of 3 day-old adult males of indicated genotypes was recorded at 25°C in condition of complete food deprivation (see S4A–S4C Fig for initial characterization). (A) The vap-dependent starvation sensitivity (white arrow) was compared to weak (Atg8a1) and strong (Agt8a2) alleles of Atg8a. Atg8a2 flies showed slightly altered development that might contribute to its greater sensitivity to starvation. (B-B’) Starvation sensitivity effect, as assayed at 25°C, is partially recapitulated by flies that were ectopically expressing an UAS-myc:Atg1 transgene (Materials and Methods) along fat cell development performed at 25°C (white arrow in B) when driven by cg-Gal4. As a control, there is no detectable starvation sensitivity (as assayed at 25°C) using identical flies (UAS-myc:Atg1 /cg-Gal4) that developed at 18°C to minimized transgene expression (white arrow in B’). Ectopic expression of Atg1 during development is therefore responsible for the sensitivity effect found in B. (C) The vap-dependent starvation sensitivity is suppressed (white arrow) by co-expressed Atg5(RI) using the broadly expressed arm-Gal4 driver. Genotypes. (A) Control: w1118/Y. Assay vap1/Y. Atg8a1/Y. Atg8a2/Y. (B, B’) Control: UAS-myc:Atg1/+ and vap1/Y and vap1/Y; cg-GAL4/+. Assay: vap1/Y. cg-GAL4/ UAS-myc:Atg1(RI)/+. (C) Control: arm-GAL4/+ and vap1/Y; arm-GAL4/+ and arm-GAL4/ UAS-Atg5(RI)/+. Assay: vap1/Y; arm-GAL4/ UAS-Atg5(RI)/+.