(A–E) The CPA complex genetically interacts with TOR.
Clonal expression of TSC1 and TSC2 reduced
cell size and increased mCherry-ATG8a puncta in the larval fat body under fed
conditions (A). These effects were suppressed by depletion of
CPSF6 (B), CPSF160 (C), or
CstF64 (D). Fat body cells were stained with DAPI. Scale
bar, 20 μm. Quantification of the relative number of mCherry-ATG8a dots
per cell (One-Way ANOVA followed by Bonferroni’s post hoc test; data are
represented as mean ± SEM; ***P<0.001) (E).
(F–G) PI3K/AKT/TORC1 signaling regulates APA and AS of
Atg1 and Atg8a. RNA from larval fat body
of indicated transgenes was subjected to qPCR or RT-PCR to detect APA and AS
isoforms. One-Way ANOVA followed by Bonferroni’s post hoc test; data are
represented as mean ± SEM; *P<0.05,
**P<0.01. (H) Rapamycin enhances phosphorylation of CPSF6.
S2R+ cells transfected with plasmids as indicated were treated with or
without Rapamycin (20 nM) for 24 hrs and then subjected to immunoprecipitation
(IP), followed by immunoblotting (IB).