Hindguts from experimental and control larvae, identified by the absence of the Tubby (Tb) marker, were dissected and processed as previously described (30 (link)). Dissections were performed in 1X PBS, dissected tissue fixed with 4% paraformaldehyde, and processed for immunohistochemistry using our standard protocols(28 (link), 30 (link)). The following primary antibodies were used: mouse anti-NimC1 (79 (link))(Gifted by Eva Kurucz, 1:30), mouse anti-Mmp1(1:1000, DSHB #3B8D12), mouse anti-phospho JNK (1:50, Cell Signaling Technology #9255), mouse anti-phospho Akt (1:1000, Cell Signaling Technology #4054), mouse anti-Histone 2A gamma variant, phosphorylated (1:100, DSHB #UNC93–5.2.1), rabbit anti-Histone H3 (tri methyl K9) (1:50, Abcam #ab8898), mouse anti-Dacapo (1:10, DSHB #NP1), rabbit anti-Dcp1(1:1000, Cell Signaling Technology #9578), mouse anti-β-gal (1:100, DSHB #40–1a), rabbit anti-β-gal (1:200, Thermo Fisher Scientific #A111–32 ). Alexa-conjugated goat-anti-mouse and anti-rabbit antibodies were used as secondary antibodies (1:1000, Thermo Fisher Scientific, #A-11031, #A-21052, #A-110356, #A-21071). All guts were imaged using SPE DM6 Leica Confocal Microscope at 40X magnification at 1.0 Zoom. Hindguts used to quantify the imaginal ring proliferation area were imaged at 10X magnification, 1.5X Zoom (30 (link)).