Figure 5.

Loop-mediated amplification of Hid levels after a pulse of hid. (a) Control en-Gal4>UAS-GFP; hs-hid flies. The expression of GFP is shown in blue. The 30 min heat shock induces high levels of Caspase-3 (green) and Hid (red), both in the anterior and posterior compartments. (b) en-Gal4>UAS-GFP UAS-dronc RNA-i; hs-hid disc showing a large reduction of Caspase-3 and Hid activity in the posterior compartment because of the suppression of dronc function. The third panel clearly indicates that most of the Hid protein visible after the heat shock is generated by the feedback loop. (c) A en-Gal4> UAS-GFP UAS-dp53 RNA-i; hs-hid disc showing reduction of Caspase-3 and Hid in the posterior compartment, where dp53 function is diminished. (d) en-Gal4> UAS-GFP UAS-puc; hs-hid disc with reduced levels of Caspase-3 and Hid in the posterior compartment. (e) The results of quantitative measurements of Caspase-3 and Hid activities in the genotypes studied. The percentage of the area of each compartment covered by the staining with anti-Caspase-3 or anti-Hid was calculated as indicated in the Materials and Methods section. There is no statistically significant difference between the values in UAS-GFP-expressing discs (n=23, P>0.05 both for anti-Caspase-3 and anti-Hid). A statistically significant reduction in the values of the posterior compartment is observed when UAS-dronc-RNA-i (n=19, P<0.0001 for both markers), UAS-dp53-RNA-i (n=20, P<0.01 for anti-Caspase-3 and P<0.001 for anti-Hid) or UAS-puc (n=27, P<0.01 for anti-Caspase-3 and P<0.001 for anti-Hid) are crossed to en-Gal4 UAS-GFP flies. ^*P<0.01, ^**P<0.001, ^***P<0.0001