Oscillation of Cyclin E and E2F target gene expression is deregulated in <i>de2f1b</i> mutant salivary glands.

<p>(A) Salivary glands of control and <i>de2f1b</i> mutant early third instar larvae (80–85 hr AEL) are stained with anti-dE2F1 (green) and anti-Cyclin E (CycE, red). Bar graph indicates quantification of the percentages of CycE and dE2F1 expressing cells that co-express dE2F1 and CycE, respectively, in control and <i>de2f1b</i> mutant salivary glands. **** = p≤ 0.0001 using two-tailed t-test. (B) Salivary glands of control and <i>de2f1b</i> mutant early (80–85 hr AEL) third instar larvae expressing PCNA-GFP (green) are stained with anti-dE2F1 (red). The region where high PCNA-GFP is observed with low dE2F1 is marked by an asterisk. (C) Salivary glands of control and <i>de2f1b</i> mutant late third instar larvae (105–110 hr AEL) are stained with anti-Cyclin E (CycE). In addition, dE2F1 activity is monitored using PCNA-GFP, which is a GFP reporter of a well-known E2F target gene, PCNA. Cytoplasmic accumulation of dE2F1 in <i>de2f1b</i> mutants is marked by an arrowhead. (D) Relative expression levels of E2F target genes between control and <i>de2f1b</i> mutant late (105–110 hr AEL) third instar larval salivary glands are determined by RT-qPCR. Values represent the mean of triplicated biological replicates and error bars represent s.d. * = p ≤ 0.05; ** = p ≤ 0.01; **** = p≤ 0.0001 using two-tailed t-test. (E) Averages of the relative standard deviation of nuclear area from control, <i>de2f1b</i> mutant, and <i>de2f1b</i> mutant knocking down <i>cycE</i> using <i>heatshock-Gal4</i> and <i>UAS-cycE</i>^<i>RNAi</i>. Values represent the mean of triplicated biological replicates and error bars represent s.d. P values represent: ns = p>0.05; *** = p≤0.001; **** = p≤0.0001 calculated using one-way ANOVA. Scale bars for all salivary glands represent 50 μm. P: proximal end. D: distal end. FB: Fat body.