Fig. 4
Requirement of Kr-h1 phosphorylation in induction of RL36 transcription. A Relative levels of RL36 transcript in the fat body of 3-day-old adult females treated with dsKr-h1 vs. dsGFP control, NPC15437 (NPC) vs. DMSO solvent control (Cont.), and dsPKCα vs. dsGFP control. *P<0.05 and **P<0.01. n=8. B Luciferase reporter assays using S2 cells co-transfected with pGL4.10-4×RL36-1647 to -1632 plus pAc5.1/Flag-Kr-h1, pAc5.1/Flag-Kr-h1S154A, pAc5.1/Flag-Kr-h1S154D, or pAc5.1/Flag empty control. Methoprene was applied at 10 μM. Co-transfection of pGL4.10-4×RL36-1647 to -1632 and pAc5.1/Flag empty vector without methoprene treatment was used as the control. Means labeled with different letters indicate significant difference at P<0.05. n=4. C ChIP assays showing relative precipitation of RL36 promoter region with the KBS motif (RPRP-KBS) in the fat body of adult females on day 0 (A0), day 3 (A3), and day 6 (A6). D RPRP-KBS in the fat body of 3-day-old adult females treated with NPC15437 (NPC) vs. DMSO solvent control (Cont.) and dsPKCα vs. dsGFP control. E RPRP-KBS in the fat body of 3-day-old adult females treated with 50 μg methoprene vs. acetone solvent control. In C–E, p-Kr-h1, phospho-Kr-h1 (Ser154) antibody; Kr-h1, Kr-h1 antibody; and IgG, non-specific rabbit IgG control. Means labeled with different letters indicate significant difference at P<0.05. n=4