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. 2014 Oct 23;10(10):e1004702. doi: 10.1371/journal.pgen.1004702

Figure 4. Responsiveness of Mcm4 and Mcm7 to JH and Met RNAi.

Figure 4

(A) Relative mRNA levels of Mcm4 and Mcm7 in the fat body of adult females treated with precocene (P10d) and those further treated with methoprene for 6, 12, 24 and 48 h (PM6h, PM12h, PM24h and PM48h, respectively). mRNA levels in precocene-treated fat bodies were used as the calibrator. *, P<0.05, **, P<0.01 and ***, P<0.001 compared to P10d; n = 4–6. (B) Mcm4 and Mcm7 mRNA abundance in fat bodies of female locusts from 0 to 10 days post adult eclosion (PAE). *, P<0.05 and **, P<0.01 compared to PAE0; n = 4–6. (C) Relative levels of Mcm4 and Mcm7 transcripts in fat bodies of dsGFP control (iGFP), Met-RNAi (iMet), and iMet further treated with methoprene for 48 h (iMet+JHA). *, P<0.05; n = 10–12. (D) Alignment of DNA element sequences containing E-box and E-box-like motifs in the upstream promoter regions of locust Mcm7 and Mcm4 with experimentally tested Met-binding consensus sequences except that of DmKr-h1 [7], [9], [34]–[36]. (E) EMSA using the Mcm4 probe listed in (D) with fat body nuclear extracts of iGFP and iMet (shown is a representative short exposure; longer exposures in some experiments showed additional two bands of less interest; see Results and Fig. S7). (F) EMSA using Mcm7 probe listed in (D) and with fat body nuclear extracts of iGFP and iMet. For both (E) and (F), arrows indicate the most likely specific bands. FP, free probe.