MAPK ERK Signaling Regulates the TGF-β1-Dependent Mosquito Response to Plasmodium falciparum
Figure 4
PD98059 inhibited TGF-β1-induced ERK phosphorylation and enhanced TGF-β1-induced NOS expression in the A. stephensi midgut.
(A) PD98059 inhibited ERK activation in mosquito midgut. Anopheles stephensi mosquitoes were fed on artificial bloodmeals supplemented with 2000 pg/ml TGF-β1 with or without 4 or 40 µM PD98059. At 20 min after completion of feeding, 100 midguts from each treatment group were dissected and prepared for immunoblot analysis. Protein lysate concentrations were measured by BCA assay; equal amounts of protein were loaded in each lane. The immunoblot is representative of 3 independent experiments. (B) Inhibition of ERK phosphorylation enhanced TGF-β1-induced NOS expression in vivo. Anopheles stephensi were fed on artificial bloodmeals supplemented with 40 µM PD98059 (PD), 2000 pg/ml TGF-β1, or 40 µM PD98059 and 2000 pg/ml TGF-β1 (TGF-β1+PD). A control group (BM) was provided an identical unsupplemented artificial bloodmeal. Previous studies confirmed that provision of small volumes of PBS and DMSO has no effect on AsNOS expression in vivo (not shown), so these treatments were not included here. AsNOS expression levels were analyzed from midguts (n = 15) collected at 24 h or 48 h after blood feeding from each treatment group. The data are represented as means±standard errors from 5 independent experiments for fold inductions compared with the PBS control. The data were analyzed by ANOVA for overall significance and Student's t-test was used for the pairwise comparison at 24 h.