CHRM3 Antibody - #DF2781

Fig. 5. Interference with CHRM3 protein inhibited the activation of the pathway caused by P1 virus. (A)Schematic diagram of shRNA and sequencing results. Red and green boxes indicate the interference sites of shRNA-1. (B)Real-time PCR analysis of CHRM3 mRNA expression in 3D4 cells transfected with shRNA-N (empty vector control), shRNA-1, or shRNA-2 at 24 h and 48 h post-transfection (n = 3). Immunoblot analysis of CHRM3 protein in CHRM3 knockdown 3D4 cells. The right panel is the intensity of the CHRM3 protein level normalized to that of β-actin (n = 3). The 3D4 cells were transfected with shRNA-N or shRNA-1 for 48 h and then infected with P1 (shRNA-1 +P1) or uninfected (shRNA-N or shRNA-1). Real-time PCR (D) and Western blot (E) were used to detect the expression of the main components of the pancreatic secretory pathway (CHRM3, Gq, PLC-β2, PRKCA, Rab3D, RhoA) at 48 h post-infection (n = 3). The gray value of the Western blot band (F) was analyzed by ImageJ software. Student's t-test was used to examine the differences between groups (∗P 

Fig. 3. Both the VP1 protein of the virus and the CHRM3 protein upregulate main components of the pancreatic secretory pathway. (A) Amplification of VP1 and CHRM3 genes and restriction digestion identification of recombinant plasmids. Western blot (B) and Immunofluorescence (C) analysis of the expression of VP1 and CHRM3 protein in HEK293 cells transfected with pcDNA3.1-HA-CHRM3 or pcDNA3.1-VP1-Flag at 48 h post-transfection. (D) Western blot analysis of the expression of VP1 and CHRM3 proteins in 3D4 cells transfected with pcDNA3.1-HA-CHRM3 or pcDNA3.1-VP1-Flag at 24 h, 48 h, and 72 h post-transfection. The right panel is the intensity of the CHRM3 protein level normalized to that of β-actin (n = 3). (E) Real-time PCR analysis of CHRM3, Gq, PLC-β2, PRKCA, Rab3D, RhoA, Rac1, and amyA mRNA expression levels in 3D4 cells transfected with pcDNA3.1-VP1-Flag, pcDNA3.1-HA-CHRM3, or pcDNA 3.1 vector (control) at 24 h, 48 h, or 72 h post-transfection (n = 3). The differences between groups were analyzed by Student's t-test (∗P