Figure 2.

Digestion of the FSHR with Endo H and plasma membrane expression upon exposure of HEK-293 cells expressing the WT FSHR to the palmitoylation inhibitor 2-bromopalmytate. (A) Representative Western blot analysis of the WT and triple mutant FSHR before (control) and after treatment of protein extracts with Endo-H. A reduction in the intensity of the 75 KDa band but not in the 80 KDa band was observed as a result of Endo-H digestion; in (B) gels were uploaded with twice the amount of protein extracts from cells expressing the triple mutant FSHR, and anti-FSHR antibody-probed membranes were then overexposed to the radiographic film. (C) Representative immunoblot and densitometric analysis of the WT FSHR from cells treated with 50 μM 2-bromopalmitate or vehicle alone (DMSO, control). A reduction in the intensity of the band corresponding to the FSHR was found as disclosed by densitometric analysis (right panel). (m): mature form of FSHR, (i) immature form of FSHR. (D) The plasma membrane expression of the WT FSHR was also reduced as a result of the brief (6 h) exposure to the palmitoylation inhibitor, as revealed by receptor binding studies.^*p < 0.05 vs. control.