Figure 8. There is no involvement of arrestins in HCA₁-mediated ERK1/2 activation.

a, HEK-293 cells stably expressing HCA₁ were transfected with specific arrestin siRNA or a nonspecific control siRNA, 72 hrs after transfection, cells were harvested, and equal amounts of total cellular lysate were separated by 10% SDS-PAGE, transferred to a PVDF membrane, and incubated with anti-arrestin2/3 antibody. b, HEK-293 cells stably expressing HCA₁-EGFP were transfected with specific arrestin siRNA or a non-specific control siRNA,72 hrs after transfection, cells were stimulated with 20 mM lactate for 60 min and examined with confocal microscopy as described under ‘Experimental Procedures.’ c, 72 hrs after transfection with specific arrestin siRNA or non-specific control siRNA, cells were stimulated with 20 mM lactate for the indicated time periods and immunoblotted using monoclonal anti-phospho-MAPK E10 (Thr202/Tyr204), and then the blots were stripped and reprobed for total ERK1/2 to control for loading. The data and pictures shown are representative of at least three independent experiments. Error bars, S.E. for three replicates. Data were analysed by Student’s t test (***P<0.001).