Disrupting PAR4 homodimers influences calcium mobilization in response to PAR4 agonist peptide. HEK293 cells were transfected with V5-PAR4-wt, V5-PAR4–2A, or V5-PAR4–4A in pCDNA3.1(1 μg). Forty-eight hours post-transfection the cells were removed from plates and loaded with Fura2-AM (A) or stained for flow cytometry (B) with an anti-V5 antibody. A, calcium mobilization was measured over time in response to the PAR4 agonist peptide (AYPGKF-NH2, 1 mm). Calcium concentration was determined as described under “Experimental Procedures.” B, cell surface expression of PAR4-wt, PAR4–2A, or PAR4–4A by flow cytometry. The mean fluorescence is from three independent experiments. The error bars indicate the standard deviation.