Following subcellular fractionation of platelet homogenates and Western blotting, two groups of alpha-subunits of trimeric G-proteins could be distinguished. Group 1 consisted of alpha(i)-2, alpha(i)-3, alpha(z), alpha(s), and alpha(q) and was predominantly localized in membranes. Group 2 consisted of alpha(16) and alpha(12) and was predominantly localized in the cytosol. Plasma membranes and dense tubular system (DTS)-membranes showed the same distribution of Group 1 alpha-subunits. An exception was alpha(q), which was virtually absent in the DTS as were Group 2 subunits. In addition, this compartment showed a doublet for alpha(z). Group 1 alpha-subunits were also found in fractions with the combined secretory granules and in separate dense granules. In addition, alpha(16) was found in these granule fractions, but secretion granules were devoid of alpha(12). These data reveal a heterogeneous distribution of alpha-subunits in platelet compartments and may indicate that G(12) and G(16) play different roles in platelets than members of the G(i) and G(s) classes and other members of the G(q) class. (C) 1997 Academic Press.