Maturation of Megakaryoblastic Cells Is Accompanied by Upregulation of G S a-L Subtype and Increased cAMP Accumulation

In platelets and megakaryoblasts G s , the trimeric G-protein that stimulates adenylyl cyclase, is present in a short, 45 kDa, and a long, 52 kDa isoform termed G s a-S and G s a-L, respectively. To assess the relative contribution of these isoforms in the cellular synthesis of cAMP, the ratio G s a-S/G s a-L was changed in the megakaryoblastic cell line DAMI by inducing cell maturation with recombinant human thrombopoietin (TPO) or the phorbol ester PMA. Flow cytometric analysis confirmed that this treatment induced a moderate (TPO) and extensive (PMA) increase in nuclear ploidy and expression of the glycoproteins-IIIa and -Ib. Northern blot analysis revealed down-regulation of total G S -mRNA after treatment of DAMI-cells with TPO and PMA. Western blot analysis showed significant (P