SummaryThrombopoietin (Tpo) regulates platelet production, but the mechanisms regulating the serum Tpo level and platelet count in circulation have been a subject of debate. Tpo was reported to be expressed mainly in liver and kidney, but we found that Tpo is expressed in all tissues examined: abundantly in liver, kidney, muscle, colon, brain and intestine, and moderately in bone marrow, spleen, lung, stomach, heart, thymus, ovary, and endothelial and leukemic cell lines. The levels of Tpo transcripts in major Tpo producing organs, liver and kidney, and in the platelet production sites bone marrow and spleen, were constant during acute thrombocytopenia induced by anti-platelet monoclonal antibody administration in mice, and during thrombocytosis induced by Tpo injection. Furthermore, we noticed that platelet count is not exactly inversely proportional to serum Tpo level. During acute thrombocytopenia, serum Tpo level transiently increased a few hours after antibody injection, and returned to the basal level just when matured megakaryocytes accumulated in bone marrow and spleen but the platelet count was still low. Matured megakaryocytes in bone marrow and spleen increased when the serum Tpo level decreased, and decreased when platelet count rebounded. Taken together with other observations, we propose here a modified version of Kuter and Rosenberg’s theory, that is, Tpo is constitutively expressed in a variety of organs throughout the body, even in acute thrombocytopenia and thrombocytosis, and that the serum Tpo level is not regulated by Tpo gene expression nor only by platelet counts in circulation, but by the total counts of both megakaryocytes in bone marrow and spleen and of platelets in circulation

The regulator of G-protein signaling (RGS) negatively regulates the ␣ subunit of G proteins by accelerating their intrinsic guanosine triphosphatase (GTPase) activity. Here are reported the isolation and characterization of a novel mouse RGS, termed RGS18, which is a new member of RGS subfamily B. Northern blot analysis showed that RGS18 messenger RNA was detected predominantly in spleen and hematopoietic cells, and immunohistochemical studies demonstrated that RGS18 was expressed in megakaryocytes, platelets, granulocytes/monocytes, and, weakly, in hematopoietic stem cells, but not in lymphocytes or erythrocytes. Although various subcellular localizations of RGS have been reported, RGS18 was found to be localized in cytoplasm in megakaryocytes. In vitro binding assays of RGS18 with megakaryocyte cell lysates with or without AlF 4 ؊ treatment demonstrated that RGS18 specifically binds to 2 ␣ subunits of the G protein, G␣i and G␣q. Furthermore, RGS18 clearly exhibited GTPase-activating protein (GAP) activity for G␣i and G␣q but not for G␣s or G␣12. In addition, chemokine stromal-derived factor 1 (SDF-1), which has been reported to stimulate megakaryocyte colony formation in the presence of thrombopoietin, affected the binding of RGS18 to G␣i but not to G␣q. Therefore, the newly isolated RGS18 turned out to be a new member of the RGS family bearing GAP activity for G␣i, which might be stimulated by SDF-1 in megakaryocytes, as well as for G␣q. Thus, RGS18 may play an important role in proliferation, differentiation, and/or migration of megakaryocytes. IntroductionThe guanosine triphosphate (GTP)-binding protein (G-protein) signaling pathway is one of the most important signaling cascades used to relay extracellular signals and sensory stimuli to eukaryotic cells. 1 Heterotrimeric G proteins, which couple heptahelical receptors to effectors in the signal transduction pathway, are composed of ␣, ␤, and ␥ subunits. 2,3 Each subunit has multiple isoforms; the ␣ subunit isoforms are grouped into 4 subfamilies: G␣i, G␣s, G␣q, and G␣12. 4 The guanosine diphosphate (GDP)-bound heterotrimer is an inactive form of the G protein. Ligand-bound activated receptors catalyze the exchange of GDP by GTP on the ␣ subunit, leading to dissociation of the ␣ from the ␤␥ dimer and signal transduction by the separated G-protein subunits (G␣ and G␤␥). The large body of research examining G-protein-mediated signaling pathways has focused mainly on the involvement of receptor phosphorylation and sequestration in the desensitization process.Recently, a new family of proteins, the regulator of the G-protein-signaling (RGS) family, has been identified. 5-7 Genetic screenings for negative regulators for the pheromone response pathway in yeast identified a protein, Sst2. 8 Further analyses revealed that Sst2 interacted directly with the G-protein ␣ subunit. 9 To date, at least 26 different RGS proteins have been described in mammals. They all contain a conserved, characteristic domain (termed RGS domain) that interacts specifically with the ac...