losartan-potassium and herbimycin

To investigate the role of protein phosphorylation in the early phase of EPO-mediated signal transduction, we EPO-stimulated a murine erythroid cell line ELM-I-1 transformed by plasmids comprised of the c-fos enhancer/promoter linked to the luciferase gene. Using this reporter gene system, we previously showed that EPO-induced activation of the c-fos promoter can be detected rapidly and sensitively as an elevation of cellular luciferase activity. In this study, we first examined the role of protein tyrosine phosphorylation. The tyrosine phosphatase inhibitor orthovanadate not only induced luciferase activity by itself but enhanced the action of EPO. On the other hand, the tyrosine kinase inhibitors erbstatin and herbimycin suppressed the effect of EPO. Next, the role of protein kinase C (PKC) in the EPO response was assessed. The PKC activator phorbol myristate acetate (PMA) not only induced luciferase activity by itself but enhanced the action of Epo. On the other hand, the PKC inhibitor 1-(5-isoquinolynyl-sulfonyl)-2-methylpiperazine (H7) suppressed the effect of Epo and PMA, whereas a nonspecific protein kinase inhibitor, N-(2-Guanidinoethyl)-5-Isoquinolinesulfornamine (HA1004) inhibited the action of neither Epo nor PMA. Another known PKC inhibitor staurosporine (STSP) did not inhibit but rather enhanced the effect of Epo. This action of STSP was blocked by H7 but not by HA1004. These results suggest that the EPO-mediated early signal transduction pathway leading to c-fos expression involves protein-tyrosine phosphorylation, is modulated by tyrosine phosphatase activity and is positively regulated by PKC.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Alkaloids; Animals; Benzoquinones; Cell Line; Erythropoietin; Genes, fos; Hydroquinones; In Vitro Techniques; Isoquinolines; Lactams, Macrocyclic; Luciferases; Mice; Phosphoproteins; Piperazines; Promoter Regions, Genetic; Protein Kinase C; Protein-Tyrosine Kinases; Quinones; Rifabutin; Signal Transduction; Staurosporine; Vanadates

Erythropoietin (epo) appears to play a significant role in influencing the proliferation and differentiation of erythroid progenitor (CFU-E) cells. To determine the mechanism of action of epo, the effect of drugs on the in vitro colony formation of CFU-E cells induced from a novel murine erythroleukemia cell line, TSA8, was examined. While cytosine arabinoside inhibited colony formation and terminal differentiation of the CFU-E cells responding to epo, herbimycin, which is a drug that inhibits src-related phosphorylation, inhibited colony formation only. The same effect of herbimycin was observed with normal CFU-E cells from mouse fetal liver cells. These results suggest that epo induces two signals, one for proliferation and the other for differentiation, and that the two signals are not linked in erythroid progenitor cells.

Topics: Animals; Anti-Bacterial Agents; Benzoquinones; Cell Differentiation; Cell Division; Cells, Cultured; Erythropoietin; Hematopoietic Stem Cells; Lactams, Macrocyclic; Leukemia, Erythroblastic, Acute; Liver; Mice; Quinones; Rifabutin