EXPRESSION OF CA2-DEPENDENT PROTEIN-KINASE TYPE-II AND TYPE-IV, AND REDUCED DNA-SYNTHESIS DUE TO THE CA2+ CALMODULIN-DEPENDENT PROTEIN-KINASE INHIBITOR KN-62 ESULFONYL)-N-METHYL-L-TYROSYL]-4-PHENYLPIPERAZINE) IN SMALL-CELL LUNG-CARCINOMA( CALMODULIN)

Because changes in intracellular Ca2+ affect progression through the m itotic cell cycle, we investigated the role of Ca2+-binding proteins i n regulating cell cycle progression. Evidence was found demonstrating that the inactivation of Ca2+/calmodulin-dependent protein kinase (CaM kinase) inhibits cell cycle progression in small cell lung carcinoma (SCLC) cells. We also demonstrated that SCLC cells express both CaM ki nase type II (CaMKII) and CaM kinase type IV (CaMKIV). Five independen t SCLC cell lines expressed proteins reactive with antibody to the CaM KII beta subunit, but none expressed detectable proteins reactive with antibody to the CaMKII alpha subunit. All SCLC cells lines tested exp ressed both the alpha and beta isoforms of CaMKIV. Immunoprecipitation of CaMKII from SCLC cells yielded multiple proteins that autophosphor ylated in the presence of Ca2+/calmodulin. Autophosphorylation was inh ibited by the CaMKII(281-302) peptide, which corresponds to the CaMKII autoinhibitory domain, and by nesulfonyl)-N-methyl-L-tyrosyl]-4-pheny lpiperazine (KN-62), a specific CaM kinase antagonist. Influx of Ca2through voltage-gated Ca2+ channels stimulated phosphorylation of CaMK II in SCLC cells, and this was inhibited by KN-62. Incubation of SCLC cells with KN-62 potently inhibited DNA synthesis, and slowed progress ion through S phase. Similar anti-proliferative effects of KN-62 occur red in SK-N-SH human neuroblastoma cells, which express both CaMKII an d CaMKIV, and in K562 human chronic myelogenous leukemia cells, which express CaMKII but not CaMKIV. The expression of both CaMKII and CaMKI V by SCLC cells, and the sensitivity of these cells to the anti-prolif erative effects of KN-62, suggest a role for CaM kinase in regulating SCLC proliferation.