Calcineurin-independent regulation of plasma membrane Ca2+ ATPase-4 in the vascular smooth muscle cell cycle

• Published in: American Journal of Physiology: Cell Physiology 2003-07, Vol.285 (1), p.C88-C95
• Main Authors: Afroze, Talat, Yang, Li L, Wang, Changsen, Gros, Robert, Kalair, Waseem, Hoque, Abu N, Mungrue, Imran N, Zhu, Ziping, Husain, Mansoor
• Format: Article
• Language: English
• Subjects: Animals; Antibodies - pharmacology; Benzylamines - pharmacology; Calcineurin - metabolism; Calcineurin Inhibitors; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases - antagonists & inhibitors; Calcium-Calmodulin-Dependent Protein Kinases - metabolism; Calcium-Transporting ATPases - genetics; Calcium-Transporting ATPases - metabolism; Carrier Proteins - genetics; Cation Transport Proteins; Cell Line; Cell Membrane - enzymology; Cell Size - physiology; Chelating Agents - pharmacology; Egtazic Acid - analogs & derivatives; Egtazic Acid - pharmacology; Enzyme Inhibitors - pharmacology; G1 Phase - physiology; Gene Expression Regulation, Enzymologic - physiology; Mice; Mice, Inbred C57BL; Muscle, Smooth, Vascular - cytology; Muscle, Smooth, Vascular - enzymology; Oligopeptides - genetics; Phenotype; Plasma Membrane Calcium-Transporting ATPases; Proto-Oncogene Proteins c-myb - immunology; Proto-Oncogene Proteins c-myb - metabolism; S Phase - physiology; Sulfonamides - pharmacology; Transfection
• ISSN: 0363-6143; 1522-1563
• DOI: 10.1152/ajpcell.00518.2002

Heart and Stroke Richard Lewar Centre of Excellence, University of Toronto, Division of Cell and Molecular Biology, Toronto General Hospital Research Institute, and Department of Medicine, University of Toronto, Toronto, Ontario, Canada M5G-2C4 Submitted 7 November 2002 ; accepted in final form 17 March 2003 Calcineurin mediates repression of plasma membrane Ca 2 + -ATPase-4 (PMCA4) expression in neurons, whereas c-Myb is known to repress PMCA1 expression in vascular smooth muscle cells (VSMC). Here, we describe a novel mouse VSMC line (MOVAS) in which 45 Ca efflux rates decreased 50%, fura 2-AM-based intracellular Ca 2 + concentrations ([Ca 2 + ] i ) increased twofold, and real-time RT-PCR and Western blot revealed a 40% decrease in PMCA4 expression levels from G 0 to G 1 /S in the cell cycle, where PMCA4 constituted 20% of total PMCA protein. Although calcineurin activity increased fivefold as MOVAS progressed from G 0 to G 1 /S, inhibition of this increase with either BAPTA or retroviral transduction with peptide inhibitors of calcineurin (CAIN), or its downstream target nuclear factor of activated T cells (NFAT) (VIVIT), had no effect on the repression of PMCA4 mRNA expression at G 1 /S. By contrast, Ca 2 + -independent activity of the calmodulin-dependent protein kinase-II (CaMK-II) increased eightfold as MOVAS progressed from G 0 to G 1 /S, and treatment with an inhibitor of CaMK-II (KN-93) or transduction of a c-Myb-neutralizing antibody significantly alleviated the G 1 /S-associated repression of PMCA4. These data show that G 1 /S-specific PMCA4 repression in proliferating VSMC is brought about by c-Myb and CaMK-II and that calcineurin may regulate cell cycle-associated [Ca 2 + ] i through alternate targets. calcineurin; c-Myb; plasma membrane Ca 2 + -ATPase-4; cell cycle Address for reprint requests and other correspondence: M. Husain, Div. of Cell and Molecular Biology, Toronto General Hospital Research Institute, Dept. of Medicine, Univ. of Toronto, EN12-221, 200 Elizabeth St., Toronto, Ontario, Canada M5G 2C4 (E-mail: mansoor.husain{at}utoronto.ca ).