Cell volume regulation in response to hypotonicity is impaired in HeLa cells expressing a protein kinase C alpha mutant lacking kinase activity

Hermoso, M; Olivero P.; Torres R.; Riveros A.; Quest, AFG; Stutzin A.

Abstract

The chloride conductance (GCl,swell) that participates in the regulatory volume decrease process triggered by osmotic swelling in HeLa cells was impaired by removal of extracellular Ca2+, depletion of intracellular Ca2+ stores with thapsigargin, or by preloading the cells with BAPTA-AM (1,2-bis(2-aminophenoxy)ethane-N,N,N?,N? -tetraacetic acid). Furthermore, overnight exposure to the phorbol ester tetradecanoyl phorbol acetate and acute incubation with inhibitors of the conventional protein kinase C (PKC) isoforms bisindolylmaleimide I and Gö6976 inhibited GCl,swell. Treatment of HeLa cells with U73122, a phospholipase C inhibitor, also prevented GCl,swell. Hypotonicity induced selective PKC? accumulation in the membrane/cytoskeleton fraction in fractionation experiments and translocation of a green fluorescent protein-PKC? fusion protein to the plasma membrane of transiently transfected HeLa cells. To further explore the role of PKCs in hypotonicity-induced GCl,swell, HeLa clones stably expressing either a kinase-dead dominant negative variant of the Ca2+-dependent PKC isoform ? (PKC? K386R) or of the atypical PKC isoform ? (PKC? K275W) were generated. GCl,swell was significantly reduced in HeLa cells expressing the dominant negative PKC? mutant but remained unaltered in cells expressing dominant negative PKC?. These findings strongly implicate PKC? as a critical regulatory element that is required for efficient regulatory volume decrease in HeLa cells.

Más información

Título según WOS: Cell volume regulation in response to hypotonicity is impaired in HeLa cells expressing a protein kinase C alpha mutant lacking kinase activity
Título según SCOPUS: Cell Volume Regulation in Response to Hypotonicity Is Impaired in HeLa Cells Expressing a Protein Kinase C? Mutant Lacking Kinase Activity
Título de la Revista: JOURNAL OF BIOLOGICAL CHEMISTRY
Volumen: 279
Número: 17
Editorial: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Fecha de publicación: 2004
Página de inicio: 17681
Página final: 17689
Idioma: English
URL: http://www.jbc.org/cgi/doi/10.1074/jbc.M304506200
DOI:

10.1074/jbc.M304506200

Notas: ISI, SCOPUS