Two Distinct Forms of MAPKAP Kinase-2 in Adult Cardiac Ventricular Myocytes
- 29 April 2000
- journal article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 39 (20), 6145-6156
- https://doi.org/10.1021/bi9928389
Abstract
Hsp27 kinase activities were studied in adult rat ventricular myocytes following sequential chromatography on Mono Q and Mono S. A basal level of activity was present following cell isolation. FPLC on Mono Q revealed three peaks of activity, peaks ‘a', ‘b', and ‘c'. A fourth peak, ‘d', was detected upon subsequent chromatography of the Mono Q flow-through on Mono S. Immunoblotting revealed that peaks ‘a', ‘b', and ‘c' contained predominantly a 49 kDa form of MAPKAP kinase-2. Peak ‘d' contained a 43 kDa form. ‘In-gel' kinase assays using hsp27 indicated both forms of MAPKAP kinase-2 were active. No other bands of hsp27 kinase activity were detected. Both forms of hsp27 kinase immunoprecipitated with a MAPKAP kinase-2 antibody and have therefore been named MAPKAP kinase-2α (p49) and MAPKAP kinase-2β (p43). MAPKAP kinase-2β chromatographed on Superose 12 as a 60.7 kDa monomer whereas the behavior of MAPKAP kinase-2α suggested both a 65.7 kDa monomer and higher molecular mass complexes. Both activities phosphorylated hsp27 on serine residues, and two-dimensional phosphopeptide mapping indicated the same sites were phosphorylated. A tumor-promoting phorbol ester, phorbol 12-myristate 13-acetate (PMA), stimulated both MAPKAP kinase-2α and MAPKAP kinase-2β activity. Inhibition of MEK activation with PD 98059 or p38α/β MAP kinase activity with SB203580 blocked activation by PMA. However, whereas PD 98059 inhibited only the PMA-stimulated activation, SB203580 inhibited both PMA-stimulated and basal hsp27 phosphorylation. These data demonstrate the presence of two forms of MAPKAP kinase-2 in adult ventricular myocytes. Both forms are activated indirectly by the ERK MAP kinase pathway and directly by p38 MAP kinase but independently regulated.Keywords
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