Phosphorylation of protein phosphatase inhibitor-1 by Cdk5.
Protein phosphatase inhibitor-1 is a prototypical mediator of cross-talk between protein kinases and protein phosphatases. Activation of cAMP-dependent protein kinase results in phosphorylation of inhibitor-1 at Thr-35, converting it into a potent inhibitor of protein phosphatase-1. Here we report that inhibitor-1 is phosphorylated in vitro at Ser-67 by the proline-directed kinases, Cdk1, Cdk5, and mitogen-activated protein kinase. By using phosphorylation state-specific antibodies and selective protein kinase inhibitors, Cdk5 was found to be the only kinase that phosphorylates inhibitor-1 at Ser-67 in intact striatal brain tissue. In vitro and in vivo studies indicated that phospho-Ser-67 inhibitor-1 was dephosphorylated by protein phosphatases-2A and -2B. The state of phosphorylation of inhibitor-1 at Ser-67 was dynamically regulated in striatal tissue by glutamate-dependent regulation of N-methyl-d-aspartic acid-type channels. Phosphorylation of Ser-67 did not convert inhibitor-1 into an inhibitor of protein phosphatase-1. However, inhibitor-1 phosphorylated at Ser-67 was a less efficient substrate for cAMP-dependent protein kinase. These results demonstrate regulation of a Cdk5-dependent phosphorylation site in inhibitor-1 and suggest a role for this site in modulating the amplitude of signal transduction events that involve cAMP-dependent protein kinase activation.
Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University, New York, New York 10021-6399, USA. bibbj@rockvax.rockefeller.edu
J A
JA
Bibb
A
A
Nishi
J P
JP
O'Callaghan
J
J
Ule
M
M
Lan
G L
GL
Snyder
A
A
Horiuchi
T
T
Saito
S
S
Hisanaga
A J
AJ
Czernik
A C
AC
Nairn
P
P
Greengard
D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000201:enzymology; D016203:CDC2 Protein Kinase / Q000378:metabolism; D019703:Calcineurin / Q000378:metabolism; D002352:Carrier Proteins; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D051360:Cyclin-Dependent Kinase 5; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D007700:Kinetics; D051379:Mice; D008810:Mice, Inbred C57BL; D016297:Mutagenesis, Site-Directed; D016202:N-Methylaspartate / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D011392:Proline / Q000378:metabolism; D054645:Protein Phosphatase 1; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D012694:Serine / Q000737:chemistry; D013997:Time Factors
D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; C021042:protein phosphatase inhibitor-1; D018698:Glutamic Acid; D012694:Serine; D016202:N-Methylaspartate; D011392:Proline; D000242:Cyclic AMP; D051360:Cyclin-Dependent Kinase 5; D017868:Cyclic AMP-Dependent Protein Kinases; D016203:CDC2 Protein Kinase; C495906:Cdk5 protein, mouse; C495907:Cdk5 protein, rat; D018844:Cyclin-Dependent Kinases; D019703:Calcineurin; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1