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Department of Anesthesiology

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Regulation of neurabin I interaction with protein phosphatase 1 by phosphorylation.

TitleRegulation of neurabin I interaction with protein phosphatase 1 by phosphorylation.
Publication TypeJournal Article
Year of Publication1999
AuthorsMcAvoy T, Allen PB, Obaishi H, Nakanishi H, Takai Y, Greengard P, Nairn AC, Hemmings HC
JournalBiochemistry
Volume38
Issue39
Pagination12943-9
Date Published1999 Sep 28
ISSN0006-2960
KeywordsAmino Acid Sequence, Animals, Brain, Cyclic AMP-Dependent Protein Kinases, Male, Microfilament Proteins, Molecular Sequence Data, Nerve Tissue Proteins, Peptide Fragments, Phosphoprotein Phosphatases, Phosphorylation, Precipitin Tests, Protein Phosphatase 1, Rats, Rats, Sprague-Dawley, Sequence Homology, Amino Acid
Abstract

Neurabin I is a brain-specific actin-binding protein. Here we show that neurabin I binds protein phosphatase 1 (PP1) and inhibits PP1 activity. Neurabin I interacted with PP1alpha in an overlay assay, in yeast two-hybrid interaction analysis, and in coprecipitation and co-immunoprecipitation experiments. Neurabin I also copurified with both the alpha and gamma isoforms of PP1. A glutathione S-transferase (GST)-neurabin I fusion protein (residues 318-661) containing the putative PP1 binding domain (residues 456-460) inhibited PP1 activity (K(i) = 2.7 +/- 1.2 nM). This fusion protein was also rapidly phosphorylated in vitro by PKA (K(m) = 6 microM) to a stoichiomtry of 1 mol/mol. The phosphorylated residue was identified as serine 461 by HPLC-MS analysis of a tryptic digest. Phosphorylation of GST-neurabin I (residues 318-661) by PKA significantly reduced its binding to PP1 by overlay and by glutathione-Sepharose coprecipitation assays. A 35-fold decrease in inhibitory potency was also observed using a S461E mutant, which mimics phosphorylation of S461. These findings identify a signaling mechanism involving the regulation of PP1 activity and localization mediated by the cAMP pathway.

Alternate JournalBiochemistry
PubMed ID10504266
Grant ListDA10044 / DA / NIDA NIH HHS / United States
MH40899 / MH / NIMH NIH HHS / United States
P01 DA010044 / DA / NIDA NIH HHS / United States