Document Detail

Cell-cycle-dependent association of protein phosphatase 1 and focal adhesion kinase.
MedLine Citation:
PMID:  11513739     Owner:  NLM     Status:  MEDLINE    
Immunofluorescence studies with protein phosphatase-1 (PP1) isoforms-specific antibodies detected PP1delta, but not alpha or gamma1, at focal adhesions. PP1delta also co-immunoprecipitated with the focal adhesion kinase (FAK) and the alphav-integrin. In the present study glutathione S-transferase (GST)-PP1delta pulled-down FAK from fibroblasts extract and the interaction domain localized between residues 159 and 295 of delta. The association was confirmed by the ability to GST-FAK-related non-kinase (FRNK) to pull-down PP1delta from fibroblasts extract. GST-FRNK also pulled-down purified muscle PP1 catalytic subunit, thus indicating direct interaction between FAK and PP1. FAK displays consensus sequences for phosphorylation by cell division cycle kinase-2-cyclin B, and might be a PP1 substrate. In fact, FAK immunoprecipitated from metabolically-labelled mitotic HeLa cells without tyrosine phosphatase inhibitors was phosphorylated on Ser only and was dephosphorylated in vitro by purified muscle PP1, with loss of phospho-Ser. No PP1 was associated with FAK immunoprecipitated from mitotic HeLa cells. However, progressively more PP1 activity was assayed in FAK-immunoprecipitates obtained from cells released from mitosis. The associated activity was maximal at 2 h from the mitotic release (when 85-90% of the cells remained round) and decreased to basal level by 8 h (when cells were all polygonal). At the same time FAK underwent dephosphorylation, which was completed by 4 h. FAK obtained from cells at 1.5 h was Ser-phosphorylated, and underwent dephosphorylation during in vitro incubation, with loss of phospho-Ser, indicating the presence of active FAK-bound phosphatase. The only FAK-associated PP1 isoform between 1 and 8 h was PP1delta. The results suggest that FAK dephosphorylation by PP1delta occurs in cells released from mitosis, and confirmed the specific association of PP1delta, as detected previously in adherent cells.
M Fresu; M Bianchi; J T Parsons; E Villa-Moruzzi
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  The Biochemical journal     Volume:  358     ISSN:  0264-6021     ISO Abbreviation:  Biochem. J.     Publication Date:  2001 Sep 
Date Detail:
Created Date:  2001-08-21     Completed Date:  2001-10-04     Revised Date:  2012-06-05    
Medline Journal Info:
Nlm Unique ID:  2984726R     Medline TA:  Biochem J     Country:  England    
Other Details:
Languages:  eng     Pagination:  407-14     Citation Subset:  IM    
Department of Experimental Pathology, University of Pisa, Via Roma 55 56126 Pisa, Italy.
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MeSH Terms
Cell Cycle
Cells, Cultured
Focal Adhesion Kinase 1
Focal Adhesion Protein-Tyrosine Kinases
Glutathione Transferase / genetics
HeLa Cells
Muscle, Skeletal / enzymology
Phosphoprotein Phosphatases / genetics,  metabolism*
Protein Phosphatase 1
Protein-Tyrosine Kinases / genetics,  metabolism*
Rats, Inbred F344
Recombinant Fusion Proteins / metabolism
Grant Support
Reg. No./Substance:
0/Recombinant Fusion Proteins; EC Transferase; EC 2.7.1.-/FAK-related nonkinase; EC Adhesion Kinase 1; EC Kinases; EC Adhesion Protein-Tyrosine Kinases; EC protein, human; EC protein, rat; EC Phosphatases; EC Phosphatase 1

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