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PMID: 10330411 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Paxillin LD4 motif binds PAK and PIX through a novel 95-kD ankyrin repeat, ARF-GAP protein: A role in cytoskeletal remodeling.

The Journal of cell biology ·Vol. 145 ·No. 4 ·1999-05-17 ·Pages 851-63

Turner CE, Brown MC, Perrotta JA, Riedy MC, Nikolopoulos SN, McDonald AR, Bagrodia S, Thomas S, Leventhal PS

Abstract

Paxillin is a focal adhesion adaptor protein involved in the integration of growth factor- and adhesion-mediated signal transduction pathways. Repeats of a leucine-rich sequence named paxillin LD motifs (Brown M.C., M.S. Curtis, and C.E. Turner. 1998. Nature Struct. Biol. 5:677-678) have been implicated in paxillin binding to focal adhesion kinase (FAK) and vinculin. Here we demonstrate that the individual paxillin LD motifs function as discrete and selective protein binding interfaces. A novel scaffolding function is described for paxillin LD4 in the binding of a complex of proteins containing active p21 GTPase-activated kinase (PAK), Nck, and the guanine nucleotide exchange factor, PIX. The association of this complex with paxillin is mediated by a new 95-kD protein, p95PKL (paxillin-kinase linker), which binds directly to paxillin LD4 and PIX. This protein complex also binds to Hic-5, suggesting a conservation of LD function across the paxillin superfamily. Cloning of p95PKL revealed a multidomain protein containing an NH2-terminal ARF-GAP domain, three ankyrin-like repeats, a potential calcium-binding EF hand, calmodulin-binding IQ motifs, a myosin homology domain, and two paxillin-binding subdomains (PBS). Green fluorescent protein- (GFP-) tagged p95PKL localized to focal adhesions/complexes in CHO.K1 cells. Overexpression in neuroblastoma cells of a paxillin LD4 deletion mutant inhibited lamellipodia formation in response to insulin-like growth fac- tor-1. Microinjection of GST-LD4 into NIH3T3 cells significantly decreased cell migration into a wound. These data implicate paxillin as a mediator of p21 GTPase-regulated actin cytoskeletal reorganization through the recruitment to nascent focal adhesion structures of an active PAK/PIX complex potentially via interactions with p95PKL.

MeSH Terms
ADP-Ribosylation Factors Amino Acid Sequence Animals Ankyrins/genetics,metabolism Binding Sites CHO Cells COS Cells Carrier Proteins/genetics,metabolism,physiology Cell Adhesion Molecules/metabolism Cell Cycle Proteins/metabolism Cell Movement Cricetinae Cytoskeletal Proteins/metabolism Cytoskeleton/physiology DNA-Binding Proteins/metabolism Focal Adhesion Protein-Tyrosine Kinases GTP Phosphohydrolases GTP-Binding Proteins/metabolism GTPase-Activating Proteins Intracellular Signaling Peptides and Proteins LIM Domain Proteins Molecular Sequence Data Paxillin Phosphoproteins/metabolism Protein Serine-Threonine Kinases/metabolism Protein-Tyrosine Kinases/metabolism Proteins/metabolism Repetitive Sequences, Nucleic Acid Sequence Analysis, DNA Subcellular Fractions Vinculin/metabolism cdc42 GTP-Binding Protein p21-Activated Kinases
Chemicals
Ankyrins Carrier Proteins Cell Adhesion Molecules Cell Cycle Proteins Cytoskeletal Proteins DNA-Binding Proteins GTPase-Activating Proteins Intracellular Signaling Peptides and Proteins LIM Domain Proteins Paxillin Phosphoproteins Proteins TGFB1I1 protein, human Vinculin Protein-Tyrosine Kinases Focal Adhesion Protein-Tyrosine Kinases PAK1 protein, human PAK2 protein, human Protein Serine-Threonine Kinases p21-Activated Kinases GTP Phosphohydrolases GTP-Binding Proteins ADP-Ribosylation Factors cdc42 GTP-Binding Protein
Authors & Affiliations
9 authors, click to expand affiliations / ORCID
Turner C E
Department of Anatomy and Cell Biology, State University of New York, Health Science Center, Syracuse, New York 13210, USA. turnerc@vax.cs.hscsyr.edu
Brown M C
Perrotta J A
Riedy M C
Nikolopoulos S N
McDonald A R
Bagrodia S
Thomas S
Leventhal P S
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.145.4.851
Email
turnerc@vax.cs.hscsyr.edu
Published
1999-05-17
Pages
851-63
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2133183
Subset
IM
Grants
NIGMS NIH HHS · R01 GM047607 · United States
NIGMS NIH HHS · GM47607 · United States
Databases
GENBANK
AF112366
External Links
PubMed source DOI: 10.1083/jcb.145.4.851 PMC Full Text View journal details
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