Paxillin binding is not the sole determinant of focal adhesion localization or dominant-negative activity of focal adhesion kinase/focal adhesion kinase-related nonkinase.
The carboxy-terminal 150 residues of the focal adhesion kinase (FAK) comprise the focal adhesion-targeting sequence, which is responsible for its subcellular localization. The mechanism of focal adhesion targeting has not been fully elucidated. We describe a mutational analysis of the focal adhesion-targeting sequence of FAK to further examine the mechanism of focal adhesion targeting and explore additional functions encoded by the carboxy-terminus of FAK. The results demonstrate that paxillin binding is dispensable for focal adhesion targeting of FAK. Cell adhesion-dependent tyrosine phosphorylation strictly correlated with the ability of mutants to target to focal adhesions. Focal adhesion targeting was also a requirement for maximal FAK-dependent tyrosine phosphorylation of paxillin and FAK-related nonkinase (FRNK)-dependent inhibition of endogenous FAK function. However, there were additional requirements for these latter functions because we identified mutants that target to focal adhesions, yet are defective for the induction of paxillin phosphorylation or the dominant-negative function of FRNK. Furthermore, the paxillin-binding activity of FRNK mutants did not correlate with their ability to inhibit FAK, suggesting that FRNK has other targets in addition to paxillin.
Cooley
M A
MA
Departments of Cell Biology & Anatomy, University of North Carolina, Chapel Hill, North Carolina 27599, USA.
Broome
J M
JM
Ohngemach
C
C
Romer
L H
LH
Schaller
M D
MD
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D002642:Chick Embryo; D003598:Cytoskeletal Proteins / Q000737:chemistry / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D014644:Genetic Variation; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D051419:Paxillin; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions / Q000378:metabolism / Q000648:ultrastructure; D014162:Transfection
D003598:Cytoskeletal Proteins; D051419:Paxillin; D010750:Phosphoproteins; C496062:Pxn protein, mouse; D011993:Recombinant Fusion Proteins; C113183:FAK-related nonkinase; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse