Cbl-associated protein is tyrosine phosphorylated by c-Abl and c-Src kinases.
BACKGROUND
The c-Cbl-associated protein (CAP), also known as ponsin, localizes to focal adhesions and stress fibers and is involved in signaling events. Phosphorylation has been described for the other two members of the sorbin homology family, vinexin and ArgBP2, but no data exist about the putative phosphorylation of CAP. According to previous findings, CAP binds to tyrosine kinase c-Abl. However, it is not known if CAP is a substrate of c-Abl or other tyrosine kinases or if phosphorylation regulates its localization.


RESULTS
We here show that CAP is Tyr phosphorylated by and interacts with both c-Abl and c-Src. One major phosphorylation site, Tyr360, and two minor contributors Tyr326 and Tyr632 were identified as Abl phosphorylation sites, whereas Src preferentially phosphorylates Tyr326 and Tyr360. Phosphorylation of CAP was not necessary for its localization to focal adhesions and stress fibers, but Tyr326Phe substitution alters the function of CAP during cell spreading.


CONCLUSIONS
This is the first demonstration of phosphorylation of CAP by any kinase. Our findings suggest that coordinated action of Src and Abl might regulate the function of CAP and reveal a functional role especially for the Src-mediated Tyr phosphorylation of CAP in cell spreading.
Fernow
Inga
I
Institute of Biochemistry, University of Giessen, Friedrichstrasse 24, 35392 Giessen, Germany. inga.fernow@web.de
Tomasovic
Ana
A
Siehoff-Icking
Ann
A
Tikkanen
Ritva
R
D019943:Amino Acid Substitution; D000081247:CSK Tyrosine-Protein Kinase; D045744:Cell Line, Tumor; D022001:Focal Adhesions / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D007328:Insulin / Q000494:pharmacology; D008840:Microfilament Proteins / Q000032:analysis / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D016315:Proto-Oncogene Proteins c-abl / Q000378:metabolism*; D022502:Stress Fibers / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism*; D014638:Vanadates / Q000494:pharmacology; D019061:src-Family Kinases
D007328:Insulin; D008840:Microfilament Proteins; C119201:ponsin; D014638:Vanadates; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D016315:Proto-Oncogene Proteins c-abl; D019061:src-Family Kinases; C000631346:CSK protein, human