Src SH2 arginine 175 is required for cell motility: specific focal adhesion kinase targeting and focal adhesion assembly function.
Src kinase is a crucial mediator of adhesion-related signaling and motility. Src binds to focal adhesion kinase (FAK) through its SH2 domain and subsequently activates it for phosphorylation of downstream substrates. In addition to this binding function, data suggested that the SH2 domain might also perform an important role in targeting Src to focal adhesions (FAs) to enable further substrate phosphorylations. To examine this, we engineered an R175L mutation in cSrc to prevent the interaction with FAK pY397. This constitutively open Src kinase mediated up-regulated substrate phosphorylation in SYF cells but was unable to promote malignant transformation. Significantly, SrcR175L cells also had a profound motility defect and an impaired FA generation capacity. Importantly, we were able to recapitulate wild-type motile behavior and FA formation by directing the kinase to FAs, clearly implicating the SH2 domain in recruitment to FAK and indicating that this targeting capacity, and not simply Src-FAK scaffolding, was critical for normal Src function.
Yeo
Myeong Gu
MG
Department of Pathology and Cell Biology, Columbia University College of Physicians and Surgeons, 630 West 168th St., New York, NY 10032, USA.
Partridge
Michael A
MA
Ezratty
Ellen J
EJ
Shen
Qiong
Q
Gundersen
Gregg G
GG
Marcantonio
Eugene E
EE
D019943:Amino Acid Substitution; D000818:Animals; D001120:Arginine / Q000737:chemistry; D002460:Cell Line; D002465:Cell Movement / Q000502:physiology*; D002645:Chickens; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D022001:Focal Adhesions / Q000502:physiology*; D016391:Genes, src; D006801:Humans; D066298:In Vitro Techniques; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D018909:src Homology Domains; D019061:src-Family Kinases / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*
D001120:Arginine; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019061:src-Family Kinases