Structural requirements for the efficient regulation of the Src protein tyrosine kinase by Csk.
Protein tyrosine kinases of the Src family are negatively regulated by phosphorylation in the C-terminal tail of the molecule. A different protein tyrosine kinase, Csk, is largely responsible for this regulation. The phosphorylated tail of c-Src engages with the SH2 domain in a conformation that is associated with low kinase activity and which involves stabilization by the SH3 domain. Inducible expression of c-Src in fission yeast is lethal unless Csk is coexpressed. Using this assay we present evidence that Src regulation by C-terminal phosphorylation does not require the myristylation signal or the unique domain at the N-terminus of the Src protein. Mutagenesis of the SH3 and SH2 domains of Csk show that neither are necessary in yeast or in vitro for efficient regulation of Src. Mutation of Tyr416 of Src, a site of autophosphorylation common to most protein tyrosine kinases, abolished the ability of Src to arrest growth of phosphorylate endogenous proteins. Tyr416 had the same effect on a shorter form of Src consisting of the kinase domain only, indicating that the mutation affects a property intrinsic to the catalytic domain. The residual activity of full-length Src mutated at Tyr416 is efficiently repressed by Csk action, suggesting that regulation by C-terminal phosphorylation does not act by preventing phosphorylation at Tyr416.
Koegl
M
M
Differentiation Programme, European Molecular Biology Laboratory, Heidelberg, Germany.
Courtneidge
S A
SA
Superti-Furga
G
G
D000081247:CSK Tyrosine-Protein Kinase; D015536:Down-Regulation; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism; D010766:Phosphorylation; D011487:Protein Conformation; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000235:genetics*; D012568:Schizosaccharomyces / Q000235:genetics; D015398:Signal Transduction; D013379:Substrate Specificity; D019061:src-Family Kinases
D009227:Myristic Acids; D019814:Myristic Acid; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D016392:Proto-Oncogene Proteins pp60(c-src); D019061:src-Family Kinases