The
linker for activation of T-ce
lls (LAT) is a pa
lmitoy
lated integra
l membrane adaptor proteinthat resides in
lipid membrane rafts and contains nine consensus putative tyrosine phosphory
lation sites,severa
l of which have been shown to serve as SH2 binding sites. Upon T-ce
ll antigen receptor (TCR/CD3) engagement, LAT is phosphory
lated by protein tyrosine kinases (PTK) and binds to the adaptorsGads and Grb2, as we
ll as to phospho
lipase C
![](/images/gifchars/gamma.gif)
1 (PLC
![](/images/gifchars/gamma.gif)
1), thereby faci
litating the recruitment of keysigna
l transduction components to drive T-ce
ll activation. The LAT tyrosine residues Y
132, Y
171, Y
191,and Y
226 have been shown previous
ly to be critica
l for binding to Gads, Grb2, and PLC
![](/images/gifchars/gamma.gif)
1. In this report,we show by generation of LAT truncation mutants that the Syk-fami
ly kinase ZAP-70 and the Tec-fami
ly kinase Itk favor phosphory
lation of carboxy-termina
l tyrosines in LAT. By direct binding studiesusing purified recombinant proteins or phosphopeptides and by mutagenesis of individua
l tyrosines inLAT to pheny
la
lanine residues, we demonstrate that Y
171 and potentia
lly Y
226 are docking sites for theVav guanine nuc
leotide exchange factor. Further, overexpression of a kinase-deficient mutant of Itk inT-ce
lls reduced both the tyrosine phosphory
lation of endogenous LAT and the recruitment of Vav toLAT comp
lexes. These data indicate that kinases from distinct PTK fami
lies are
like
ly responsib
le forLAT phosphory
lation fo
llowing T-ce
ll activation and that Itk kinase activity promotes recruitment ofVav to LAT.