ABERRANT TCR-MEDIATED SIGNALING IN CD45-NULL THYMOCYTES INVOLVES DYSFUNCTIONAL REGULATION OF LCK, FYN, TCR-ZETA AND ZAP-70

CD45 is a transmembrane phosphotyrosine phosphatase expressed on all n ucleated hemopoietic cells, Targeting of CD45 exon 9 has generated a m ouse line completely lacking CD45 expression (CD45-null) in which ther e are severe abnormalities in T cell development, Defects in TCR-media ted signals underlying these abnormalities have now been investigated using CD45-null T cells. No T cell proliferation was detected in respo nse to a CD3 mAb, In thymocytes the p56(lck) and p596(fyn) tyrosine ki nases were hyperphosphorylated, and p56(lck) was in its inactive confo rmation. Both basal and TCR-stimulated tyrosine phosphorylation of TCR -zeta and CD3-epsilon were much reduced, and TCR stimulation induced a n abnormal p18 phosphoisomer of TCR-zeta previously noted in T cells s timulated by altered peptide ligands, These defects were associated wi th the failure of ZAP-70 kinase recruitment to the TCR-zeta chain. TCR coupling to the tyrosine phosphorylation of several proteins, includi ng HS1 and p120(cbl), was also much reduced. However, TCR-induced sign aling was not ablated, and significant inositol phosphate and calcium signals were observed in CD45-null thymocytes, Our molecular analysis suggests that the threshold for TCR signal transduction is greatly inc reased in CD45-null T cells, thus explaining the profound defects in t hymic development.