T-CELL RECEPTOR AND CD8-DEPENDENT TYROSINE PHOSPHORYLATION EVENTS IN CYTOTOXIC T-LYMPHOCYTES - ACTIVATION OF P56(LCK) BY CD8 BINDING TO CLASS-I PROTEIN

Tyrosine phosphorylation of proteins plays a central role in T cell ac tivation. Mitogens or anti-receptor antibodies have been employed to s tudy these signaling events, but the extent to which these mimic recep tor interactions with native ligands is unclear, Cytotoxic T lymphocyt es can be activated for functional responses using purified, native cl ass I ligands presented on a surface. Previous work showed that stimul ation with fluid-phase anti-T cell receptor (TCR) monoclonal antibody (mAb) activates CDS to mediate adhesion to class I proteins and that a ctivated CD8 generates a co-stimulatory signal upon binding to class I . Changes in tyrosine phosphorylation of substrates and activity of th e p56(lck) kinase have now been examined in this two-step process, The observed changes are small in comparison to those found using more po tent nonphysiological stir-null, but may more accurately reflect the e vents required for activation of functional responses. Fluid-phase ant i-TCR mAb caused increased tyrosine phosphorylation of a discrete subs et of cellular substrates. Increased phosphorylation of additional sub strates occurred upon CD8 binding to class I, resulting in a phosphory lation pattern comparable to that found in cells stimulated with class I alloantigen. Anti-TCR mAb alone caused increased tyrosine phosphory lation of p56(lck). When CDS bound to class I, phosphorylation of p56( lck) decreased to below the basal level Pound in unstimulated cells, a ccompanied by a substantial increase in kinase activity, These results are consistent with the two-step model for TCR activation of CD8/clas s I interactions and directly demonstrate that CD8 binding to class I leads to up-regulation of p56(lck) activity.