EVIDENCE THAT PROTEIN-TYROSINE KINASE P56-LCK REGULATES THE ACTIVITY OF PHOSPHATIDYLINOSITOL-3'-KINASE IN INTERLEUKIN-2-DEPENDENT T-CELLS

The relative levels of phosphatidylinositol-3'-kinase (PI3K) activity were measured in interleukin-2 (IL-2)-dependent helper (HT-2) and cyto lytic (CTLL-2) T-cell clones that had been stably transfected with exp ression plasmids encoding either the normal p56-Lck kinase or a mutant version of this kinase, p56-Lck(Y505F), that has constitutively high levels of kinase activity. Stimulation of untransfected T-cells or of transfected T-cells containing increased levels of normal p56-Lck resu lted in an approximate doubling of the relative amounts of total cellu lar PI3K activity. In contrast, T-cells producing the activated versio n of p56-Lck contained levels of PI3K activity comparable to or slight ly higher than those found IL-2-stimulated control cells, even in the absence of IL-2. Assessments of the relative levels of PI3K activity i n immunoprecipitates prepared with the use of anti-phosphotyrosine-spe cific antibodies revealed constitutively high levels of anti-phosphoty rosine-immunoprecipitable PI3K activity in T-cells containing p56-Lck( Y505F), as opposed to T-cells containing normal p56-Lck where increase s in anti-PY-immunoprecipitable PI3K activity were IL-2-inducible. IL- 2 stimulation of T-cells containing the normal p56-Lck kinase resulted in marked increases in the relative amounts of PI3K activity and p85 that were coimmunoprecipitated when using anti-p56-Lck antibodies. In contrast, PI3K activity and the p85 subunit of PI3K could be coimmunop recipitated from T-cells producing the activated p56-Lck(Y505F) kinase even in the absence of IL-2 stimulation, implying constitutive associ ation of PI3K with the activated Lck kinase. Taken together with previ ous studies showing that IL-2 induces rapid increases in the activitie s of both p56-Lck and PI3K in T-cells, these findings suggest that p56 -Lck lies immediately upstream of PI3K in a signal transduction cascad e initiated by the binding of IL-2 to its specific receptor on T-lymph ocytes.