SIGNALS DETERMINING PROTEIN-TYROSINE KINASE AND GLYCOSYL-PHOSPHATIDYLINOSITOL-ANCHORED PROTEIN TARGETING TO A GLYCOLIPID-ENRICHED MEMBRANE-FRACTION

Glycosyl-phosphatidylinositol (GPI)-anchored membrane proteins and cer tain protein tyrosine kinases associate with a Triton X-100-insoluble, glycolipid-enriched membrane fraction in MDCK cells. Also, certain pr otein tyrosine kinases have been shown to associate with GPI-anchored proteins in other cell types. To characterize the interaction between GPI-anchored proteins and protein tyrosine kinases, GPI-anchored prote ins were coexpressed with p56(lck) in HeLa cells. Both proteins were s hown to target independently to the glycolipid-enriched membranes. Coi mmunoprecipitation of GPI-anchored proteins and p56(lck) occurred only when both proteins were located in the glycolipid-enriched membranes, and gentle disruption of these membranes abolished the interaction. T he GPI anchor was found to be the targeting signal for this membrane f raction in GPI-anchored proteins. Analysis of mutants indicated that p 56(lck) was nearly quantitatively palmitoylated at Cys-5 but not palmi toylated at Cys-3. The nonpalmitoylated cysteine at position 3 was ver y important for association of p56(lck) with the membrane fraction, wh ile palmitoylation at Cys-5 promoted only a low level of interaction. Because other src family protein tyrosine kinases that are associated with GPI-anchored proteins always contain a Cys-3, we propose that thi s residue, in addition to the N-terminal myristate, is part of a commo n signal targeting these proteins to a membrane domain that has been l inked to transmembrane signaling.