The protein-tyrosine kinase Lck associates with and is phosphorylated by Cdc2.
The protein-tyrosine kinase Lck is essential for signaling through the T-cell antigen receptor. Treatment of T-cells with a variety of extracellular stimuli increases the phosphorylation of Lck on serine residues. This results in shifts in the apparent molecular weight of Lck to forms that exhibit reduced electrophoretic mobility on SDS-polyacrylamide gels. We found that as a result of arresting cells in mitosis, forms of Lck were generated that migrated with slower mobilities on SDS-polyacrylamide gels. This suggested that a serine/threonine kinase, active at mitosis, was phosphorylating Lck. Using antibodies to Lck and to the cyclin-dependent serine kinase, Cdc2, as well as the cyclin-dependent kinase affinity resin, Suc1-agarose, we detected a stable interaction between Lck and Cdc2. The interaction was mediated through the Src homology 3 domain of Lck and was selective, as only the active form of Cdc2 was found to associate with Lck. Moreover, Cdc2 was able to phosphorylate Lck in vitro and shift its electrophoretic mobility to a more slowly migrating form. An association between active Cdc2 and the Src-related kinases Lyn and Fyn was also demonstrated, although Cdc2 was not found associated with the tyrosine kinases, Csk and Syk. These results demonstrate that at mitosis, Cdc2 associates with and phosphorylates Lck.
Pathan
N I
NI
Department of Biology, Purdue University, West Lafayette, Indiana 47907, USA.
Geahlen
R L
RL
Harrison
M L
ML
D000818:Animals; D000906:Antibodies; D016590:Aphidicolin / Q000494:pharmacology; D016203:CDC2 Protein Kinase / Q000302:isolation & purification / Q000378:metabolism*; D002460:Cell Line; D002846:Chromatography, Affinity; D006801:Humans; D019169:Jurkat Cells; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D051379:Mice; D008938:Mitosis; D015739:Nocodazole / Q000494:pharmacology; D010766:Phosphorylation; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D018411:Spodoptera; D014162:Transfection; D019061:src-Family Kinases / Q000302:isolation & purification / Q000378:metabolism*
D000906:Antibodies; D011993:Recombinant Fusion Proteins; D016590:Aphidicolin; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D019061:src-Family Kinases; D016203:CDC2 Protein Kinase; D015739:Nocodazole