Stimulation of phosphorylation of Tyr394 by hydrogen peroxide reactivates biologically inactive, non-membrane-bound forms of Lck.
Lck, a lymphocyte-specific tyrosine protein kinase, is bound to cellular membranes as the result of myristoylation and palmitoylation of its amino terminus. Its activity is inhibited by phosphorylation of tyrosine 394. The Tyr-505 --> Phe mutant of Lck (F505Lck) exhibits elevated biological activity and constitutive phosphorylation of Tyr-394 in vivo. Mutations at sites of fatty acylation that prevent F505Lck from associating with cellular membranes abolish the biological activity as a protein kinase in vivo and in vitro, and eliminate the phosphorylation of Tyr-394. Here, we show that exposure of cells expressing cytoplasmic or nuclear forms of F505Lck to H2O2, a general inhibitor of tyrosine protein phosphatases, restores the catalytic activity of these mutant proteins through stimulation of phosphorylation of Tyr-394. H2O2 treatment induced the phosphorylation of Tyr-394 therefore need not occur by autophosphorylation. Thus, there appear to be two mechanisms through which the phosphorylation of Lck at Tyr-394 can occur. One is restricted to the plasma membrane and does not require the presence of oxidants. The other is operational in the nucleus as well as the cytosol and is responsive to oxidants.
Yurchak
L K
LK
Molecular Biology and Virology Laboratory, Salk Institute, San Diego, California 92186, USA.
Hardwick
J S
JS
Amrein
K
K
Pierno
K
K
Sefton
B M
BM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002384:Catalysis; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D004793:Enzyme Reactivators / Q000494:pharmacology*; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D008969:Molecular Sequence Data; D010766:Phosphorylation; D051381:Rats; D013379:Substrate Specificity; D014443:Tyrosine / Q000378:metabolism*; D019061:src-Family Kinases / Q000378:metabolism*
D004793:Enzyme Reactivators; D014443:Tyrosine; D006861:Hydrogen Peroxide; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D019061:src-Family Kinases