Phospholipase C-gamma1 interacts with conserved phosphotyrosyl residues in the linker region of Syk and is a substrate for Syk.
Antigen receptor ligation on lymphocytes activates protein tyrosine kinases and phospholipase C-gamma (PLC-gamma) isoforms. Glutathione S-transferase fusion proteins containing the C-terminal Src-homology 2 [SH2(C)] domain of PLC-gamma1 bound to tyrosyl phosphorylated Syk. Syk isolated from antigen receptor-activated B cells phosphorylated PLC-gamma1 on Tyr-771 and the key regulatory residue Tyr-783 in vitro, whereas Lyn from the same B cells phosphorylated PLC-gamma1 only on Tyr-771. The ability of Syk to phosphorylate PLC-gamma1 required antigen receptor ligation, while Lyn was constitutively active. An mCD8-Syk cDNA construct could be expressed as a tyrosyl-phosphorylated chimeric protein tyrosine kinase in COS cells, was recognized by PLC-gamma1 SH2(C) in vitro, and induced tyrosyl phosphorylation of endogenous PLC-gamma1 in vivo. Substitution of Tyr-525 and Tyr-526 at the autophosphorylation site of Syk in mCD8-Syk substantially reduced the kinase activity and the binding of this variant chimera to PLC-gamma1 SH2(C) in vitro; it also failed to induce tyrosyl phosphorylation of PLC-gamma1 in vivo. In contrast, substitution of Tyr-348 and Tyr-352 in the linker region of Syk in mCD8-Syk did not affect the kinase activity of this variant chimera but almost completely eliminated its binding to PLC-gamma1 SH(C) and completely eliminated its ability to induce tyrosyl phosphorylation of PLC-gamma1 in vivo. Thus, an optimal kinase activity of Syk and an interaction between the linker region of Syk with PLC-gamma1 are required for the tyrosyl phosphorylation of PLC-gamma1.
Law
C L
CL
Department of Microbiology, University of Washington, Seattle, USA.
Chandran
K A
KA
Sidorenko
S P
SP
Clark
E A
EA
D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000201:enzymology / Q000378:metabolism*; D001483:Base Sequence; D002460:Cell Line; D017124:Conserved Sequence; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D051966:Phospholipase C gamma; D010766:Phosphorylation; D010957:Plasmids; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011947:Receptors, Antigen, B-Cell / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D000072377:Syk Kinase; D010738:Type C Phospholipases / Q000235:genetics / Q000378:metabolism*; D014443:Tyrosine / Q000235:genetics; D051746:ZAP-70 Protein-Tyrosine Kinase; D018909:src Homology Domains / Q000235:genetics*
D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; D011947:Receptors, Antigen, B-Cell; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human; D010738:Type C Phospholipases; D051966:Phospholipase C gamma