GIT1 mediates Src-dependent activation of phospholipase Cgamma by angiotensin II and epidermal growth factor.
Critical events for vasoconstrictor and growth factor signal transduction include stimulation of phospholipase Cgamma (PLCgamma) and elevation of intracellular calcium. c-Src has been proposed as a common mediator for these signals activated by both G protein-coupled receptors (GPCRs) and tyrosine kinase-coupled receptors (TKRs). Here we show that the GPCR kinase-interacting protein-1 (GIT1) is a substrate for c-Src that undergoes tyrosine phosphorylation in response to angiotensin II (AngII) and EGF in vascular smooth muscle and 293 cells. GIT1 associates with PLCgamma via the PLCgamma Src homology 2 and 3 domains constitutively, and the interaction is unaltered by AngII and EGF. GIT1 interaction with PLCgamma is required for PLCgamma activation based on inhibition of tyrosine phosphorylation and calcium mobilization after GIT1 knockdown with antisense GIT1 oligonucleotides. GIT1 interacts with PLCgamma via a novel Spa homology domain (SHD) and a coiled-coil domain. Deletion mutation analysis showed that GIT1(SHD) is required for AngII- and EGF-mediated PLCgamma activation (measured by phosphorylation of Tyr783 and inositol 1,4,5-trisphosphate formation). We propose that GIT1 is a novel regulator of PLCgamma function that mediates PLCgamma activation by c-Src and integrates signal transduction by GPCRs and TKRs.
Haendeler
Judith
J
Center for Cardiovascular Research and Department of Medicine, University of Rochester, Rochester, New York 14642, USA.
Yin
Guoyong
G
Hojo
Yukihiro
Y
Saito
Yuji
Y
Melaragno
Matthew
M
Yan
Chen
C
Sharma
Virendra K
VK
Heller
Manfred
M
Aebersold
Ruedi
R
Berk
Bradford C
BC
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D018797:Cell Cycle Proteins; D002460:Cell Line; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D004789:Enzyme Activation; D004815:Epidermal Growth Factor / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000502:physiology*; D017353:Gene Deletion; D006801:Humans; D015151:Immunoblotting; D013058:Mass Spectrometry; D008954:Models, Biological; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009130:Muscle, Smooth / Q000166:cytology; D009841:Oligonucleotides / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D051966:Phospholipase C gamma; D010750:Phosphoproteins; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; C485647:GIT1 protein, human; D020690:GTPase-Activating Proteins; C485648:Git1 protein, rat; D009841:Oligonucleotides; D010455:Peptides; D010750:Phosphoproteins; D000804:Angiotensin II; D014443:Tyrosine; D004815:Epidermal Growth Factor; D019061:src-Family Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D002118:Calcium