An ITIM-like motif within the CCK2 receptor sequence required for interaction with SHP-2 and the activation of the AKT pathway.
SHP-2 is a tyrosine phosphatase which functions as a positive regulator downstream of RTKs, activating growth-stimulatory signalling pathways. To date, very few G protein-coupled receptors (GPCRs) have been shown to be connected to SHP-2 and very little is known about the positive role of SHP-2 in GPCR signalling. The CCK2 receptor (CCK2R), a GPCR, is now recognized to mediate mitogenic effects of gastrin on gastrointestinal cells. In the present study, we demonstrate the role of SHP-2 in the activation of the AKT pathway by the CCK2R in COS-7 cells transfected with the CCK2R and in a pancreatic cancer cell line expressing the endogenous receptor. Using surface plasmon resonance analysis, we identified a highly conserved ITIM motif, containing the tyrosine residue 438, located in the C-terminal intracellular tail of the CCK2R which directly interacts with the SHP-2 SH2 domains. The interaction was confirmed by pull down assays and co-immunoprecipitation of the receptor with SHP-2. This interaction was transiently increased following gastrin stimulation of the CCK2R and correlated with the tyrosine phosphorylation of SHP-2. Mutational analysis of the key ITIM residue 438 confirmed that the CCK2R ITIM sequence is required for interaction with SHP-2 and the activation of the AKT pathway.
Vatinel
Sébastien
S
INSERM U 531, IFR 31, Institut Louis Bugnard, BP 84225, 31432 Toulouse cedex 4, France.
Ferrand
Audrey
A
Lopez
Fréderic
F
Kowalski-Chauvel
Aline
A
Estève
Jean-Pierre
JP
Fourmy
Daniel
D
Dufresne
Marlène
M
Seva
Catherine
C
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D005755:Gastrins / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells; D010190:Pancreatic Neoplasms / Q000378:metabolism*; D010766:Phosphorylation; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism* / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D051381:Rats; D043887:Receptor, Cholecystokinin B / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism
D005755:Gastrins; D047908:Intracellular Signaling Peptides and Proteins; D043887:Receptor, Cholecystokinin B; D014443:Tyrosine; D051057:Proto-Oncogene Proteins c-akt; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D017027:Protein Tyrosine Phosphatases; C516725:Ptpn11 protein, mouse; C516727:Ptpn11 protein, rat