title: Src-dependent TrkA transactivation is required for pituitary adenylate cyclase-activating polypeptide 38-mediated Rit activation and neuronal differentiation.
abstract: Pituitary adenylate cyclase-activating polypeptide (PACAP) is a potent neuropeptide that possesses both neurotrophic and neurodevelopmental effects. Recently, the Rit GTPase was found to be activated by a novel Galpha/cAMP/exchange protein activated by cyclic AMP (Epac)-dependent signaling pathway and required for PACAP-dependent cAMP response element-binding protein activation and neuronal differentiation. However, Epac did not function as a Rit guanine nucleotide exchange factor (GEF), and the nature of the PACAP regulatory cascade remained unclear. Here, we show that PACAP-mediated Rit activation involves Src family kinase-dependent TrkA receptor transactivation. PACAP receptor (PACR1) stimulation triggered both G(i)alpha and G(s)alpha/cAMP/Epac regulatory cascades resulting in Src kinase activity, which in turn induced TrkA kinase tyrosine phosphorylation. Importantly, Src inhibition, or the lack of functional Trk receptors, was found to inhibit PACAP-mediated Rit activation, whereas constitutively active Src alone was sufficient to stimulate Rit-guanosine triphosphate levels. A single tyrosine (Y(499)) phosphorylation event was identified as critical to both PACAP-mediated transactivation and TrkA-dependent Rit activation. Accordingly, PACAP stimulation resulted in TrkA-dependent phosphorylation of both the Shc adaptor and son of sevenless (SOS)1/2 GEFs, and Rit activation was inhibited by RNA interference silencing of SOS1/2, implicating a TrkA/Shc/SOS signaling complex in Rit regulation. Together, these observations expand upon the nature of PACR1-mediated transactivation and identify TrkA-Rit signaling as a key contributor to PACAP-dependent neuronal differentiation.
authors.affiliation: Department of Molecular and Cellular Biochemistry, University of Kentucky College of Medicine, Lexington, KY 40536-0509, USA. gshi2@uky.edu
authors.forename: Geng-Xian
authors.initials: GX
authors.lastname: Shi
authors.forename: Ling
authors.initials: L
authors.lastname: Jin
authors.forename: Douglas A
authors.initials: DA
authors.lastname: Andres
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D045744:Cell Line, Tumor; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000378:metabolism; D015151:Immunoblotting; D051379:Mice; D009154:Mutation; D041681:NIH 3T3 Cells; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D016716:PC12 Cells; D037342:Pertussis Toxin / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide / Q000494:pharmacology*; D011743:Pyrimidines / Q000494:pharmacology; D034622:RNA Interference; D051381:Rats; D020917:Receptor, trkA / Q000235:genetics / Q000378:metabolism*; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D020664:Son of Sevenless Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism*; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: C412373:AG 1879; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D011743:Pyrimidines; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; D020664:Son of Sevenless Proteins; D037342:Pertussis Toxin; D020917:Receptor, trkA; D019061:src-Family Kinases; C486777:Rit protein, rat; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D019205:GTP-Binding Protein alpha Subunits, Gs; D018631:ras Proteins