Trking Down an Inside Job

Abstract

Nerve growth factor (NGF) binds to the TrkA receptor tyrosine kinase at the cell surface, leading to receptor autophosphorylation and activation of intracellular signaling pathways. However, ligand binding to some G protein (heterotrimeric guanine nucleotide-binding protein)-coupled receptors (GPCRs), such as the adenosine A 2A receptor or the pituitary adenylate cyclase-activating polypeptide (PACAP) PAC1 receptor, can independently stimulate TrkA phosphorylation and activation of TrkA signaling pathways. Rajagopal et al. used phosphotyrosine-specific antibodies on pan-Trk immunoprecipitates of PC12-615 cells to investigate TrkA phosphorylation in response to adenosine and observed that forms of the TrkA receptor that are not targeted to the plasma membrane were phosphorylated first. Inhibitors of transcription or translation blocked phosphorylation in response to the adenosine agonist CGS 21680 but not phosphorylation induced by NGF; reverse transcription polymerase chain reaction (RT-PCR) indicated that CGS 21680 treatment did not elicit NGF production. Cell surface biotinylation experiments indicated that CGS 21680 activated an intracellular pool of Trk receptors. Immunofluorescence analysis in combination with confocal microscopy revealed that, whereas NGF treatment stimulated TrkA phosphorylation at the plasma membrane, CGS 21680 activated perinuclear TrkA associated with the Golgi apparatus. Moreover, pharmacological inhibition of Golgi vesicle fusion abolished perinuclear TrkA immunoreactivity and blocked CGS 21680-dependent TrkA phosphorylation and the downstream phosphorylation of Akt. PACAP stimulated a similar pattern of perinuclear immunofluorescence in primary cultures of basal forebrain neurons. Thus, GPCR-dependent activation of TrkA appears to take place in an intracellular compartment, a finding the authors discuss in the context of retrograde signaling from internalized TrkA receptors. R. Rajagopal, Z.-Y. Chen, F. S. Lee, M. V. Chao, Transactivation of Trk neurotrophin receptors by G-protein-coupled receptor ligands occurs on intracellular membranes. J. Neurosci . 24 , 6650-6658 (2004). [Abstract] [Full Text]