TrkA glycosylation regulates receptor localization and activity

The human nerve growth factor receptor (TrkA) contains four potential N-gly cosylation sites that are highly conserved within the Trk family of neurotr ophin receptors, and nine additional sites that are less well conserved. Us ing a microscale deglycosylation assay, we show here that both conserved an d variable N-glycosylation sites are used during maturation of TrkA. Glycos ylation at these sites serves two distinct functions, First, glycosylation is necessary to prevent Ligand-independent activation of TrkA, Unglycosylat ed TrkA core protein is phosphorylated even in the absence of ligand stimul ation and displays constitutive kinase activity as well as constitutive int eraction with the signaling molecules Shc and PLC-gamma. Second, glycosylat ion is required to localize TrkA to the cell surface, where it can trigger the Ras/Raf/MAP kinase cascade. Using confocal microscopy, we show that ung lycosylated active Trk receptors are trapped intracellularly. Furthermore, the unglycosylated active TrkA receptors are unable to activate kinases in the Ras-MAP kinase pathway, MEK and Erk. Consistent Kith these biochemical observations, unglycosylated TrkA core protein does not promote neuronal di fferentiation in Trk PC12 cells even at high levels of constitutive catalyt ic activity, (C) 1999 John Wiley & Sons, Inc.