Glycosylation at ASN211 regulates the activation state of the discoidin domain receptor 1 (DDR1) [Protein Structure and Folding]
February 7th, 2014 by Fu, H.-L., Valiathan, R. R., Payne, L., Kumarasiri, M., Mahasenan, K. V., Mobashery, S., Huang, P., Fridman, R.
Discoidin Domain Receptor 1 (DDR1) belongs to a unique family of receptor tyrosine kinases (RTKs) that signal in response to collagens. DDR1 undergoes autophosphorylation in response to collagen binding with a slow and sustained kinetics that is unique among members of the RTK family. DDR1 dimerization precedes receptor activation suggesting a structural inhibitory mechanism to prevent unwarranted phosphorylation. However, the mechanism(s) that maintains the autoinhibitory state of the DDR1 dimers is unknown. Here we report that N-glycosylation at the N211 residue plays a unique role in the control of DDR1 dimerization and autophosphorylation. Using site-directed mutagenesis we found that mutations that disrupt the conserved N211DS N-glycosylation motif, but not other N-glycosylation sites (N260, N371, N394), result in collagen I-independent constitutive phosphorylation. Mass spectrometry revealed that the N211Q mutant undergoes phosphorylation at Y484, Y520, Y792, and Y797. The N211Q traffics to the cells surface and its ectodomain displays collagen I binding with an affinity similar to that of the wild type DDR1 ectodomain. However, unlike the wild type receptor, the N211Q mutant exhibits enhanced receptor dimerization and sustained activation upon ligand withdrawal. Taken together, these data suggest that N-glycosylation at the highly conserved N211DS motif evolved to act as a negative repressor of DDR1 phosphorylation in the absence of ligand. The presence of glycan moieties at that site may help to lock the collagen-binding domain in the inactive state and prevent unwarranted signaling by receptor dimers. These studies provide a novel insight into the structural mechanisms that regulate DDR activation.