Regulation of the inositol 1,4,5-trisphosphate receptor type I by O-GlcNAc glycosylation

The inositol 1,4,5-trisphosphate (InsP(3)) receptor type I (InsP(3)R-I) is the principle channel for intracellular calcium (Ca2+) release in many cell types, including central neurons. It is regulated by endogenous compounds like Ca2+ and ATP, by protein partners, and by posttranslational modification. We report that the InsP(3)R-I is modified by O-linked glycosylation of serine or threonine residues with beta-N-acetylglucosamine (O-GlcNAc). The level of O-GlcNAcylation can be altered in vitro by the addition of the enzymes which add [OGT (O-GlcNActransferase)] or remove (O-GlcNAcase) this sugar or by loading cells with UDP-GlcNAc. We monitored the effects of this modification on InsP(3)R function at the single-channel level and on intracellular Ca2+ transients. Single-channel activity was monitored with InsP(3)R incorporated into bilayers; Ca2+ signaling was monitored using cells loaded with a Ca2+- sensitive fluorophore. We found that channel activity was decreased by the addition of O-GlcNAc and that this decrease was reversed by removal of the sugar. Similarly, cells loaded with UDP-GlcNAc had an attenuated response to uncaging of InsP(3). These results show that O-GlcNAcylation is an important regulator of the InsP(3)R-I and suggest a mechanism for neuronal dysfunction under conditions in which O-GlcNAc is high, such as diabetes or physiological stress.