Glycosylation of the murine estrogen receptor-alpha

O-linked N-acetylglucosamine (O-GlcNAc) is a highly dynamic and abundant mo dification found on nuclear and cytoplasmic proteins of nearly all eukaryot es. O-GlcNAc addition is required for life at the single cell level and is analogous to protein phosphorylation in most respects. In a previous study (M.S. Jiang, G.W. Hart, A subpopulation of estrogen receptors are modified by O-linked N-acetylglucosamine. J. Biol. Chem. 270 (1997) 2421-2428), we d emonstrated that a subpopulation of the murine estrogen receptor-alpha (mER -alpha) is modified by O-GlcNAc at Th-575. Here we mutated mER-alpha to con vert Thr(575) and Ser(576) to Val and Ala, respectively. Surprisingly, this glycosylation-site mutant is still extensively modified by O-GlcNAc. Analy ses of glycopeptides identified two additional sites of modification on mER -alpha, at Ser(10) and Thr(50) near the N-terminus. The major glycosylation sites are within or near PEST regions, suggesting that O-GlcNAc may regula te mER-alpha turnover. (C) 2001 Elsevier Science Ltd. All rights reserved.