Alternative O-glycosylation/O-phosphorylation of the murine estrogen receptor beta

Estrogen receptor beta, a homologue to estrogen receptor ct, is a new membe r of the steroid hormone receptor family. Recently, we documented that estr ogen receptor ct, like other transcription factors, is modified by O-linked N-acetylglucosamine (O-GlcNAc), a ubiquitous transitory posttranslational modification on nuclear and cytoplasmic proteins. Here, we report that estr ogen receptor beta is alternatively modified by either O-GlcNAc or O-phosph ate. Lectin chromatography of in vitro translated protein first suggested t hat murine estrogen receptor beta (mER-beta) is O-GlcNAcylated. Structural characterization of the carbohydrate moieties on mER-beta, overexpressed in insect Sf9 cells, confirmed the presence of O-GlcNAc. mER-beta, overexpres sed in mammalian cells, is also O-GlcNAcylated, The major site of O-GlcNAc on mER-beta from Sf9 cells is Ser(16) near the N-terminus. Concomitant anal yses also documented the O-phosphorylation of mER-beta at Ser(16). MALDI-TO F mass spectrometry showed alternative occupancy of this locus by these two abundant and dynamic posttranslational modifications. The localization of a major O-GlcNAc/O-phosphate site in proximity of the transactivation domai n and as part of a PEST region (target sequences for rapid protein degradat ion) on mER-beta suggests that these modifications may play a role in regul ating estrogen receptor beta transactivation and turnover.