Decreased UDP-GlcNAc levels abrogate proliferation control in EMeg32-deficient cells

The hexosamine pathway provides UDP-N-acetylhexosamine donor substrates use d in cytosolic and Golgi-mediated glycosylation of proteins and for formati on of glycosylphosphatidylinositol (GPT) anchors, which tether proteins to the outer plasma membrane, We have recently identified the murine glucosami ne-6-phosphate (GlcN6P) acetyltransferase, EMeg32, as a developmentally reg ulated enzyme on the route to UDP-N-acetylglucosamine (UDP-GlcNAc), Here we describe embryos and cells that have the EMeg32 gene inactivated by homolo gous recombination, Homozygous mutant embryos die at around embryonic day ( E) 7.5 with a general proliferative delay of development. In vitro differen tiated EMeg32(-/-) ES cells show reduced proliferation. Mouse embryonic fib roblasts (MEFs) deficient for EMeg32 exhibit defects in proliferation and a dhesiveness, which could be complemented by stable re-expression of EMeg32 or by nutritional restoration of intracellular UDP-GlcNAc levels. Reduced U DP-GlcNAc levels predominantly translated into decreased O-GlcNAc modificat ions of cytosolic and nuclear proteins, Interestingly, growth-impaired EMeg 32(-/-)MEFs withstand a number of apoptotic stimuli and express activated P KB/AKT, Thus, EMeg32-dependent UDP-GlcNAc levels influence cell cycle progr ession and susceptibility to apoptotic stimuli.