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

Abstract

The hexosamine pathway provides UDPN-acetylhexosamine donor substrates used in cytosolic and Golgi-mediated glycosylation of proteins and for formation of glycosylphosphatidylinositol (GPI) anchors, which tether proteins to the outer plasma membrane. We have recently identified the murine glucosamine-6-phosphate (GIcN6P) acetyltransferase, EMeg32, as a developmentally regulated enzyme on the route to UDP-N-acetylglucosamine (UDPGIcNAc). Here we describe embryos and cells that have the EMeg32 gene inactivated by homologous recombination. Homozygous mutant embryos die at around embryonic day (E) 7.5 with a general proliferative delay of development. In vitro differentiated EMeg32-(/)- ES cells show reduced proliferation. Mouse embryonic fibroblasts (MEFs) deficient for EMeg32 exhibit defects in proliferation and adhesiveness, which could be complemented by stable re-expression of EMeg32 or by nutritional restoration of intracellular UDP-GIcNAc levels. Reduced UDP-GIcNAc levels predominantly translated into decreased O-GIcNAc modifications of cytosolic and nuclear proteins. Interestingly, growth-impaired EMeg32-(/)- MEFs withstand a number of apoptotic stimuli and express activated PKB/AKT. Thus, EMeg32-dependent UDP-GIcNAc levels influence cell cycle progression and susceptibility to apoptotic stimuli.