<i>Ogt</i>-Dependent X-Chromosome-Linked Protein Glycosylation Is a Requisite Modification in Somatic Cell Function and Embryo Viability

TLDR

Ogt encodes an O‑GlcNAc transferase that modifies nuclear and cytosolic proteins, but its essential role in embryonic stem cell viability has made mammalian Ogt deficiency studies difficult, requiring conditional mutagenesis. The authors used oocyte‑ and spermatid‑specific Cre‑loxP mutagenesis to segregate mutant Ogt alleles into parental gametes, creating an in vivo genetic system that enables study of female heterozygotes carrying X‑linked genes essential during embryogenesis. Loss of O‑GlcNAc in neurons, thymocytes, and fibroblasts caused tau hyperphosphorylation, T‑cell apoptosis, and fibroblast growth arrest with altered expression of c‑Fos, c‑Jun, c‑Myc, Sp1, and p27, demonstrating that mammalian cells require functional Ogt and that O‑GlcNAc regulates phosphorylation and expression in key signaling pathways.

Abstract

The Ogt gene encodes a glycosyltransferase that links N-acetylglucosamine to serine and threonine residues (O-GlcNAc) on nuclear and cytosolic proteins. Efforts to study a mammalian model of Ogt deficiency have been hindered by the requirement for this X-linked gene in embryonic stem cell viability, necessitating the use of conditional mutagenesis in vivo. We have extended these observations by segregating Ogt mutation to distinct somatic cell types, including neurons, thymocytes, and fibroblasts, the latter by an approach developed for inducible Ogt mutagenesis. We show that Ogt mutation results in the loss of O-GlcNAc and causes T-cell apoptosis, neuronal tau hyperphosphorylation, and fibroblast growth arrest with altered expression of c-Fos, c-Jun, c-Myc, Sp1, and p27. We further segregated the mutant Ogt allele to parental gametes by oocyte- and spermatid-specific Cre-loxP mutagenesis. By this we established an in vivo genetic approach that supports the ontogeny of female heterozygotes bearing mutant X-linked genes required during embryogenesis. Successful production and characterization of such female heterozygotes further indicates that mammalian cells commonly require a functional Ogt allele. We find that O-GlcNAc modulates protein phosphorylation and expression among essential and conserved cell signaling pathways.