O-GlcNAcylation: A key regulator of hepatic proliferation and differentiation
O-GlcNAcylation is a posttranslational modification involved in various physiological processes, including development, regeneration, inflammation, proliferation, metabolism, and pathogenesis of various diseases. O-GlcNAcylation is regulated by two enzymes, O-GlcNAc transferase (OGT), which adds a glucosamine sugar on serine or threonine residues of proteins, and O-GlcNAcase (OGA), which removes the sugar molecule. The process of O-GlcNAcylation plays a role in regulating the cellular fate of various cell types, including hematopoietic stem cells and preadipocytes. In these studies, we demonstrated that O-GlcNAcylation regulates hepatic differentiation and liver regeneration and protects against the development of hepatocellular carcinoma (HCC). To investigate the role of O-GlcNAcylation in liver pathobiology, we studied mice with knockout (KO) of OGT or OGA specifically in hepatocytes, which led to a significant decrease or increase in hepatic O-GlcNAcylation, respectively. Our studies, aided by single-cell RNA sequencing, revealed that a decrease in hepatocyte O-GlcNAcylation resulted in loss of hepatic differentiation, specifically in the periportal hepatocytes and disruption of liver zonation. The lack of hepatic differentiation in OGT-KO mice was accompanied by cell death, increased inflammation, and the initiation of fibrosis. Further, we found that while OGA-KO mice had normal liver regeneration, OGT-KO mice exhibited defects in the termination of liver regeneration, resulting in sustained cell proliferation, significant hepatomegaly, hepatic dysplasia, and the appearance of small dysplastic nodules 28 days after PHX. These mice also showed sustained increases in the expression of cyclins, along with a significant induction of pro-inflammatory and pro-fibrotic gene expression in the liver. RNA-Seq studies revealed the inactivation of HNF4α, a known termination signal and master regulator of hepatic differentiation, in OGT-KO mice 28 days after PHX. This lack of hepatocyte differentiation was confirmed by a significant decrease in HNFα target genes in OGT-KO mice. Immunoprecipitation experiments also revealed that HNF4α is O-GlcNAcylated in normal differentiated hepatocytes. To investigate the effect of O-GlcNAcylation modulation in chronic liver disease, we used two models including the choline-deficient ethionine supplemented (CDE) diet model and the DEN-induced HCC model. OGT-KO mice challenged with the CDE diet showed enhanced biliary epithelial cell proliferation with increased hedgehog signaling, while OGT-knockout mice exacerbated DEN-induced HCC development with increased YAP signaling. In contrast, increasing hepatic O-GlcNAcylation inhibited DEN-induced HCC. In summary, our studies demonstrate that O-GlcNAcylation is a critical regulator of hepatic differentiation, which influences hepatocyte proliferation, and a potential therapeutic target in chronic liver diseases, including HCC.
Toxicology, CDE Diet, HCC, HNF4alpha, Liver Regeneration, O-GlcNAc, Single-Cell RNA-Sequencing
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