Abstract
Background: O-GlcNAcylation is a posttranslational modification associated with various physiological and pathophysiological processes including diabetes, cancer, neurodegeneration and inflammation. However, the biological mechanisms underlying the role of specific O-GlcNAc sites and their link to phenotypes remain largely unexplored due to lack of suitable in vivo models. TGF-β activated kinase-1 binding protein-1 (TAB1) is a scaffolding protein required for TGF-β activated kinase-1 (TAK1) mediated signalling. A single O-GlcNAc site has been identified on human TAB1 that modulates TAK1-mediated cytokine release in cells. Methods: Here, we report the generation of the Tab1 S393A mouse model using a constitutive knock-in strategy. The Tab1 S393A mice carry a Ser393Ala (S393A) mutation that leads to loss of O-GlcNAcylation site on TAB1. Results: We did not observe any obvious phenotype in Tab1 S393A mice. Loss of O-GlcNAcylation on TAB1 has no consequences on TAB1 protein level or on TAB1-TAK1 interaction. Conclusions: The homozygous Tab1 S393A mice are viable and develop with no obvious abnormalities, providing a powerful tool to further investigate the role of O-GlcNAc on TAB1 in the inflammatory response in the context of a whole organism.
Highlights
O-GlcNAcylation is a dynamic and reversible post-translational modification on serine and threonine residues of intracellular proteins
The Ser393 site, equivalent to Ser395 in the human transforming growth factor (TGF)-β activated kinase-1 binding protein-1 (TAB1) sequence, was abolished by introducing a Ser393Ala mutation using a classical recombinational approach. We demonstrate that this mutation causes the loss of the O-GlcNAc modification on TAB1 without affecting TAB1 protein levels or its interaction with TGF-β activated kinase-1 (TAK1)
We found that TAB1 interacts with TAK1 in both WT and transgenic animals, suggesting that O-GlcNAcylation of TAB1 is not required for its interaction with TAK1 (Figure 3D)
Summary
O-GlcNAcylation is a dynamic and reversible post-translational modification on serine and threonine residues of intracellular proteins. It is catalysed by two enzymes, O-linked N-acetylglucosaminyltransferase (OGT) and β-N-acetylglucosaminidase (O-GlcNAcase, OGA), responsible for the addition and removal of the O-GlcNAc group, respectively (Hart et al, 2011). The TAK1-TAB1 complex has been found to be important in vivo for the survival of activated macrophages upon lipopolysaccharide (LPS) stimulation and for the modulation of immune response in T and B cells (Mihaly et al, 2014; Sato et al, 2005). A single O-GlcNAc site has been identified on TAB1 that modulates TAK1-mediated cytokine release in cells. Conclusions: The homozygous Tab1S393A mice are viable and develop with no obvious abnormalities, providing a powerful tool to further investigate the role of O-GlcNAc on TAB1 in the inflammatory response in the context of a whole organism
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