O-GlcNAc modification of proteins affects volume regulation in Jurkat cells

Abstract

An increasing amount of recent research has demonstrated that the hexosamine biosynthesis pathway (HBP) plays a significant role in the modulation of intracellular signaling transduction pathways, and affects cellular processes via modification of protein by O-linked beta-N-acetylglucosamine (O-GlcNAc). Besides the many known and postulated effects of protein O-GlcNAc modifications, there is little available data on the role of O-GlcNAc in cellular volume regulation. Our objective was to test the effect of increased O-GlcNAc levels on hypotonia-induced volume changes in Jurkat cells. We pretreated Jurkat cells for 1 h with glucosamine (GlcN), PUGNAc (O-(2-acetamido-2-deoxy-D-glucopyranosylidene)-amino-N-phenylcarbamate) an inhibitor of O-GlcNAcase, or a high level of glucose to induce elevated levels of O-GlcNAc. We found that the response of Jurkat cells to hypotonic stress was significantly altered. The hypotonia induced cell-swelling was augmented in both GlcN and PUGNAc-treated cells and, to a lesser extent, in high glucose concentration-treated cells. Evaluated by NMR measurements, GlcN and PUGNAc treatment also significantly reduced intracellular water diffusion. Taken together, increased cell swelling and reduced water diffusion caused by elevated O-GlcNAc show notable analogy to the regulatory volume changes seen by magnetic resonance methods in nervous and other tissues in different pathological states. In conclusion, we demonstrate for the first time that protein O-GlcNAc could modulate cell volume regulation.

MicroRNA-15b Suppresses Th17 Differentiation and Is Associated with Pathogenesis of Multiple Sclerosis by Targeting O-GlcNAc Transferase

“…In T lymphocytes, O-GlcNAc modification is dramatically increased after activation by mitogen (63), whereas active OGT is requisite for activating TCRs (28,64), and T cell-specific deletion of OGT leads to a reduction in the total T cell number due to increased apoptosis (64,65). Alteration of O-GlcNAc modification is also a component of T cell expression of cytokines and regulation of the cell volume (66,67). Later studies put a strong emphasis on the relationship between OGT-mediated protein O-GlcNAcylation and T cell functions (68).…”

Cell-Penetrant, Nanomolar O-GlcNAcase Inhibitors Selective against Lysosomal Hexosaminidases

“…A number of hOGA inhibitors have been described and used to demonstrate the involvement of O -GlcNAc in the insulin response (Akimoto et al., 2007), regulation of Tau phosphorylation (Yuzwa et al., 2008), Akt activation (Vosseller et al., 2002), and, recently, in regulation of cellular volume (Nagy et al., 2009). However, many of these inhibitors possess limited selectivity over the structurally related lysosomal hexosaminidases A/B (HexA/B) and/or are only active on cells in the high micromolar range, increasing the risk of off-target effects, as suggested recently by Macauley et al.…”

Lithium Induces ER Stress and N-Glycan Modification in Galactose-Grown Jurkat Cells

“…We have previously found that Jurkat cell extracts exhibit Olinked N-acetylglucosamine (O-GlcNAc) positivity when detected by CTD110.6, an anti-O-GlcNAc antibody [24]. Galactose treatment alone did not significantly increase the amount of positive proteins in Jurkat cells ( Figure 6 and Figure S1B).…”