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Glucose mediates the translocation of NeuroD1 by O-linked glycosylation.

ABSTRACT: O-Linked GlcNAc modification of nuclear and cytosolic proteins has been shown to regulate the function of many cellular proteins. Increased O-linked glycosylation, observed under chronic hyperglycemia conditions, has been implicated in the pathogenesis of diabetes. However, the exact role of O-GlcNAc modification in regulating glucose homeostasis remains to be established. We report here that the subcellular localization of the pancreatic beta cell-specific transcription factor NeuroD1 is regulated by O-linked glycosylation in the mouse insulinoma cell line MIN6. Under low glucose conditions, NeuroD1 is mainly in the cytosol. However, treatment of MIN6 cells with high glucose results in O-linked GlcNAc modification of NeuroD1 and its subsequent translocation into the nucleus. Consistent with these data, treatment of MIN6 cells with O-(2-acetamido-2-deoxy-d-glucopyranosylidene)-amino N-phenylcarbamate, an inhibitor of O-GlcNAcase, causes Neuro-D1 localization to the nucleus and induction of insulin gene expression even on low glucose. Furthermore, we demonstrate that NeuroD1 interacts with the O-GlcNAc transferase, OGT only at high concentrations of glucose and depletion of OGT by using small interfering RNA oligos interferes with the nuclear localization of NeuroD1 on high glucose. On low glucose NeuroD1 interacts with the O-GlcNAcase and becomes deglycosylated, which is likely to be important for export of Neuro-D1 into cytosol in the presence of low glucose. In summary, the presented data suggest that glucose regulates the subcellular localization of NeuroD1 in pancreatic beta cells via O-linked GlcNAc modification of NeuroD1 by OGT.

SUBMITTER: Andrali SS

PROVIDER: S-EPMC2096475 | biostudies-literature | 2007 May

REPOSITORIES: biostudies-literature

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Glucose mediates the translocation of NeuroD1 by O-linked glycosylation.

Andrali Sreenath S SS Qian Qingwen Q Ozcan Sabire S

The Journal of biological chemistry 20070402 21

O-Linked GlcNAc modification of nuclear and cytosolic proteins has been shown to regulate the function of many cellular proteins. Increased O-linked glycosylation, observed under chronic hyperglycemia conditions, has been implicated in the pathogenesis of diabetes. However, the exact role of O-GlcNAc modification in regulating glucose homeostasis remains to be established. We report here that the subcellular localization of the pancreatic beta cell-specific transcription factor NeuroD1 is regula ...[more]

PMID: 17403669

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Project description:Zika virus (ZIKV) is an emerging mosquito-borne flavivirus. Recent ZIKV outbreaks have produced serious human disease, including neurodevelopmental malformations (congenital Zika syndrome) and Guillain-Barré syndrome. These outcomes were not associated with ZIKV infection prior to 2013, raising the possibility that viral genetic changes could contribute to new clinical manifestations. All contemporary ZIKV isolates encode an N-linked glycosylation site in the envelope (E) protein (N154), but this glycosylation site is absent in many historical ZIKV isolates. Here, we investigated the role of E protein glycosylation in ZIKV pathogenesis using two contemporary Asian-lineage strains (H/PF/2013 and PRVABC59) and the historical African-lineage strain (MR766). We found that glycosylated viruses were highly pathogenic in Ifnar1-/- mice. In contrast, nonglycosylated viruses were attenuated, producing lower viral loads in the serum and brain when inoculated subcutaneously but remaining neurovirulent when inoculated intracranially. These results suggest that E glycosylation is advantageous in the periphery but not within the brain. Accordingly, we found that glycosylation facilitated infection of cells expressing the lectins dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN) or DC-SIGN-related (DC-SIGNR), suggesting that inefficient infection of lectin-expressing leukocytes could contribute to the attenuation of nonglycosylated ZIKV in mice.IMPORTANCE It is unclear why the ability of Zika virus (ZIKV) to cause serious disease, including Guillain-Barré syndrome and birth defects, was not recognized until recent outbreaks. One contributing factor could be genetic differences between contemporary ZIKV strains and historical ZIKV strains. All isolates from recent outbreaks encode a viral envelope protein that is glycosylated, whereas many historical ZIKV strains lack this glycosylation. We generated nonglycosylated ZIKV mutants from contemporary and historical strains and evaluated their virulence in mice. We found that nonglycosylated viruses were attenuated and produced lower viral loads in serum and brains. Our studies suggest that envelope protein glycosylation contributes to ZIKV pathogenesis, possibly by facilitating attachment to and infection of lectin-expressing leukocytes.

Project description:The organization and clustering of cell surface proteins plays a critical role in controlling receptor signaling; however, the biophysical mechanisms regulating these parameters are not well understood. Elucidating these mechanisms is highly significant to our understanding of immune function in health and disease, given the importance of B cell receptor (BCR) signaling in directing B cells to produce antibodies for the clearance of pathogens, and the potential deleterious effects of dysregulated BCR signaling, such as in B cell malignancies or autoimmune disease. One of main inhibitory co-receptors on B cells is CD22, a sialic-acid binding protein, which interacts homotypically with other sialylated CD22 molecules, as well as heterotypically with IgM and CD45. Although the importance of CD22 in attenuating BCR signaling is well established, we still do not fully understand what mediates CD22 organization and association to BCRs. CD22 is highly glycosylated, containing 12 N-linked glycosylation sites on its extracellular domain, the function of which remain to be resolved. We were interested in how these glycosylation sites mediate homotypic vs. heterotypic interactions. To this end, we mutated five out of the six N-linked glycosylation residues on CD22 localized closest to the sialic acid binding site. Glycan site N101 was not mutated as this resulted in lack of CD22 expression. We used dual-color super-resolution imaging to investigate the impact of altered glycosylation of CD22 on the nanoscale organization of CD22 and its association with BCR. We show that mutation of these five glycosylation sites increased the clustering tendency of CD22 and resulted in higher density CD22 nanoclusters. Consistent with these findings of altered CD22 organization, we found that mutation of N-glycan sites attenuated CD22 phosphorylation upon BCR stimulation, and consequently, increased BCR signaling. Importantly, we identified that these sites may be ligands for the soluble secreted lectin, galectin-9, and are necessary for galectin-9 mediated inhibition of BCR signaling. Taken together, these findings implicate N-linked glycosylation in the organization and function of CD22, likely through regulating heterotypic interactions between CD22 and its binding partners.

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Glucose mediates the translocation of NeuroD1 by O-linked glycosylation.

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Glucose mediates the translocation of NeuroD1 by O-linked glycosylation.

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