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The orphan nuclear receptor Nor1/Nr4a3 is a negative regulator of ?-cell mass.

ABSTRACT: The Nr4a subfamily of nuclear receptor comprises three members in mammalian cells: Nur77/Nr4a1, Nurr1/Nr4a2, and Nor1/Nr4a3. Nr4a proteins play key roles in the regulation of glucose homeostasis in peripheral metabolic tissues. However, their biological functions in ?-cells remain relatively uncharacterized. Here we sought to investigate the potential role of Nor1 in the regulation of ?-cell mass and, in particular, ?-cell survival/apoptosis. We used histological analysis to examine the consequences of genetic deletion of either Nur77 and Nor1 on ?-cell mass, investigated the expression patterns of Nr4as in human islets and INS cells and performed gain- and loss-of-function experiments to further characterize the role of Nor1 in ?-cell apoptosis. Surprisingly, Nor1 knockout mice displayed increased ?-cell mass, whereas mice with genetic deletion of Nur77 did not exhibit any significant differences compared with their WT littermates. The increase in ?-cell mass in Nor1 knockout mice was accompanied by improved glucose tolerance. A gene expression study performed in both human islets and INS cells revealed that Nor1 expression is significantly increased by pro-inflammatory cytokines and, to a lesser extent, by elevated concentrations of glucose. Nor1 overexpression in both INS and human islet cells caused apoptosis, whereas siRNA-mediated Nor1 knockdown prevented cytokine-induced ?-cell death. Finally, Nor1 expression was up-regulated in islets of individuals with type 2 diabetes. Altogether, our results uncover that Nor1 negatively regulates ?-cell mass. Nor1 represents a promising molecular target in diabetes treatment to prevent ?-cell destruction.

SUBMITTER: Close AF

PROVIDER: S-EPMC6442030 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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The orphan nuclear receptor Nor1/Nr4a3 is a negative regulator of β-cell mass.

Close Anne-Françoise AF Dadheech Nidheesh N Villela Bárbara Scoralick BS Rouillard Claude C Buteau Jean J

The Journal of biological chemistry 20190129 13

The Nr4a subfamily of nuclear receptor comprises three members in mammalian cells: Nur77/Nr4a1, Nurr1/Nr4a2, and Nor1/Nr4a3. Nr4a proteins play key roles in the regulation of glucose homeostasis in peripheral metabolic tissues. However, their biological functions in β-cells remain relatively uncharacterized. Here we sought to investigate the potential role of Nor1 in the regulation of β-cell mass and, in particular, β-cell survival/apoptosis. We used histological analysis to examine the conseque ...[more]

PMID: 30696767

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Project description:BackgroundNor1/NR4A3 is a member of the NR4A subfamily of nuclear receptors that play essential roles in regulating gene expression related to development, cell homeostasis and neurological functions. However, Nor1 is still considered an orphan receptor, as its natural ligand remains unclear for mediating transcriptional activation. Yet other activation signals may modulate Nor1 activity, although their precise role in the development and maintenance of the nervous system remains elusive.MethodsWe used transcriptional reporter assays, gene expression profiling, protein turnover measurement, and cell growth assays to assess the functional relevance of Nor1 and SUMO-defective variants in neuronal cells. SUMO1 and SUMO2 conjugation to Nor1 were assessed by immunoprecipitation. Tubulin stability was determined by acetylation and polymerization assays, and live-cell fluorescent microscopy.ResultsHere, we demonstrate that Nor1 undergoes SUMO1 conjugation at Lys-89 within a canonical ψKxE SUMOylation motif, contributing to the complex pattern of Nor1 SUMOylation, which also includes Lys-137. Disruption of Lys-89, thereby preventing SUMO1 conjugation, led to reduced Nor1 transcriptional competence and protein stability, as well as the downregulation of genes involved in cell growth and metabolism, such as ENO3, EN1, and CFLAR, and in microtubule cytoskeleton dynamics, including MAP2 and MAPT, which resulted in reduced survival of neuronal cells. Interestingly, Lys-89 SUMOylation was potentiated in response to nocodazole, a microtubule depolymerizing drug, although this was insufficient to rescue cells from microtubule disruption despite enhanced Nor1 gene expression. Instead, Lys-89 deSUMOylation reduced the expression of microtubule-severing genes like KATNA1, SPAST, and FIGN, and enhanced α-tubulin cellular levels, acetylation, and microfilament organization, promoting microtubule stability and resistance to nocodazole. These effects contrasted with Lys-137 SUMOylation, suggesting distinct regulatory mechanisms based on specific Nor1 input SUMOylation signals.ConclusionsOur study provides novel insights into Nor1 transcriptional signaling competence and identifies a hierarchical mechanism whereby selective Nor1 SUMOylation may govern neuronal cytoskeleton network dynamics and resistance against microtubule disturbances, a condition strongly associated with neurodegenerative diseases.

Project description:BackgroundNeuron-derived orphan receptor (Nor) 1, nuclear receptor (Nur) 77, and nuclear receptor-related protein (Nurr) 1 constitute the NR4A family of orphan nuclear receptors which were recently found to modulate hepatic glucose production, insulin signalling in adipocytes, and oxidative metabolism in skeletal muscle. In this study, we assessed whether common genetic variation within the NR4A3 locus, encoding Nor-1, contributes to the development of prediabetic phenotypes, such as glucose intolerance, insulin resistance, or beta-cell dysfunction.MethodsWe genotyped 1495 non-diabetic subjects from Southern Germany for the five tagging single nucleotide polymorphisms (SNPs) rs7047636, rs1526267, rs2416879, rs12686676, and rs10819699 (minor allele frequencies >or= 0.05) covering 100% of genetic variation within the NR4A3 locus (with D' = 1.0, r2 >or= 0.9) and assessed their association with metabolic data derived from the fasting state, an oral glucose tolerance test (OGTT), and a hyperinsulinemic-euglycemic clamp (subgroup, N = 506). SNPs that revealed consistent associations with prediabetic phenotypes were subsequently genotyped in a second cohort (METSIM Study; Finland; N = 5265) for replication.ResultsAll five SNPs were in Hardy-Weinberg equilibrium (p >or= 0.7, all). The minor alleles of three SNPs, i.e., rs1526267, rs12686676, and rs10819699, consistently tended to associate with higher insulin release as derived from plasma insulin at 30 min(OGTT), AUCC C-peptide-to-AUC Gluc ratio and the AUC Ins30-to-AUC Gluc30 ratio with rs12686676 reaching the level of significance (p <or= 0.03, all; additive model). The association of the SNP rs12686676 with insulin secretion was replicated in the METSIM cohort (p <or= 0.03, additive model). There was no consistent association with glucose tolerance or insulin resistance in both study cohorts.ConclusionWe conclude that common genetic variation within the NR4A3 locus determines insulin secretion. Thus, NR4A3 represents a novel candidate gene for beta-cell function which was not covered by the SNP arrays of recent genome-wide association studies for type 2 diabetes mellitus.

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The orphan nuclear receptor Nor1/Nr4a3 is a negative regulator of ?-cell mass.

Publications

The orphan nuclear receptor Nor1/Nr4a3 is a negative regulator of β-cell mass.

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