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Phosphoenolpyruvate Carboxykinase, a Key Enzyme That Controls Blood Glucose, Is a Target of Retinoic Acid Receptor-Related Orphan Receptor ?.

ABSTRACT: Phosphoenolpyruvate carboxykinase (PEPCK) catalyzes a committed and rate-limiting step in hepatic gluconeogenesis, and its activity is tightly regulated to maintain blood glucose levels within normal limits. PEPCK activity is primarily regulated through hormonal control of gene transcription. Transcription is additionally regulated via a cAMP response unit, which includes a cAMP response element and four binding sites for CCAAT/enhancer-binding protein (C/EBP). Notably, the cAMP response unit also contains a putative response element for retinoic acid receptor-related orphan receptor ? (ROR?). In this paper, we characterize the effect of the ROR? response element on cAMP-induced transcription. Electrophoresis mobility shift assay indicates that ROR? binds this response element in a sequence-specific manner. Furthermore, luciferase reporter assays indicate that ROR? interacts with C/EBP at the PEPCK promoter to synergistically enhance transcription. We also found that cAMP-induced transcription depends in part on ROR? and its response element. In addition, we show that suppression of ROR? by siRNA significantly decreased PEPCK transcription. Finally, we found that a ROR? antagonist inhibits hepatic gluconeogenesis in an in vitro glucose production assay. Taken together, the data strongly suggest that PEPCK is a direct ROR? target. These results define possible new roles for ROR? in hepatic gluconeogenesis.

SUBMITTER: Matsuoka H

PROVIDER: S-EPMC4575163 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Phosphoenolpyruvate Carboxykinase, a Key Enzyme That Controls Blood Glucose, Is a Target of Retinoic Acid Receptor-Related Orphan Receptor α.

Matsuoka Hiroshi H Shima Akiho A Kuramoto Daisuke D Kikumoto Daisuke D Matsui Takashi T Michihara Akihiro A

PloS one 20150918 9

Phosphoenolpyruvate carboxykinase (PEPCK) catalyzes a committed and rate-limiting step in hepatic gluconeogenesis, and its activity is tightly regulated to maintain blood glucose levels within normal limits. PEPCK activity is primarily regulated through hormonal control of gene transcription. Transcription is additionally regulated via a cAMP response unit, which includes a cAMP response element and four binding sites for CCAAT/enhancer-binding protein (C/EBP). Notably, the cAMP response unit al ...[more]

PMID: 26383638

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Project description:ObjectivesThyroid hormone (T3) and high glucose concentrations are critical components of β-cell maturation and function. In the present study, we asked whether T3 and glucose signaling pathways coordinately regulate transcription of genes important for β-cell function and proliferation.MethodsRNA-seq analysis was performed on cadaveric human islets from five different donors in response to low and high glucose concentrations and in the presence or absence of T3. Gene expression was also studies in sorted human β-cells, mouse islets and Ins-1 cells by RT-qPCR. Silencing of the thyroid hormone receptors (THR) was conducted using lentiviruses. Proliferation was assessed by ki67 immunostaining in primary human/mouse islets. Chromatin immunoprecipitation and proximity ligation assay were preformed to validate interactions of ChREBP and THR.ResultsWe found glucose-mediated expression of carbohydrate response element binding protein alpha and beta (ChREBPα and ChREBPβ) mRNAs and their target genes are highly dependent on T3 concentrations in rodent and human β-cells. In β-cells, T3 and glucose coordinately regulate the expression of ChREBPβ and PCK1 (phosphoenolpyruvate carboxykinase-1) among other important genes for β-cell maturation. Additionally, we show the thyroid hormone receptor (THR) and ChREBP interact, and their relative response elements are located near to each other on mutually responsive genes. In FACS-sorted adult human β-cells, we found that high concentrations of glucose and T3 induced the expression of PCK1. Next, we show that overexpression of Pck1 together with dimethyl malate (DMM), a substrate precursor, significantly increased β-cell proliferation in human islets. Finally, using a Cre-Lox approach, we demonstrated that ChREBPβ contributes to Pck1-dependent β-cell proliferation in mouse β-cells.ConclusionsWe conclude that T3 and glucose act together to regulate ChREBPβ, leading to increased expression and activity of Pck1, and ultimately increased β-cell proliferation.

Dataset Information

Phosphoenolpyruvate Carboxykinase, a Key Enzyme That Controls Blood Glucose, Is a Target of Retinoic Acid Receptor-Related Orphan Receptor ?.

Publications

Phosphoenolpyruvate Carboxykinase, a Key Enzyme That Controls Blood Glucose, Is a Target of Retinoic Acid Receptor-Related Orphan Receptor α.

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