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ChREBP Regulates Itself and Metabolic Genes Implicated in Lipid Accumulation in ?-Cell Line.

ABSTRACT: Carbohydrate response element binding protein (ChREBP) is an important transcription factor that regulates a variety of glucose-responsive genes in hepatocytes. To date, only two natural isoforms, Chrebp? and Chrebp?, have been identified. Although ChREBP is known to be expressed in pancreatic ? cells, most of the glucose-responsive genes have never been verified as ChREBP targets in this organ. We aimed to explore the impact of ChREBP expression on regulating genes linked to accumulation of lipid droplets, a typical feature of ?-cell glucotoxicity. We assessed gene expression in 832/13 cells overexpressing constitutively active ChREBP (caChREBP), truncated ChREBP with nearly identical amino acid sequence to Chrebp?, or dominant negative ChREBP (dnChREBP). Among multiple ChREBP-controlled genes, ChREBP was sufficient and necessary for regulation of Eno1, Pklr, Mdh1, Me1, Pdha1, Acly, Acaca, Fasn, Elovl6, Gpd1, Cpt1a, Rgs16, Mid1ip1,Txnip, and Chrebp?. Expression of Chrebp? and Srebp1c were not changed by caChREBP or dnChREBP. We identified functional ChREBP binding sequences that were located on the promoters of Chrebp? and Rgs16. We also showed that Rgs16 overexpression lead to increased considerable amounts of lipids in 832/13 cells. This phenotype was accompanied by reduction of Cpt1a expression and slight induction of Fasn and Pklr gene in these cells. In summary, we conclude that Chrebp? modulates its own expression, not that of Chrebp?; it also regulates the expression of several metabolic genes in ?-cells without affecting SREBP-1c dependent regulation. We also demonstrate that Rgs16 is one of the ChREBP-controlled genes that potentiate accumulation of lipid droplets in ?-cells.

SUBMITTER: Sae-Lee C

PROVIDER: S-EPMC4725739 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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ChREBP Regulates Itself and Metabolic Genes Implicated in Lipid Accumulation in β-Cell Line.

Sae-Lee Chanachai C Moolsuwan Kanya K Chan Lawrence L Poungvarin Naravat N

PloS one 20160125 1

Carbohydrate response element binding protein (ChREBP) is an important transcription factor that regulates a variety of glucose-responsive genes in hepatocytes. To date, only two natural isoforms, Chrebpα and Chrebpβ, have been identified. Although ChREBP is known to be expressed in pancreatic β cells, most of the glucose-responsive genes have never been verified as ChREBP targets in this organ. We aimed to explore the impact of ChREBP expression on regulating genes linked to accumulation of lip ...[more]

PMID: 26808438

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Project description:Methods: We performed mRNA sequence analysis by deep sequencing, in triplicate, using Illumina HiSeq to assess the impact of PHLPP1 knockdown in regulating gene expression in RAW 264.7 cells treated with/without OxLDL. The transcriptome libraries were constructed using the NEB adapters and were sequenced on at 150 nucleotide read length using the paired-end chemistry. The raw reads were subjected to Adapter and low-quality reads removal by Trimmomatic -0.36v. The raw reads were subjected to contamination [structural RNA / low complexity sequences] removal by mapping with bowtie 2-2.2.1. The decontaminated data set was mapped to the mouse genome. Reads mapping to gene list were counted using feature count module of sub reads package. The read counts were normalized in DESeq2-3.5, and subject to differential expression analysis. Differentially expressed genes were selected based on log2-ratio change with p-value <0.05 (Student’s t-test, unpaired). Hierarchical clustering was performed with the programs Cluster (uncentered correlation; average linkage clustering) and Treeview. Results: To gain a deeper understanding of the role of PHLPP1 in lipid metabolism in macrophages, we performed mRNA Seq analysis to assess the gene expression changes in a mouse macrophage cell line. The experiment has been carried out in triplicates. Alignment of the reads showed 97% alignment to the reference mouse genome. Identification of the differentially expressed genes were then carried out by DESeq2 version 1.2.10 with stringent criteria (FDR <0.05% with 2 fold up and down regulation) which reveals in control treated vs. control (121-down, 396-Up), Test vs. Control - (326-down, 569 Up) and Test treated vs. control treated (1314-down, 1328 up) of DEG to be altered in expression respectively. Inference: RNA-seq analysis revealed that PHLPP1 knockdown parallel with OxLDL treatment resulted in diminished expression profile of cholesterol biosynthesis and lipid metabolism genes.

Dataset Information

ChREBP Regulates Itself and Metabolic Genes Implicated in Lipid Accumulation in ?-Cell Line.

Publications

ChREBP Regulates Itself and Metabolic Genes Implicated in Lipid Accumulation in β-Cell Line.

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