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The histone demethylase Jhdm1a regulates hepatic gluconeogenesis.

Pan D, Mao C, Zou T, Yao AY, Cooper MP, Boyartchuk V, Wang YX - PLoS Genet. (2012)

Bottom Line: In vivo, silencing of Jhdm1a promotes liver glucose synthesis, while its exogenous expression reduces blood glucose level.Importantly, the regulation of gluconeogenesis by Jhdm1a requires its demethylation activity.This is achieved, at least in part, by its USF1-dependent association with the C/EBPα promoter and its subsequent demethylation of dimethylated H3K36 on the C/EBPα locus.

View Article: PubMed Central - PubMed

Affiliation: Program in Gene Function and Expression and Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.

ABSTRACT
Hepatic gluconeogenesis is required for maintaining blood glucose homeostasis; yet, in diabetes mellitus, this process is unrestrained and is a major contributor to fasting hyperglycemia. To date, the impacts of chromatin modifying enzymes and chromatin landscape on gluconeogenesis are poorly understood. Through catalyzing the removal of methyl groups from specific lysine residues in the histone tail, histone demethylases modulate chromatin structure and, hence, gene expression. Here we perform an RNA interference screen against the known histone demethylases and identify a histone H3 lysine 36 (H3K36) demethylase, Jhdm1a, as a key negative regulator of gluconeogenic gene expression. In vivo, silencing of Jhdm1a promotes liver glucose synthesis, while its exogenous expression reduces blood glucose level. Importantly, the regulation of gluconeogenesis by Jhdm1a requires its demethylation activity. Mechanistically, we find that Jhdm1a regulates the expression of a major gluconeogenic regulator, C/EBPα. This is achieved, at least in part, by its USF1-dependent association with the C/EBPα promoter and its subsequent demethylation of dimethylated H3K36 on the C/EBPα locus. Our work provides compelling evidence that links histone demethylation to transcriptional regulation of gluconeogenesis and has important implications for the treatment of diabetes.

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Related in: MedlinePlus

Jhdm1a regulates gluconeogenesis in vivo.(A) Jhdm1a knockdown or scramble adenoviruses were transduced into the liver of wild-type male C57BL/6J mice (n = 5 per group). Mice fed ad libitum were sacrificed at Day 5 after viral infusion. (Left) mRNA levels of PEPCK, G6Pase and Jhdm1a in the liver were measured and normalized to U36b4. **, P<0.005. (Right) PEPCK and G6Pase protein. (B) Blood insulin levels at fed state were measured at Day 5. (C) Jhdm1a knockdown or scramble adenoviruses were transduced into the liver of wild-type male C57BL/6J mice (n = 10 per group). At Day 5, mice were i.p. injected with pyruvate (2 g/kg body weight) after a starvation for 16 hr and blood glucose levels were measured. *, P<0.05. (D and E) Adenoviruses expressing wild-type Jhdm1a, H212A point mutant, or GFP were transduced into the liver of male ob/ob mice (n = 5 per group). Gene expression was measured on Day 5 and blood glucose levels were measured on Day 3 after a 5-hr fasting. Changes of blood glucose level relative to Day 0 are presented. *, P<0.03; **, P<0.01. (F) Hepatic Jhdm1a mRNA levels in male C57BL/6J mice (n = 5 per group) fed ad libitum, or fasted for 5 hr or 20 hr. (G) Male C57BL/6J mice (n = 4) were i.p. injected with glucagon (300 µg/kg), insulin (0.75 U/Kg), or PBS. Hepatic Jhdm1a mRNA levels were examined 6 hr after injection. (H) Hepatic Jhdm1a mRNA levels in lean mice and diabetic ob/ob mice (n = 3 per group).
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pgen-1002761-g003: Jhdm1a regulates gluconeogenesis in vivo.(A) Jhdm1a knockdown or scramble adenoviruses were transduced into the liver of wild-type male C57BL/6J mice (n = 5 per group). Mice fed ad libitum were sacrificed at Day 5 after viral infusion. (Left) mRNA levels of PEPCK, G6Pase and Jhdm1a in the liver were measured and normalized to U36b4. **, P<0.005. (Right) PEPCK and G6Pase protein. (B) Blood insulin levels at fed state were measured at Day 5. (C) Jhdm1a knockdown or scramble adenoviruses were transduced into the liver of wild-type male C57BL/6J mice (n = 10 per group). At Day 5, mice were i.p. injected with pyruvate (2 g/kg body weight) after a starvation for 16 hr and blood glucose levels were measured. *, P<0.05. (D and E) Adenoviruses expressing wild-type Jhdm1a, H212A point mutant, or GFP were transduced into the liver of male ob/ob mice (n = 5 per group). Gene expression was measured on Day 5 and blood glucose levels were measured on Day 3 after a 5-hr fasting. Changes of blood glucose level relative to Day 0 are presented. *, P<0.03; **, P<0.01. (F) Hepatic Jhdm1a mRNA levels in male C57BL/6J mice (n = 5 per group) fed ad libitum, or fasted for 5 hr or 20 hr. (G) Male C57BL/6J mice (n = 4) were i.p. injected with glucagon (300 µg/kg), insulin (0.75 U/Kg), or PBS. Hepatic Jhdm1a mRNA levels were examined 6 hr after injection. (H) Hepatic Jhdm1a mRNA levels in lean mice and diabetic ob/ob mice (n = 3 per group).

Mentions: Based on our discovery of the regulatory role of Jhdm1a in vitro, we tested whether Jhdm1a regulates gluconeogenesis in live animals. We obtained five lentiviral Jhdm1a knockdown constructs from Open Biosystems and tested their knockdown efficiency by RT-QPCR in mouse cell culture. We transferred two best ones into an adenoviral vector, generated adenoviruses, and further confirmed that they were able to reduce ectopically expressed Jhdm1a protein level in vitro (Figure S5). The viruses were infused into the liver of wild-type C57BL/6J mice via tail vein injection and endogenous Jhdm1a expression was decreased, which led to a significantly increase in hepatic expression of PEPCK and G6Pase in both fed and fasting states, compared with the scramble control (Figure 3A, Figure S6 and S7). A corresponding enhanced PEPCK and G6Pase protein production was observed (Figure 3A). Blood insulin levels examined at fed state were not significantly different (Figure 3B). Although the Jhdm1a knockdown mice were still able to maintain normal glycemia, they displayed higher glucose production upon injection of the gluconeogenic substrate pyruvate (Figure 3C). We next ectopically expressed either wild-type Jhdm1a or the H212A point mutant in the liver of diabetic ob/ob mice. Expression of the wild-type Jhdm1a, but not the H212A point mutant, decreased the expression of PEPCK and G6Pase (Figure 3D). Accordingly, we observed a statistically significant reduction of blood glucose level in ob/ob mice expressing wild-type Jhdm1a (Figure 3E). Thus, Jhdm1a indeed has a physiological role in hepatic gluconeogenesis in vivo, and this role is mediated by its histone demethylation activity.


The histone demethylase Jhdm1a regulates hepatic gluconeogenesis.

Pan D, Mao C, Zou T, Yao AY, Cooper MP, Boyartchuk V, Wang YX - PLoS Genet. (2012)

Jhdm1a regulates gluconeogenesis in vivo.(A) Jhdm1a knockdown or scramble adenoviruses were transduced into the liver of wild-type male C57BL/6J mice (n = 5 per group). Mice fed ad libitum were sacrificed at Day 5 after viral infusion. (Left) mRNA levels of PEPCK, G6Pase and Jhdm1a in the liver were measured and normalized to U36b4. **, P<0.005. (Right) PEPCK and G6Pase protein. (B) Blood insulin levels at fed state were measured at Day 5. (C) Jhdm1a knockdown or scramble adenoviruses were transduced into the liver of wild-type male C57BL/6J mice (n = 10 per group). At Day 5, mice were i.p. injected with pyruvate (2 g/kg body weight) after a starvation for 16 hr and blood glucose levels were measured. *, P<0.05. (D and E) Adenoviruses expressing wild-type Jhdm1a, H212A point mutant, or GFP were transduced into the liver of male ob/ob mice (n = 5 per group). Gene expression was measured on Day 5 and blood glucose levels were measured on Day 3 after a 5-hr fasting. Changes of blood glucose level relative to Day 0 are presented. *, P<0.03; **, P<0.01. (F) Hepatic Jhdm1a mRNA levels in male C57BL/6J mice (n = 5 per group) fed ad libitum, or fasted for 5 hr or 20 hr. (G) Male C57BL/6J mice (n = 4) were i.p. injected with glucagon (300 µg/kg), insulin (0.75 U/Kg), or PBS. Hepatic Jhdm1a mRNA levels were examined 6 hr after injection. (H) Hepatic Jhdm1a mRNA levels in lean mice and diabetic ob/ob mice (n = 3 per group).
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3375226&req=5

pgen-1002761-g003: Jhdm1a regulates gluconeogenesis in vivo.(A) Jhdm1a knockdown or scramble adenoviruses were transduced into the liver of wild-type male C57BL/6J mice (n = 5 per group). Mice fed ad libitum were sacrificed at Day 5 after viral infusion. (Left) mRNA levels of PEPCK, G6Pase and Jhdm1a in the liver were measured and normalized to U36b4. **, P<0.005. (Right) PEPCK and G6Pase protein. (B) Blood insulin levels at fed state were measured at Day 5. (C) Jhdm1a knockdown or scramble adenoviruses were transduced into the liver of wild-type male C57BL/6J mice (n = 10 per group). At Day 5, mice were i.p. injected with pyruvate (2 g/kg body weight) after a starvation for 16 hr and blood glucose levels were measured. *, P<0.05. (D and E) Adenoviruses expressing wild-type Jhdm1a, H212A point mutant, or GFP were transduced into the liver of male ob/ob mice (n = 5 per group). Gene expression was measured on Day 5 and blood glucose levels were measured on Day 3 after a 5-hr fasting. Changes of blood glucose level relative to Day 0 are presented. *, P<0.03; **, P<0.01. (F) Hepatic Jhdm1a mRNA levels in male C57BL/6J mice (n = 5 per group) fed ad libitum, or fasted for 5 hr or 20 hr. (G) Male C57BL/6J mice (n = 4) were i.p. injected with glucagon (300 µg/kg), insulin (0.75 U/Kg), or PBS. Hepatic Jhdm1a mRNA levels were examined 6 hr after injection. (H) Hepatic Jhdm1a mRNA levels in lean mice and diabetic ob/ob mice (n = 3 per group).
Mentions: Based on our discovery of the regulatory role of Jhdm1a in vitro, we tested whether Jhdm1a regulates gluconeogenesis in live animals. We obtained five lentiviral Jhdm1a knockdown constructs from Open Biosystems and tested their knockdown efficiency by RT-QPCR in mouse cell culture. We transferred two best ones into an adenoviral vector, generated adenoviruses, and further confirmed that they were able to reduce ectopically expressed Jhdm1a protein level in vitro (Figure S5). The viruses were infused into the liver of wild-type C57BL/6J mice via tail vein injection and endogenous Jhdm1a expression was decreased, which led to a significantly increase in hepatic expression of PEPCK and G6Pase in both fed and fasting states, compared with the scramble control (Figure 3A, Figure S6 and S7). A corresponding enhanced PEPCK and G6Pase protein production was observed (Figure 3A). Blood insulin levels examined at fed state were not significantly different (Figure 3B). Although the Jhdm1a knockdown mice were still able to maintain normal glycemia, they displayed higher glucose production upon injection of the gluconeogenic substrate pyruvate (Figure 3C). We next ectopically expressed either wild-type Jhdm1a or the H212A point mutant in the liver of diabetic ob/ob mice. Expression of the wild-type Jhdm1a, but not the H212A point mutant, decreased the expression of PEPCK and G6Pase (Figure 3D). Accordingly, we observed a statistically significant reduction of blood glucose level in ob/ob mice expressing wild-type Jhdm1a (Figure 3E). Thus, Jhdm1a indeed has a physiological role in hepatic gluconeogenesis in vivo, and this role is mediated by its histone demethylation activity.

Bottom Line: In vivo, silencing of Jhdm1a promotes liver glucose synthesis, while its exogenous expression reduces blood glucose level.Importantly, the regulation of gluconeogenesis by Jhdm1a requires its demethylation activity.This is achieved, at least in part, by its USF1-dependent association with the C/EBPα promoter and its subsequent demethylation of dimethylated H3K36 on the C/EBPα locus.

View Article: PubMed Central - PubMed

Affiliation: Program in Gene Function and Expression and Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.

ABSTRACT
Hepatic gluconeogenesis is required for maintaining blood glucose homeostasis; yet, in diabetes mellitus, this process is unrestrained and is a major contributor to fasting hyperglycemia. To date, the impacts of chromatin modifying enzymes and chromatin landscape on gluconeogenesis are poorly understood. Through catalyzing the removal of methyl groups from specific lysine residues in the histone tail, histone demethylases modulate chromatin structure and, hence, gene expression. Here we perform an RNA interference screen against the known histone demethylases and identify a histone H3 lysine 36 (H3K36) demethylase, Jhdm1a, as a key negative regulator of gluconeogenic gene expression. In vivo, silencing of Jhdm1a promotes liver glucose synthesis, while its exogenous expression reduces blood glucose level. Importantly, the regulation of gluconeogenesis by Jhdm1a requires its demethylation activity. Mechanistically, we find that Jhdm1a regulates the expression of a major gluconeogenic regulator, C/EBPα. This is achieved, at least in part, by its USF1-dependent association with the C/EBPα promoter and its subsequent demethylation of dimethylated H3K36 on the C/EBPα locus. Our work provides compelling evidence that links histone demethylation to transcriptional regulation of gluconeogenesis and has important implications for the treatment of diabetes.

Show MeSH
Related in: MedlinePlus