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Deficiency of Capicua disrupts bile acid homeostasis.

Kim E, Park S, Choi N, Lee J, Yoe J, Kim S, Jung HY, Kim KT, Kang H, Fryer JD, Zoghbi HY, Hwang D, Lee Y - Sci Rep (2015)

Bottom Line: We also found that levels of proinflammatory cytokine genes were up-regulated in Cic-L(-/-) liver.Moreover, induction of tumor necrosis factor alpha (Tnfα) expression and decrease in the levels of FOXA2, C/EBPβ, and RXRα were found in Cic-L(-/-) liver before BA was accumulated, suggesting that inflammation might be the cause for the cholestasis in Cic-L(-/-) mice.Our findings indicate that CIC is a critical regulator of BA homeostasis, and that its dysfunction might be associated with chronic liver disease and metabolic disorders.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, Pohang University of Science and Technology, Pohang, Gyeongbuk 790-784, Republic of Korea.

ABSTRACT
Capicua (CIC) has been implicated in pathogenesis of spinocerebellar ataxia type 1 and cancer in mammals; however, the in vivo physiological functions of CIC remain largely unknown. Here we show that Cic hypomorphic (Cic-L(-/-)) mice have impaired bile acid (BA) homeostasis associated with induction of proinflammatory cytokines. We discovered that several drug metabolism and BA transporter genes were down-regulated in Cic-L(-/-) liver, and that BA was increased in the liver and serum whereas bile was decreased within the gallbladder of Cic-L(-/-) mice. We also found that levels of proinflammatory cytokine genes were up-regulated in Cic-L(-/-) liver. Consistent with this finding, levels of hepatic transcriptional regulators, such as hepatic nuclear factor 1 alpha (HNF1α), CCAAT/enhancer-binding protein beta (C/EBPβ), forkhead box protein A2 (FOXA2), and retinoid X receptor alpha (RXRα), were markedly decreased in Cic-L(-/-) mice. Moreover, induction of tumor necrosis factor alpha (Tnfα) expression and decrease in the levels of FOXA2, C/EBPβ, and RXRα were found in Cic-L(-/-) liver before BA was accumulated, suggesting that inflammation might be the cause for the cholestasis in Cic-L(-/-) mice. Our findings indicate that CIC is a critical regulator of BA homeostasis, and that its dysfunction might be associated with chronic liver disease and metabolic disorders.

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Changes in levels of several hepatic transcription factors and proinflammatory cytokine genes in liver of Cic-L-/- mice at P18.(a) Western blot image showing the decrease in FOXA2, HNF1α, C/EBP1β, and RXRα protein levels in Cic-L-/- liver at P18. The right panel is a bar graph for quantitative analysis on levels of hepatic transcription factors based on Western blot image. The asterisk indicates non-specific bands. *P<0.05 and **P<0.01. All error bars show s.e.m. (b) qRT-PCR analysis of Hnf1α, C/ebpβ,Rxrα, and Foxa2 expression levels using liver total RNAs prepared from 18 day-old WT and Cic-L-/- mice (n = 5 per each genotype). All error bars show s.e.m. (c) qRT-PCR analysis of Il-1β, Il-6, and Tnfα expression levels using liver total RNAs prepared from 18 day-old WT and Cic-L-/- mice (n = 7 per each genotype). *P<0.05. All error bars show s.e.m.
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f4: Changes in levels of several hepatic transcription factors and proinflammatory cytokine genes in liver of Cic-L-/- mice at P18.(a) Western blot image showing the decrease in FOXA2, HNF1α, C/EBP1β, and RXRα protein levels in Cic-L-/- liver at P18. The right panel is a bar graph for quantitative analysis on levels of hepatic transcription factors based on Western blot image. The asterisk indicates non-specific bands. *P<0.05 and **P<0.01. All error bars show s.e.m. (b) qRT-PCR analysis of Hnf1α, C/ebpβ,Rxrα, and Foxa2 expression levels using liver total RNAs prepared from 18 day-old WT and Cic-L-/- mice (n = 5 per each genotype). All error bars show s.e.m. (c) qRT-PCR analysis of Il-1β, Il-6, and Tnfα expression levels using liver total RNAs prepared from 18 day-old WT and Cic-L-/- mice (n = 7 per each genotype). *P<0.05. All error bars show s.e.m.

Mentions: Many previous studies have shown that expression of genes involved in drug metabolism is coordinately regulated by several nuclear receptors and liver-enriched transcription factors222324252627282930. It is also known that cholestasis is closely associated with induction of proinflammatory modulators and decreased expression and activity of hepatic transcriptional regulators1314. In order to explain for the suppressed expression of several drug metabolism genes in liver and impaired BA homeostasis in Cic deficient mice, we examined hepatic levels of several liver-enriched transcription factors and nuclear receptors including FOXA2, HNF1α, C/EBPβ, RXRα, HNF4α, and FOXO1 by western blot analysis using liver total extracts from WT and Cic-L-/- mice at P18. We found that levels of FOXA2, HNF1α, C/EBPβ, and RXRα, but not HNF4α and FOXO1, were significantly down-regulated in livers of Cic-L-/- mice (Fig. 4a), suggesting that down-regulation of drug metabolism genes might result from decreased levels of a subset of liver-enriched transcription factors and nuclear receptors in Cic-L-/- mice. Interestingly, their mRNA levels were not significantly altered in Cic-L-/- mice (Fig. 4b), indicating that levels of these transcription factors are down-regulated at the post-transcriptional level. To validate that the down-regulation of several genes involved in drug metabolism and BA transport in Cic-L-/- liver was, at least in part, due to the decreased levels of hepatic transcriptional regulators, we examined promoter occupancy of FOXA2 and RXRα for their target genes whose levels were significantly down-regulated in Cic-L-/- liver (Figs. 2C and 3D). We carried out chromatin immunoprecipitation (ChIP) using either anti-FOXA2 or anti-RXRα antibody followed by qPCR for promoter regions that contain binding motifs for either FOXA2 or RXRα. ChIP-qPCR analyses revealed that less amount of FOXA2 and RXRα is associated with promoter regions of all the tested genes (Oatp2, Fmo3, and Ugt2b34 as FOXA2 targets16, and Cyp2b10 and Sult2a1 as RXRα targets3132) in liver of Cic-L-/- mice compared with WT (Fig. S4), suggesting that reduction in the levels of several hepatic transcriptional regulators leads to decrease in their promoter occupancy of target genes, thereby contributing to the down-regulation of a subset of genes mediating drug metabolism and BA transport in Cic-L-/- mice.


Deficiency of Capicua disrupts bile acid homeostasis.

Kim E, Park S, Choi N, Lee J, Yoe J, Kim S, Jung HY, Kim KT, Kang H, Fryer JD, Zoghbi HY, Hwang D, Lee Y - Sci Rep (2015)

Changes in levels of several hepatic transcription factors and proinflammatory cytokine genes in liver of Cic-L-/- mice at P18.(a) Western blot image showing the decrease in FOXA2, HNF1α, C/EBP1β, and RXRα protein levels in Cic-L-/- liver at P18. The right panel is a bar graph for quantitative analysis on levels of hepatic transcription factors based on Western blot image. The asterisk indicates non-specific bands. *P<0.05 and **P<0.01. All error bars show s.e.m. (b) qRT-PCR analysis of Hnf1α, C/ebpβ,Rxrα, and Foxa2 expression levels using liver total RNAs prepared from 18 day-old WT and Cic-L-/- mice (n = 5 per each genotype). All error bars show s.e.m. (c) qRT-PCR analysis of Il-1β, Il-6, and Tnfα expression levels using liver total RNAs prepared from 18 day-old WT and Cic-L-/- mice (n = 7 per each genotype). *P<0.05. All error bars show s.e.m.
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f4: Changes in levels of several hepatic transcription factors and proinflammatory cytokine genes in liver of Cic-L-/- mice at P18.(a) Western blot image showing the decrease in FOXA2, HNF1α, C/EBP1β, and RXRα protein levels in Cic-L-/- liver at P18. The right panel is a bar graph for quantitative analysis on levels of hepatic transcription factors based on Western blot image. The asterisk indicates non-specific bands. *P<0.05 and **P<0.01. All error bars show s.e.m. (b) qRT-PCR analysis of Hnf1α, C/ebpβ,Rxrα, and Foxa2 expression levels using liver total RNAs prepared from 18 day-old WT and Cic-L-/- mice (n = 5 per each genotype). All error bars show s.e.m. (c) qRT-PCR analysis of Il-1β, Il-6, and Tnfα expression levels using liver total RNAs prepared from 18 day-old WT and Cic-L-/- mice (n = 7 per each genotype). *P<0.05. All error bars show s.e.m.
Mentions: Many previous studies have shown that expression of genes involved in drug metabolism is coordinately regulated by several nuclear receptors and liver-enriched transcription factors222324252627282930. It is also known that cholestasis is closely associated with induction of proinflammatory modulators and decreased expression and activity of hepatic transcriptional regulators1314. In order to explain for the suppressed expression of several drug metabolism genes in liver and impaired BA homeostasis in Cic deficient mice, we examined hepatic levels of several liver-enriched transcription factors and nuclear receptors including FOXA2, HNF1α, C/EBPβ, RXRα, HNF4α, and FOXO1 by western blot analysis using liver total extracts from WT and Cic-L-/- mice at P18. We found that levels of FOXA2, HNF1α, C/EBPβ, and RXRα, but not HNF4α and FOXO1, were significantly down-regulated in livers of Cic-L-/- mice (Fig. 4a), suggesting that down-regulation of drug metabolism genes might result from decreased levels of a subset of liver-enriched transcription factors and nuclear receptors in Cic-L-/- mice. Interestingly, their mRNA levels were not significantly altered in Cic-L-/- mice (Fig. 4b), indicating that levels of these transcription factors are down-regulated at the post-transcriptional level. To validate that the down-regulation of several genes involved in drug metabolism and BA transport in Cic-L-/- liver was, at least in part, due to the decreased levels of hepatic transcriptional regulators, we examined promoter occupancy of FOXA2 and RXRα for their target genes whose levels were significantly down-regulated in Cic-L-/- liver (Figs. 2C and 3D). We carried out chromatin immunoprecipitation (ChIP) using either anti-FOXA2 or anti-RXRα antibody followed by qPCR for promoter regions that contain binding motifs for either FOXA2 or RXRα. ChIP-qPCR analyses revealed that less amount of FOXA2 and RXRα is associated with promoter regions of all the tested genes (Oatp2, Fmo3, and Ugt2b34 as FOXA2 targets16, and Cyp2b10 and Sult2a1 as RXRα targets3132) in liver of Cic-L-/- mice compared with WT (Fig. S4), suggesting that reduction in the levels of several hepatic transcriptional regulators leads to decrease in their promoter occupancy of target genes, thereby contributing to the down-regulation of a subset of genes mediating drug metabolism and BA transport in Cic-L-/- mice.

Bottom Line: We also found that levels of proinflammatory cytokine genes were up-regulated in Cic-L(-/-) liver.Moreover, induction of tumor necrosis factor alpha (Tnfα) expression and decrease in the levels of FOXA2, C/EBPβ, and RXRα were found in Cic-L(-/-) liver before BA was accumulated, suggesting that inflammation might be the cause for the cholestasis in Cic-L(-/-) mice.Our findings indicate that CIC is a critical regulator of BA homeostasis, and that its dysfunction might be associated with chronic liver disease and metabolic disorders.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, Pohang University of Science and Technology, Pohang, Gyeongbuk 790-784, Republic of Korea.

ABSTRACT
Capicua (CIC) has been implicated in pathogenesis of spinocerebellar ataxia type 1 and cancer in mammals; however, the in vivo physiological functions of CIC remain largely unknown. Here we show that Cic hypomorphic (Cic-L(-/-)) mice have impaired bile acid (BA) homeostasis associated with induction of proinflammatory cytokines. We discovered that several drug metabolism and BA transporter genes were down-regulated in Cic-L(-/-) liver, and that BA was increased in the liver and serum whereas bile was decreased within the gallbladder of Cic-L(-/-) mice. We also found that levels of proinflammatory cytokine genes were up-regulated in Cic-L(-/-) liver. Consistent with this finding, levels of hepatic transcriptional regulators, such as hepatic nuclear factor 1 alpha (HNF1α), CCAAT/enhancer-binding protein beta (C/EBPβ), forkhead box protein A2 (FOXA2), and retinoid X receptor alpha (RXRα), were markedly decreased in Cic-L(-/-) mice. Moreover, induction of tumor necrosis factor alpha (Tnfα) expression and decrease in the levels of FOXA2, C/EBPβ, and RXRα were found in Cic-L(-/-) liver before BA was accumulated, suggesting that inflammation might be the cause for the cholestasis in Cic-L(-/-) mice. Our findings indicate that CIC is a critical regulator of BA homeostasis, and that its dysfunction might be associated with chronic liver disease and metabolic disorders.

Show MeSH
Related in: MedlinePlus