Hammerhead-type FXR agonists induce an enhancer RNA Fincor that ameliorates nonalcoholic steatohepatitis in mice
- Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, United States
- Department of Biochemistry and Molecular Biology, McGovern Medical School, University of Texas Health Science Center, United States
Figures
Figure 1 with 2 supplements
Activation of farnesoid X receptor (FXR) by GW4064 induces FXR-induced non-coding RNA (Fincor), a novel enhancer RNA (eRNA), in mouse liver.
(A) Volcano plot from RNA-seq showing significantly induced eRNAs (Fincor is highlighted) in the livers of C57BL/6 male mice treated with GW4064 (i.p. injection, 30 mg/kg, 1 hr) or vehicle. The x …
Figure 1—figure supplement 1
Examples of FXR-regulated eRNAs and time course expression of Fincor after FXR activation.
(A, B) Examples of farnesoid X receptor (FXR)-regulated eRNAs produced near the genes Hes1 (A) and Slc35g1 (B) were shown. (C) Time course expression of FXR-induced non-coding RNA (Fincor). C57BL/6 …
Figure 1—figure supplement 2
RNA-seq analysis of the differentially expressed genes in the GW4064-treated mouse liver.
(A) Volcano plot showing the differential expressed genes (DEGs) after GW4064 treatment (DESeq2 FDR <0.05). The numbers refer to the number of genes up- or downregulated. (B) Bar plot showing the …
Figure 2 with 1 supplement
Ligand-activated farnesoid X receptor (FXR) directly activates transcription of enhancer RNAs including FXR-induced non-coding RNA (Fincor) in the liver.
(A) Left: experimental outline. Fxr floxed (Fxr-Flox) and Fxr liver-specific knockout (Fxr-LKO) male mice were fasted overnight and treated with vehicle or GW4064, and livers were collected 1 hr …
Figure 2—figure supplement 1
Validation of Fincor expression and the analysis of the epigenetic features (presence of histone marks and binding of FXR and RXR⍺) at the enhancers that display FXR-induced eRNAs.
(A) Farnesoid X receptor (FXR) protein levels in the livers isolated from Fxr floxed (Fxr-Flox) and Fxr liver-specific knockout (Fxr-LKO) mice are shown. (B) Validation of hepatic FXR-dependent …
Figure 3
Farnesoid X receptor-induced non-coding RNA (Fincor) is a liver-specific nucleus-enriched eRNA.
(A) Expression levels of Fincor in various tissues after GW4064 treatment. Data from C57BL/6 male mice fasted overnight and i.p. injected with vehicle or GW4064 (30 mg/kg) for 1 hr (n=2/group). Data …
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Figure 3—source data 1
Original file for the gel images shown in Figure 3C.
- https://cdn.elifesciences.org/articles/91438/elife-91438-fig3-data1-v1.zip
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Figure 3—source data 2
Original file for the gel images shown in Figure 3C with highlighted bands and sample labels.
- https://cdn.elifesciences.org/articles/91438/elife-91438-fig3-data2-v1.zip
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Figure 3—source data 3
Original file for the immunoblot image shown in Figure 3F.
- https://cdn.elifesciences.org/articles/91438/elife-91438-fig3-data3-v1.zip
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Figure 3—source data 4
Original file for the immunoblot image shown in Figure 3F with highlighted bands and sample labels.
- https://cdn.elifesciences.org/articles/91438/elife-91438-fig3-data4-v1.zip
Figure 4
Farnesoid X receptor-induced non-coding RNA (Fincor) is induced by the hammerhead class of non-steroidal farnesoid X receptor (FXR) agonists, including GW4064 and tropifexor.
(A) The chemical structures of the FXR agonists including the hammerhead class of synthetic FXR agonists, non-hammerhead-type synthetic agonist, semi-synthetic BA, and natural BA. BA: bile acid. (B) …
Figure 5 with 1 supplement
Generation of CRISPR/Cas9-mediated farnesoid X receptor-induced non-coding RNA (Fincor) liver-specific knockdown mice.
(A) Experimental scheme: male Cas9 mice were infected with adenovirus expressing sgRNA for Fincor or a control for 1 week. Then the liver and serum were collected from these mice after 4–5 hr of …
Figure 5—figure supplement 1
Generation of farnesoid X receptor-induced non-coding RNA (Fincor) liver-specific knockdown mice by CRISPR-Cas9 method.
(A) Left: experimental scheme for the farnesoid X receptor-induced non-coding RNA (Fincor) loss-of-function experiments. Right: illustration demonstrating the sequence targeted by the sgRNA in …
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Figure 5—figure supplement 1—source data 1
Original file for the gel image shown in Figure 5—figure supplement 1.
- https://cdn.elifesciences.org/articles/91438/elife-91438-fig5-figsupp1-data1-v1.zip
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Figure 5—figure supplement 1—source data 2
Original file for the gel image shown in Figure 5—figure supplement 1 with highlighted bands and sample labels.
- https://cdn.elifesciences.org/articles/91438/elife-91438-fig5-figsupp1-data2-v1.zip
Figure 6
In diet-induced nonalcoholic steatohepatitis (NASH) mice, tropifexor-mediated beneficial effects on reducing hepatic steatosis are largely independent of farnesoid X receptor-induced non-coding RNA (Fincor).
(A–E) Male Cas9 mice were fed a NASH diet for 12 weeks. The mice were randomly assigned to three groups and infected with adenovirus expressing sgRNA for Fincor or control. Three days later, the …
Figure 7 with 1 supplement
In diet-induced nonalcoholic steatohepatitis (NASH) mice, tropifexor-mediated beneficial effects on reducing liver fibrosis and inflammation are diminished by farnesoid X receptor-induced non-coding RNA (Fincor) downregulation.
(A) Representative images from hematoxylin and eosin (H&E), TUNEL, Sirius Red, and F4/80 staining of liver sections from the same cohort of mice described in Figure 6. Scale bar (50 μm). Image …
Figure 7—figure supplement 1
The effects of farnesoid X receptor-induced non-coding RNA (Fincor) downregulation on nonalcoholic steatohepatitis (NASH) pathologies.
Male Cas9 mice were fed with a NASH diet for 12 weeks. Then these mice were randomly assigned to two groups and infected with adenovirus expressing sgRNA for Fincor or control. The tissues were …
Figure 8
Hepatic expression of farnesoid X receptor-induced non-coding RNA (Fincor) is elevated in liver disease associated with inflammation and fibrosis.
(A) C57BL/6 mice were fed with a high-fat diet for 12 weeks. The liver RNAs were extracted and Fincor expression was measured (n=5/group). (B) C57BL/6 mice were fed with a high-fat diet with high …
Additional files
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Supplementary file 1
Sequencing data generated in this study and a list of Farnesoid X receptor (FXR)-associated eRNAs.
- https://cdn.elifesciences.org/articles/91438/elife-91438-supp1-v1.xlsx
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Supplementary file 2
Differentially expressed genes in the FXR-induced non-coding RNA (Fincor)-downregulated mice liver.
- https://cdn.elifesciences.org/articles/91438/elife-91438-supp2-v1.xlsx
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Supplementary file 3
The binding of transcription factors at the FXR-induced non-coding RNA (Fincor) locus.
- https://cdn.elifesciences.org/articles/91438/elife-91438-supp3-v1.docx
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Supplementary file 4
Predicted RNA binding proteins (RBPs) binding to FXR-induced non-coding RNA (Fincor).
- https://cdn.elifesciences.org/articles/91438/elife-91438-supp4-v1.docx
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Supplementary file 5
Primer sequences used in this study.
- https://cdn.elifesciences.org/articles/91438/elife-91438-supp5-v1.docx
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MDAR checklist
- https://cdn.elifesciences.org/articles/91438/elife-91438-mdarchecklist1-v1.docx
