TAZ inhibits glucocorticoid receptor and coordinates hepatic glucose homeostasis in normal physiological states

  1. Simiao Xu
  2. Yangyang Liu
  3. Ruixiang Hu
  4. Min Wang
  5. Oliver Stöhr
  6. Yibo Xiong
  7. Liang Chen
  8. Hong Kang
  9. Lingyun Zheng
  10. Songjie Cai
  11. Li He
  12. Cunchuan Wang
  13. Kyle D Copps
  14. Morris F White
  15. Ji Miao  Is a corresponding author
  1. Division of Endocrinology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Branch of the National Clinical Research Center for Metabolic Disease, China
  2. Division of Endocrinology, Boston Children’s Hospital, Harvard Medical School, United States
  3. Cancer Center, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, China
  4. Department of Gastrointestinal Surgery, The First Affiliated Hospital of Jinan University, China
  5. Department of Biliary-Pancreatic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, China
  6. Department of Pathology, Beth Israel Deaconess Medical Center, United States
  7. College of Science, Northeastern University, United States
  8. Department of Systemic Biology, Harvard Medical School, United States
  9. Transplantation Research Center, Brigham and Women’s Hospital, Harvard Medical School, United States
  10. Department of Pediatrics, Harvard Medical School, United States
10 figures, 4 tables and 1 additional file

Figures

Figure 1 with 3 supplements
humanTAZ protein is regulated by fasting and feeding in hepatocytes.

8- to 12-week-old C57BL/6J mice were ad libitum-fed (non-fasted) or fasted for 24 hr (fasted). (A–D) Hepatic proteins were measured by immunoblotting whole-cell lysates (A; quantified results are shown in B) or nuclear (Nuc) or cytoplasmic (Cyto) extracts (C; quantified results are shown in D). (E) Immunohistochemical staining for TAZ in the livers of mice fed ad libitum or fasted for 24 hr (CV, central vein; PV, portal vein; scale bar, 500 μM). (F, G) Protein levels were measured by immunoblotting lysates of pericentral and periportal mouse hepatocytes (F) or hepatocytes isolated from mice that were ad libitum-fed or fasted for 24 hr (G). (H) mRNA expression of hepatic mRNA expression of hepatic Taz was measured using real-time qRT-PCR. Data are means and SEMs; control values were set to 1; n = 6. Representative results of 2–3 independent experiments are shown. Data were analyzed by unpaired Student’s t-test; **p<0.01, ***p<0.001.

Figure 1—figure supplement 1
Hepatic gene expression in ad libitum-fed and fasted (24 hr) mouse livers.

(A) Gluconeogenic genes. (B) TAZ target gene involved in proliferation.

Figure 1—figure supplement 2
Validation of immunohistochemical staining for TAZ in control and L-TAZ KO liver sections.

Immunohistochemistry using an anti-TAZ antibody in L-TAZ KO and control floxed (Flox) mouse liver sections (CV, central vein; PV, portal vein; scale bar, 50 μM).

Figure 1—figure supplement 3
mRNA and protein expression of TAZ in primary mouse hepatocytes.

Primary mouse hepatocytes were incubated in low glucose media (1 g/L) without fetal bovine serum (FBS) for overnight and then placed in high glucose media (4.5 g/L) with FBS for the indicated times. Protein levels (A) and mRNA expression (B) were measured. Data are means and SEMs of triplicated wells. Data were analyzed by one-way ANOVA.

Figure 2 with 5 supplements
Knockdown or knockout of hepatic TAZ increases gluconeogenic gene expression and blood glucose concentrations in mice.

(A–G) 8- to 12-week-old C57BL/6J mice were administered AdshTAZ or AdshControl (AdshCon), then sacrificed 8 days later in the ad libitum-fed state, when the mRNA expression of hepatic genes (A, C) and the corresponding protein levels (B, D, and E [quantified results of D]) were measured. (F) Ad libitum-fed and fasting blood glucose concentrations were measured. (G) Six days after adenoviral injection, the mice underwent pyruvate tolerance testing (PTT) (left) and the areas under the curves (AUCs) were calculated (right); p<0.001 AdshTAZ versus AdshCon. (H–M) 10- to 12-week-old, age- and sex-matched L-TAZ KO and floxed (Flox) control littermates were studied. (H, I) TAZ protein was measured in whole liver cell lysates (H) or isolated hepatocytes (I) by immunoblotting. (J, K) Ad libitum-fed and fasting blood glucose concentrations were measured. (L, M) Mice underwent PTT; p<0.001 L-TAZ KO versus Flox. Data are means and SEMs; control values were set to 1 (A, C, E); n = 7–10. Representative results of 2–3 independent experiments are shown. Data were analyzed by unpaired Student’s t-test (A, C, E–F, G [right], J–K, L–M [right]) and two-way ANOVA (G [left] and L–M [left]); *p<0.05, **p<0.01, ***p<0.001.

Figure 2—figure supplement 1
Information on mice administered AdshTAZ or AdshCon.

Body (A), epididymal white adipose tissue (WAT) (B), and liver weight (C) were measured. (D) H&E staining of liver sections (scale bar, 50 μM). Data were analyzed by unpaired Student’s t-test; **p<0.01.

Figure 2—figure supplement 2
TAZ knockdown has no effects on protein expression of key gluconeogenic factors and insulin and glucagon signaling in mice.

Hepatic proteins were measured by immunoblotting whole-cell lysates.

Figure 2—figure supplement 3
TAZ knockdown has no effects on plasma insulin and glucagon concentrations in the ad libitum-fed state.

Plasma insulin (A) and glucagon (B) were measured. Data were analyzed by unpaired Student’s t-test.

Figure 2—figure supplement 4
Information on L-TAZ KO and flox mice.

Body (A) and liver weight (B) were measured. (C) H&E staining of liver sections (scale bar, 50 μM). Data were analyzed by two-way ANOVA (A) and unpaired Student’s t-test (B).

Figure 2—figure supplement 5
L-TAZ KO has no effects on insulin sensitivity.

(A) Hepatic proteins were measured by immunoblotting whole-cell lysates. (B, C) Insulin sensitivity was assessed by intraperitoneal insulin tolerance test (ITT). Glucose percentages relative to the basal blood glucose concentration (B) are plotted. Data were analyzed by two-way ANOVA.

Figure 3 with 2 supplements
Overexpression of hepatic TAZ inhibits gluconeogenic gene expression and reduces blood glucose concentration in mice.

8- to 12-week-old C57BL/6J mice were administered AdGFP or AdTAZ, then sacrificed 5 days later, after a 24 hr fast, when the hepatic protein levels in nuclear extracts (A [representative image, left; quantified results, right]), isolated pericentral hepatocytes (B), or whole-cell lysates (E, F [quantified results of E]) were measured. Blood glucose (C) and mRNA expression of hepatic genes (D) were measured. (G, H) Five days after adenoviral injection, the mice underwent glucagon challenge (G) or pyruvate tolerance testing (PTT) (H); p<0.001 L-TAZ KO versus Flox. Data are means and SEMs; control values were set to 1 (A, D, F); n = 5–10. Representative results of 2–3 independent experiments are shown. Data were analyzed by unpaired Student’s t-test (A, C, D, F, G [right], H [right]) and two-way ANOVA (G [left], H [left]); *p<0.05, **p<0.01, ***p<0.001.

Figure 3—figure supplement 1
Immunohistochemical staining for TAZ in livers of mice administered AdTAZ or control AdGFP and fasted for 24 hr.

CV, central vein; PV, portal vein; scale bar, 500 μM.

Figure 3—figure supplement 2
Information on mice administered AdTAZ or control AdGFP.

Mouse body weight (A), epididymal white adipose tissue weight (WAT) weight (B), liver weight (C), and plasma insulin (F, left) and glucagon (F, right) levels were measured. (D) H&E staining of liver sections (scale bar, 50 μM). (E) Hepatic proteins were measured by immunoblotting whole-cell lysates. Data were analyzed by unpaired Student’s t-test; *p<0.05.

Figure 4 with 3 supplements
The inhibition of gluconeogenic gene expression by TAZ is hepatocyte-autonomous.

Primary mouse hepatocytes were isolated from 8- to 12-week-old C57BL/6J mice. (A–D) To knock down TAZ, cells were infected with AdshTAZ or AdshControl (AdshCon). (E–H) To overexpress TAZ, cells were infected with AdTAZ or AdGFP. Cells were treated with glucagon (20 nM) for 3 hr (B, F) or dexamethasone (Dex, 100 nM) for 6 hr (C, G). (A [left]–C, F, G) Gene expression was measured using real-time RT-PCR. (A [right], E) TAZ protein levels were measured by immunoblotting whole-cell lysates and the quantified results of (E) are shown on the right. (D, H) Glucose production, in the presence of glucagon (20 nM), was assessed by measuring the glucose concentration in the media at the indicated times. Data are means and SEMs of three wells; control values were set to 1. Data were analyzed by unpaired Student’s t-test (A, E) and two-way ANOVA (B–D and F–H). *p<0.05, **p<0.01, ***p<0.001 versus control adenovirus-treated cells; in (B–D) and (F–H), #p<0.05, ###p<0.001 versus similarly treated controls. Representative results of 2–5 independent experiments are shown.

Figure 4—figure supplement 1
The effect of TAZ on the induction of glycogen synthase by dexamethasone (Dex).

Primary mouse hepatocytes were infected with AdTAZ or control AdGFP and were treated with Dex (100 nM) for 6 hr. Gene expression was measured using real-time RT-PCR. Data were analyzed by two-way ANOVA. **p<0.01 versus control adenovirus-treated cells; ##p<0.01 versus similarly treated controls.

Figure 4—figure supplement 2
Protein expression in primary mouse hepatocytes in the presence of TAZ knockdown or overexpression.

Cells were infected with AdshTAZ or AdshCon (A) or AdTAZ or AdGFP (B). Protein levels were measured by immunoblotting whole-cell lysates.

Figure 4—figure supplement 3
TAZ knockdown or overexpression in primary hepatocytes has no effects on insulin or glucagon signaling.

Cells were infected with AdshTAZ or AdshCon (A, B) or AdTAZ or AdGFP (C, D). Protein levels were measured by immunoblotting whole-cell lysates. Cells were treated with glucagon (20 nM) for 30 min (A, C) or insulin (20 nM) for 10 min (B, D).

Figure 5 with 3 supplements
TAZ interacts with glucocorticoid receptor (GR) and inhibits the transactivation of gluconeogenic genes by GR.

(A–D, G, K, L, N, O) HepG2 cells were co-transfected with expression vectors, luciferase reporters, and an internal control (Renilla), and treated with dexamethasone (Dex; 100 nM), as indicated. Relative luciferase activity (RLA) is presented after normalization to the Renilla activity. (E, F, M, Q) 293A cells were transfected with expression vectors, which was followed by immunoprecipitation and immunoblotting, as indicated. (H–J, R, S) Primary mouse hepatocytes were infected with adenoviruses, with or without glucagon or Dex treatment. (R, S) Cells were treated with RU486 30 min prior to glucagon treatment. Protein levels (H) and gene expression (I, J, R, S) were measured by immunoblotting and real-time RT-PCR, respectively. (P) Chromatin immunoprecipitation (ChIP) assays were performed using an anti-GR antibody or a control IgG. The relative enrichment of GR was assessed using real-time RT-PCR, and primers for the indicated regions of the Pck1 and G6pc genes were performed using an anti-GR antibody or a control IgG. Data are means and SEMs of three or four wells or immunoprecipitation reactions. Representative results of 2–5 independent experiments are shown. Data were analyzed by one-way ANOVA (A–D, G, K, L, N, O) and two-way ANOVA (I, J, P, R, S). *p<0.05, **p<0.01, ***p<0.001, #p<0.05, ##p<0.01, ###p<0.001. In (A–D, G, K, L, N, O), * denotes comparisons with wells treated with GR/Dex or GR/PGC1α/Dex alone; in (I, J R, S), # denotes comparisons with controls administered the same viruses; in (P), # denotes a comparison with IgG.

Figure 5—source data 1

A conserved I/LPKY motif is identified in glucocorticoid receptor (GR) from various species.

Highlighted in red.

https://cdn.elifesciences.org/articles/57462/elife-57462-fig5-data1-v2.eps
Figure 5—figure supplement 1
TAZ inhibits PCK-Luc activity.

(A, C) HepG2 cells were co-transfected with expression vectors, luciferase reporters, and an internal control (Renilla), and treated with dexamethasone (Dex) (100 nM), as indicated. Relative luciferase activity (RLA) is presented. (B) Cells were co-transfected with a flag-TAZ expression vector and a TAZ knockdown vector, or a control vector, as indicated, and TAZ expression was measured by immunoblotting whole-cell lysates. Data are means and SEMs of triplicated wells; controls were set to 1. Data were analyzed by one-way ANOVA. **p<0.01 denotes comparisons with wells treated with GR/PGC1α/HNF4α/Dex alone.

Figure 5—figure supplement 2
Effects of TAZ mutants on TEAD transactivation.

293A A cells were co-transfected with expression vectors, luciferase reporters, and an internal control (Renilla), and treated with dexamethasone (Dex) (100 nM), as indicated. Relative luciferase activity (RLA) is presented. Data are means and SEMs of triplicated wells; control was set to 1. Data were analyzed by one-way ANOVA. **p<0.01 and ***p<0.001 denotes comparisons with wells transfected with TEAD4 alone.

Figure 5—figure supplement 3
TAZ has no effects on glucocorticoid receptor (GR) nuclear localization or the amounts of GR dimer.

(A) 293A A cells were transfected with expression vectors for GR fused with a GFP, TAZ, or a control empty vector (Con), and treated with dexamethasone (Dex) (100 nM) or vehicle for 6 hr prior to fixation and DAPI staining. GFP-GR distribution was examined by a fluorescent microscope. (B) 293A A cells were transfected with expression vectors for GR, TAZ, or a control empty vector for 48 hr. Cells were then treated with dithiobis (succinimidyl propionate) (DSP) (3 µM) for 30 minutes on ice, and GR dimer was measured by immunoblotting whole-cell lysates.

Figure 6 with 2 supplements
TAZ represses glucocorticoid receptor (GR) transactivation of gluconeogenic genes in mouse liver.

(A, G) Endogenous TAZ was immunoprecipitated from liver nuclear extracts prepared from ad libitum-fed C57BL/6J mice (A) or mice treated with RU486 or vehicle (G), and the amounts of GR in the immunoprecipitates were measured by immunoblotting. C57BL/6J mice were administered AdshTAZ or AdshCon, as in Figure 2 (B, C, H) or AdTAZ or AdGFP, as in Figure 3 (D–F and K–N). (I, J) L-TAZ KO and control mice. (H–N) Mice were treated with RU486 (RU) or vehicle (V), as indicated. The relative enrichment of GR (B, D, N), acetylated-histone 4 (Ac-H4) (C, E, N), and Pol II (F) in the liver extracts was assessed using chromatin immunoprecipitation (ChIP) assays. (H, I, K) Blood glucose concentration. (L) Hepatic gene expression. (J, M) Pyruvate tolerance testing (PTT). Data are means and SEMs; n = 5–8; for ChIP, the data are the results of triplicate or quadruplicate immunoprecipitations. Data were analyzed by two-way ANOVA (B–F, I, J, M, N) and unpaired Student’s t-test (H, K, L). *p<0.05, **p<0.01, ***p<0.001, #p<0.05, ##p<0.01, ###p<0.001; in (B–F, N), # denotes comparisons with IgG; in (I, J), # denotes comparisons with vehicle-treated controls of the same genotype; NS, not significant.

Figure 6—figure supplement 1
TAZ does not bind to gluconeogenic gene promoters.

8- to 12-week-old male C57BL/6J mice were administered adenoviruses expressing TAZ (flag-tagged) or GFP for 5 days. Chromatin immunoprecipitation (ChIP) assays were performed from liver extracts using indicated antibodies. Data are means and SEMs of 3–4 immunoprecipitates. Data were analyzed by two-way ANOVA; * p<0.05, **p<0.01, #p<0.05, ##p<0.01, ###p<0.001; # denotes comparisons with IgG.

Figure 6—figure supplement 2
Blood glucose concentrations and hepatic gene expression of mice administered AdTAZ and dexamethasone (Dex).

8- to 12-week-old C57BL/6J J mice were administered adenoviruses expressing TAZ or GFP for 5 days. Mice were treated with Dex or vehicle. Blood glucose (A) and hepatic gene expression (B) were measured. Data were analyzed by one-way ANOVA; *p<0.05, **p<0.01, ***p<0.001.

Figure 7 with 1 supplement
The inhibition of gluconeogenic gene expression by TAZ requires its WW domain.

C57BL/6J mice (A–D) or (E) C57BL/6J mice were ad libitum-fed (non-fasted) or fasted for 24 hr. (A) Blood glucose. (B) Pyruvate tolerance testing (PTT). (C) Hepatic gene expression. (D, E) Chromatin immunoprecipitation (ChIP) assays were conducted using the indicated antibodies. (F) Regulation of gluconeogenic gene expression by TAZ in the fed and fasting states. Data are means and SEMs; n = 5–8; except for (D) and (E), in which the data are the results of triplicate or quadruplicate immunoprecipitations. Data were analyzed by one-way ANOVA (A, B [right], C), two-way ANOVA (B [left], D, E). *p<0.05, **p<0.01, ***p<0.001, #p<0.05, ##p<0.01, ###p<0.001. In (B), * denotes comparisons with AdGFP and # denotes comparisons between AdS89A and AdTAZΔWW; in (D, E), # denotes comparisons with IgG.

Figure 7—figure supplement 1
Body and liver mass and hepatic TAZ expression in mice administered AdTAZ mutants or AdGFP.

Mice were administered adenoviruses expressing TAZ mutants (flag-tagged) or GFP for 5 days. Body weight (A) and liver weight (B) were measured, and hepatic protein levels (C) were measured by immunoblotting whole -cell lysates. Data were analyzed by one-way ANOVA; **p<0.01, ###p<0.001.

Figure 8 with 1 supplement
TAZ regulates glucose production in insulin-resistant states.

(A–C) 8- to 10-week-old L-DKO or flox controls were administered the indicated adenoviruses. (A) Blood glucose. (B) Pyruvate tolerance testing (PTT). (C) Hepatic gene expression. (D) 4- to 6-week-old C57BL/6J mice were fed chow or a high-fat diet (HFD) for 2 months. Mice that were ad libitum-fed or fasted for 24 hr were compared, and hepatic TAZ expression was measured by immunoblotting whole-cell lysates. (E, F) 4- to 6-week-old L-TAZ KO and floxed control littermates were fed an HFD for 2 months. Mice underwent PTT (E, left) or insulin tolerance test (ITT) (F, left) and the areas under the curves were calculated (right). Data are means and SEMs; n = 5–8. Data were analyzed by unpaired Student’s t-test (A, C, E [right], F [right]) or two-way ANOVA (B , E [left], F [left]); *p<0.05, **p<0.01, ***p<0.001; in (B), # denotes comparisons with flox mice administered the same virus.

Figure 8—figure supplement 1
L-DKO and flox mice administered AdTAZS89A.

(A) Hepatic proteins were measured by immunoblotting nuclear (Nuc) or cytoplasmic (Cyto) extracts of L-DKO and flox control mice. (B, C) Female L-DKO and flox controls were administered adenoviruses expressing TAZ or GFP. Body (B) and liver (C) weight were measured. Data were analyzed by unpaired Student’s t-test. *p<0.05, ***p<0.001.

Author response image 1
PTT data plotted using SEM and SD.

Figures 2G (A), 2L (C), and 2M (E), which are plotted using SEM, are compared with Figures R1B, R1D, and R1F, respectively, which are plotted using SD.

Author response image 2
Hepatic TAZ protein expression in hyperglycemic and insulin resistant mouse models.

Hepatic TAZ protein expression was measured by immunoblotting liver cytoplasmic (Cyto) and nuclear (Nucl) fractions.

Tables

Appendix 1—key resources table
Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional information
Genetic reagent (Mus musculus)C57BL/6JJackson LaboratoryStock #: 000664 RRID:IMSR_JAX:000664
Genetic reagent (M. musculus)Tazflox/flox: Yapflox/floxJackson LaboratoryStock #: 030532RRID:IMSR_JAX:030532
Genetic reagent (M. musculus)Albumin-CreJackson LaboratoryStock #: 003574RRID:IMSR_JAX:003574
Genetic reagent (M. musculus)Tazflox/flox: Alb-CreThis paperSee ‘Animals and treatments’ in Materials and methods
Cell line (Homo sapiens)HepG2ATCCCat. #: HB-8065RRID:CVCL 0027
Cell line (H. sapiens)293AThermo Fisher ScientificCat. #: R70507RRID:CVCL_6910
Commercial assay or kitBLOCK-iT U6 RNAi entry vector kitThermo Fisher ScientificCat. #: K494500
Commercial assay or kitBLOCK-iT U6 adenoviral RNAi expression systemThermo Fisher ScientificCat. #: K494100
Commercial assay or kitStellux Chemiluminscence rodent insulin ELISA kitAlpcoCat. #:80-INSMR-CH01
Commercial assay or kitMouse glucagon ELISA kitAlpcoCat. #:48-GLUHU-E01
Commercial assay or kitDual-luciferase reporter assay kitPromegaCat. #: E1960
Commercial assay or kitVIP substrate Kit, HRPVector LaboratoriesCat. #: SK-4600 RRID:AB_2336848PMID:28123024
Commercial assay or kitMutagenesis kitAgilentCat. #: 210,519
Commercial assay or kitcDNA synthesis kitThermo Fisher ScientificCat. #: 4368813
Commercial assay or kitNE-PER Nuclear and Cytoplasmic Extraction kitThermo Fisher ScientificCat. #: 78835
Commercial assay or kitAmplex glucose oxidase assay kitThermo Fisher ScientificCat. #: A22189
Recombinant DNA reagentpCMV-TOPO TAZ (human)AddgeneCat. #: 24809 RRID:Addgene_24809PMID:18568018
Recombinant DNA reagentpcDNA3-flag-TAZ (human)This paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpcDNA3-flag-TAZ S51A (human)This paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpcDNA3-flag-TAZ S89A (human)This paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpCMV-TOPO TAZΔWW (human)AddgeneCat. #: 24811RRID:Addgene_24811PMID:18568018
Recombinant DNA reagentpCMV-TOPO TAZΔCC (human)AddgeneCat. #: 24816 RRID:Addgene_24816PMID:18568018
Recombinant DNA reagentpcDNA3-flag-TAZΔWW (human)This paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpcDNA3-flag- TAZΔCC (human)This paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpcDNA3-flag-TAZ WWThis paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpGL3-3XGRE-LucThis paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpGL3-G6PC-Luc (human)Dr. Pere Puigserver
Recombinant DNA reagentpGL3-PCK1-Luc (human)Dr. Pere Puigserver
Recombinant DNA reagentpcDNA3-HNF4 α (mouse)Dr. Pere Puigserver
Recombinant DNA reagentpcDNA3-PGC1α (mouse)Dr. Pere Puigserver
Recombinant DNA reagentpEGFP-GRAddgeneCat. #: 47504 RRID:Addgene_47504
Recombinant DNA reagentpcDNA3-GR (human)This paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpcDNA3-GR4A (human)This paperSee ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagent8xGTIIC-LucAddgeneCat. #: 34615 RRID:Addgene_34615PMID:21654799
Recombinant DNA reagentpcDNA3-Flag-YAP1 (human)AddgeneCat. #: 18881 RRID:Addgene_18881PMID:18280240
Recombinant DNA reagentpRK5-TEAD1 (human)AddgeneCat. #: 33109 RRID:Addgene_33109PMID:18579750
Recombinant DNA reagentpAd-Track-CMV-GFPAddgeneCat. #: 16405 RRID:Addgene_16405PMID:9482916Construct to establish adenovirus
Recombinant DNA reagentpAd-Track-CMV-Flag-TAZ (human)This paperConstruct to establish adenovirus expressing TAZ; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpAd-Track-CMV-Flag-TAZΔWW (human)This paperConstruct to establish adenovirus expressing TAZΔWW; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentpAd-Track-CMV-Flag-TAZS89A (human)This paperConstruct to establish adenovirus expressing TAZS89A; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentU6-shLamin (human)This paperControl for shTAZ; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentU6-shTAZ (human)This paperConstruct to knockdown TAZ in HepG2 cells; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentU6-shLacZThis paperConstruct to establish adenovirus expressing shControl; control for shTAZ; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentU6-shTAZ (mouse)This paperConstruct to establish adenovirus expressing shTAZ; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentAd-shLacZThis paperControl adenovirus expressing shLacZ; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentAd-shTAZ (mouse)This paperAdenovirus expressing shTAZ; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentAd-Track-CMV-GFPThis paperControl adenovirus expressing GFP, generated from pAd-Track-CMV-GFP vector; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentAd-Track-CMV-flag-TAZThis paperAdenovirus generated from pAd-Track-CMV-TAZ; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentAd-Track-CMV-flag-TAZΔWWThis paperAdenovirus generated from pAd-Track-CMV-TAZΔWW; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Recombinant DNA reagentAd-Track-CMV-flag-TAZS89AThis paperAdenovirus generated from pAd-Track-CMV-TAZS89A; see ‘Plasmid and adenoviral vector constructs’ in Materials and methods
Sequence-based reagentshLamin (human)This paperCTGGACTTCCAGAAGAACA
Sequence-based reagentshLacZThis paperCTACACAAATCAGCGATTT
Sequence-based reagentshTAZ (human)This paperGCTCAGATCCTTTCCTCAATG
Sequence-based reagentshTAZ (mouse)This paperGCCAGAGATACTTCCTTAATC
Sequence-based reagentMouse-Tbp-FThis paperqRT-PCR primerACCTTCACCAATGACTCCTATG
Sequence-based reagentMouse-Tbp-RThis paperqRT-PCR primerTGACTGCAGCAAATCGCTTGG
Sequence-based reagentMouse-Cry61-FThis paperqRT-PCR primerCAAGAAATGCAGCAAGACCA
Sequence-based reagentMouse-Cry61-RThis paperqRT-PCR primerGGCCGGTATTTCTTGACACT
Sequence-based reagentMouse-Ctgf-FThis paperqRT-PCR primerTCCACCCGAGTTACCAATGA
Sequence-based reagentMouse-Ctgf -RThis paperqRT-PCR primerCAAACTTGACAGGCTTGGC
Sequence-based reagentMouse-G6pc-FThis paperqRT-PCR primerTGGCTTTTTCTTTCCTCGAA
Sequence-based reagentMouse-Pck1-FThis paperqRT-PCR primerTCGGAGACTGGTTCAACCTC
Sequence-based reagentMouse-Pck1-RThis paperqRT-PCR primerGAGGGACAGCAGCACCAT
Sequence-based reagentMouse-Taz-FThis paperqRT-PCR primerACAGGTGAAAATTCCGGTCA
Sequence-based reagentMouse-Taz -RThis paperqRT-PCR primerGAAGGCAGTCCAGGAAATCA
Sequence-based reagentMouse-Yap-FThis paperqRT-PCR primerAAGCCATGACTCAGGATGGA
Sequence-based reagentMouse-Yap-RThis paperqRT-PCR primerGTTCATGGCAAAACGAGGGTC
Sequence-based reagentMouse-Ctgf-ChIP-FThis paperqChIP primerTTCCTGGCGAGCTAAAGTGT
Sequence-based reagentMouse-Ctgf-ChIP-RThis paperqChIP primerCCTTCCTGCCTCATCAACTC
Sequence-based reagentMouse-G6pc-ChIP-GRE-FThis paperqChIP primerAGCACTGTCAAGCAGTGTGC
Sequence-based reagentMouse-G6pc-ChIP-GRE-FThis paperqChIP primerGCAAAACAGGCACACAAAAA
Sequence-based reagentMouse-G6pc-ChIP-HNF4E-FThis paperqChIP primerCCCTGAACATGTTTGCATCA
Sequence-based reagentMouse-G6pc-ChIP-HNF4E-RThis paperqChIP primerGTAGGTCAATCCAGCCCTGA
Sequence-based reagentMouse-Pck1-ChIP-Con-FThis paperqChIP primerTGGGAGACACACATCTTATTCCA
Sequence-based reagentMouse-Pck1-ChIP-Con-RThis paperqChIP primerGTCCCTCTATAGACTTCCAGCACA
Sequence-based reagentMouse-Pck1-ChIP-GRE-FThis paperqChIP primerTGCAGCCAGCAACATATGAA
Sequence-based reagentMouse-Pck1-ChIP-GRE-FThis paperqChIP primerTGATGCAAACTGCAGGCTCT
Sequence-based reagentMouse-Pck1-ChIP-HNF4E-FThis paperqChIP primerTAAGGCAAGAGCCTGCAGTT
Sequence-based reagentMouse-Pck1-ChIP-HNF4E-FThis paperqChIP primerAGGCCCCTCTATCAGCCATA
Antibody(Rabbit polyclonal) anti-TAZCell Signaling TechnologyCat. #: 4883 RRID:AB_1904158PMID:29533785IB (1:1000)
Antibody(Rabbit monoclonal) anti-p-TAZ (Ser89)Cell Signaling TechnologyCat. #: 59971 RRID:AB_2799578IB (1:1000)
Antibody(Rabbit polyclonal) anti-AKTCell Signaling TechnologyCat. #: 9272 RRID:AB_329827PMID:23653460IB (1:1000)
Antibody(Rabbit polyclonal) anti-p-AKT (Thr308)Cell Signaling TechnologyCat. #: 9275 RRID:AB_329828PMID:23715867IB (1:1000)
Antibody(Rabbit polyclonal) anti-p-AKT (Ser473)AbclonalCat. #: AP0098 RRID:AB_2770899IB (1:1000)
Antibody(Mouse monoclonal) anti-beta-actinSanta Cruz BiotechnologyCat. #: sc-47778 RRID:AB_2714189PMID:28017329IB (1:3000)
Antibody(Rabbit monoclonal) anti-CREBCell Signaling TechnologyCat. #: 9197RRID:AB_331277PMID:24080368IB (1:1000)
Antibody(Rabbit polyclonal) anti-p-CREB (Ser133)AbclonalCat. #: AP0333RRID:AB_2771008IB (1:1000)
Antibody(Mouse monoclonal) anti-FlagAbclonalCat. #: AE005RRID:AB_2770401IB (1:10000)IP (1 µg/IP)ChIP (1–2 µg/IP)
Antibody(Rabbit polyclonal) anti-FlagCell Signaling TechnologyCat. #: 2368 RRID:AB_2217020PMID:25514086IB (1:1000)
Antibody(Rabbit monoclonal) anti-FoxO1Cell Signalling TechnologyCat. #: 2880 RRID:AB_2106495PMID:24248465IB (1:1000)
Antibody(Rabbit monoclonal) anti-p-FoxO1 (Ser256)Cell Signaling TechnologyCat. #: 84192 RRID:AB_2800035PMID:31583122IB (1:1000)
Antibody(Rabbit polyclonal) anti-G6PCAbcamCat. #: ab83690 RRID:AB_1860503PMID:25774555IB (1:1000)
Antibody(Mouse monoclonal) anti-GAPDHSanta Cruz BiotechnologyCat. #: sc-32233 RRID:AB_627679PMID:24105481IB (1:1000)
Antibody(Mouse monoclonal) anti-GLULBD BiosciencesCat. #: 610517 RRID:AB_397879PMID:17120293IB (1:1000)
Antibody(Rabbit polyclonal) anti-GRAbclonalCat. #: A2164 RRID:AB_2764182IB (1:3000)
AntibodyGoat polyclonal anti-HMGCRSanta Cruz BiotechnologyCat. #: sc-27578 RRID:AB_2118199PMID:26824363IB (1:1000)
Antibody(Rabbit polyclonal) anti-HNF4αSanta Cruz BiotechnologyCat. #: sc-8987 RRID:AB_2116913PMID:29937200IB (1:000)
Antibody(Rabbit polyclonal) anti-PGC1αAbclonalCat. #: A12348 RRID:AB_2759191IB (1:000)
Antibody(Rabbit monoclonal) anti-TubulinCell SignalingTechnologyCat. #: 2125 RRID:AB_2619646PMID:28343940IB (1:5000)
Antibody(Mouse monoclonal) anti-VinculinSanta Cruz BiotechnologyCat. #: sc-73614 RRID:AB_1131294PMID:29017056IB (1:5000)
Antibody(Rabbit polyclonal) anti-YAPCell Signaling TechnologyCat. #: 4912 RRID:AB_2218911PMID:28323616IB (1:000)
Antibody(Mouse polyclonal) anti-IRS1Provided by Dr. Morris WhitePMID:29867232IB (1:000)
Antibody(Mouse polyclonal) anti-IRS2Provided by Dr. Morris WhitePMID:29867232IB (1:000)
Antibody(Rabbit polyclonal) anti-Ac-Histone4AbclonalCat. #: A15233 RRID:AB_2762128ChIP (1 µg/IP)
Antibody(Mouse monoclonal) anti-GRSanta Cruz BiotechnologyCat. #: Sc-393232 RRID:AB_2687823PMID:28467930ChIP (1–2 µg/IP)
Antibody(Rabbit polyclonal) anti-TAZAbclonalCat. #: A8202 RRID:AB_2721146ChIP (1–2 µg/IP)IHC (1: 200)
AntibodyGoat anti-(Rabbit polyclonal) IgG-HPRThermo Fisher ScientificCat. #: 31460RRID:AB_228341PMID:24932808IB (1:5000–20,000)
AntibodyGoat anti-(Mouse polyclonal) IgG-HRPThermo Fisher ScientificCat. #: 31430RRID:AB_228307PMID:10359649IB (1:5000–20,000)
AntibodyRabbit anti-goat (Rabbit polyclonal) IgG-HRPSanta Cruz BiotechnologyCat. #: sc-2768RRID:AB_656964PMID:23970784IB (1:5000–15,000)
Chemical compound, drugGlucagonSigmaCat. #: G2044
Chemical compound, drugRU486SigmaCat. #: M8046
Chemical compound, drugDexamethasoneSigmaCat. #: D4902For cell culture studies
Chemical compound, drugDexamethasoneSigmaCat. #: 2915For in vivo studies
Chemical compound, drugSodium pyruvateSigmaCat. #: P5280
Chemical compound, drugProtease inhibitor cocktail tabletSigmaCat. #: S8820
Chemical compound, drugPhosphatase inhibitor cocktail tabletSigmaCat. #: 4906837001
Chemical compound, drugBovine insulinSigmaCat. #: I0516For cell culture studies
Chemical compound, drugHuman insulinHumulin R U-100Eli LilyCat. #: HI-210For in vivo studies
Chemical compound, drugPercollCytivaCat. #: 17089109
Chemical compound, drugTrizolThermo Fisher ScientificCat. #: 15596018
Chemical compound, drugDSPThermo Fisher ScientificCat. #: PG82081
OtherSYBR Green PCR master mixBiolineCat. #: BIO-84050
OtherCollagen Type I Rat TailCorningCat. #: 354,236
OtherCollagenase Type IWorthington Biochemical CorporationCat. #: LS004196
OtherPVDF membraneSigmaCat. #: IPVH00010
OtherECLThermo Fisher ScientificCat. #: A43841
OtherAgarose A/G beadsSanta Cruz BiotechnologyCat. #: sc-2003 RRID:AB_10201400PMID:28392145
OtherDMEM cell culture mediaThermo Fisher ScientificCat. #: 11965118
OtherM199 cell culture mediaThermo Fisher ScientificCat. #:11043023
OtherDMEM low glucose cell culture media, no phenol redThermo Fisher ScientificCat. #:11054020
OtherLipofectamine 2000Thermo Fisher ScientificCat. #: 11668019Transfection reagent
Author response table 1
Blood glucose concentrations (mg/dl) of AdshTAZ- and AdshCon-infected mice during the PTT .
Time (min)
Treatment0153060120
AdshCon769112010573
AdshCon7311613911964
AdshCon761091008671
AdshCon67939810372
AdshCon6311911910457
AdshCon781041208368
AdshCon669612210667
AdshTAZ9612916116891
AdshTAZ79130149114103
AdshTAZ80122135116105
AdshTAZ105195195116108
AdshTAZ8614016214694
AdshTAZ8513015512289
AdshTAZ10212912712594
AveAdshCon71.3104.0116.9100.967.4
AdshTAZ90.4139.3154.9129.697.7
SEMAdshCon2.2224.2205.3024.6982.103
AdshTAZ3.9819.4968.3167.6152.826
SDAdshCon5.88011.16514.02912.4295.563
AdshTAZ10.53325.12422.00320.1487.477
Two-way ANOVAp0.09420.0002<0.00010.00330.0017
Author response table 2
Blood glucose concentrations (mg/dl) of female L-TAZ KO and floxed control mice during the PTT.
Time (min)
Treatment0153060120
Flox (F)5671756334
Flox (F)4485987060
Flox (F)52791038651
Flox (F)5489976352
Flox (F)51821038547
Flox (F)52891017653
Flox (F)5387898347
Flox (F)5481956659
L-TAZ KO (F)58971208663
L-TAZ KO (F)661021148673
L-TAZ KO (F)58891069159
L-TAZ KO (F)57931118364
L-TAZ KO (F)56931088862
L-TAZ KO (F)58961098753
L-TAZ KO (F)57891059665
L-TAZ KO (F)63941088859
L-TAZ KO (F)629212810663
L-TAZ KO (F)629111310260
AveFlox (F)52.082.995.174.050.4
L-TAZ KO (F)59.793.4112.291.362.1
SEMFlox (F)1.2682.1423.3083.4642.890
L-TAZ KO (F)1.0441.3432.2402.3991.629
SDFlox (F)3.5866.0589.3579.7988.176
L-TAZ KO (F)3.3024.24870.847.5875.152
Two-way ANOVAp0.08360.0067<0.0001<0.0010.002
Author response table 3
Blood glucose concentrations (mg/dl) of male L-TAZ KO and floxed control mice during the PTT.
Time (min)
Treatment0153060120
Flox (M)46651026952
Flox (M)4484977351
Flox (M)5189957956
Flox (M)5478986957
Flox (M)53741017760
Flox (M)46821087756
Flox (M)5475976855
Flox (M)51891037958
L-TAZ KO (M)56921298562
L-TAZ KO (M)56941189559
L-TAZ KO (M)59971088860
L-TAZ KO (M)6610511010278
L-TAZ KO (M)551061109266
L-TAZ KO (M)61931159268
L-TAZ KO (M)58991058771
AveFlox (M)49.979.5100.173.955.6
L-TAZ KO (M)58.798.0113.691.666.3
SEMFlox (M)1.4072.8971.4931.6631.051
L-TAZ KO (M)1.4432.1383.0462.1702.552
SDFlox (M)3.9808.1944.2244.7042.973
L-TAZ KO (M)3.8175.6578.0595.7406.751
Two-way ANOVAp0.0173<0.0001<0.0001<0.0001<0.0026

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  1. Simiao Xu
  2. Yangyang Liu
  3. Ruixiang Hu
  4. Min Wang
  5. Oliver Stöhr
  6. Yibo Xiong
  7. Liang Chen
  8. Hong Kang
  9. Lingyun Zheng
  10. Songjie Cai
  11. Li He
  12. Cunchuan Wang
  13. Kyle D Copps
  14. Morris F White
  15. Ji Miao
(2021)
TAZ inhibits glucocorticoid receptor and coordinates hepatic glucose homeostasis in normal physiological states
eLife 10:e57462.
https://doi.org/10.7554/eLife.57462