Journal: The Journal of Biological Chemistry

Article Title: Rab2A modulates liver fibroblast growth factor 21 (FGF21) expression and systemic metabolism via apolipoprotein B–CREBH signaling

doi: 10.1016/j.jbc.2025.110977

Figure Lengend Snippet: Liver-specific inhibition of Rab2A increases FGF21 expression and secretion . A , serum FGF21 levels in shNC and shRab2A mice (male, n = 5 per group; samples from PMID: 35061665 ). B , serum FGF21 levels in Flox and LCK mice under “Random feed” and “Fasted” conditions (male, n = 5 per group). C , serum FGF21 levels in Flox and LCK mice after 12 weeks of HFHCD feeding (male, n = 6 versus 5). D , hepatic Fgf21 mRNA levels in “Random feed” Flox and LCK mice at 10 weeks of age (male, n = 5 per group). E , hepatic Fgf21 mRNA levels in “Fasted” Flox and LCK mice at 16 weeks of age (male, n = 5 per group). F , hepatic Fgf21 mRNA levels in Flox and LCK mice after 12 weeks of HFHCD feeding (male, n = 6 versus 5). G , representative immunoblots of hepatic FGF21 protein in “Fasted” Flox and LCK mice at 16 weeks of age. H , quantification of the immunoblots in ( G ). I , representative immunoblots of hepatic FGF21 protein in Flox and LCK mice after 12 weeks of HFHCD feeding. J , quantification of the immunoblots in ( I ). Data are presented as mean ± SD. p Values were determined using the unpaired two-tailed Student’s t test. FGF21, fibroblast growth factor 21; HFHCD, high-fat–high-cholesterol diet; LCK, liver-specific Rab2A knockout.

Article Snippet: The following primary and secondary antibodies were used: goat anti FGF21 (R&D Systems; AF3057), mouse anti-Rab2A (Proteintech; 67501-1-Ig), mouse anti-tubulin (DUONENG-BIO; AB0178801), mouse anti-actin (Zen-Bioscience; 200068-8F10), rabbit anti-CREBH (Kerafast; EWS101), rabbit anti-GAPDH (Proteintech; 10494-1-AP), rabbit anti-APOB (Proteintech; 20578-1-AP), mouse anti-CHC (Santa Cruz; sc-12734), rabbit anti-GRP94 (Proteintech; 14700-1-AP), mouse anti-GRASP55 (Santa Cruz; sc-271840), HRP-conjugated goat anti-mouse IgG (Jackson ImmunoResearch; 115-035-003), HRP-conjugated goat anti-rabbit IgG (Jackson ImmunoResearch; 111-035-003), and HRP-conjugated rabbit anti-goat IgG (Jackson ImmunoResearch; 305-035-003).

Techniques: Inhibition, Expressing, Western Blot, Two Tailed Test, Knock-Out

Journal: The Journal of Biological Chemistry

Article Title: Rab2A modulates liver fibroblast growth factor 21 (FGF21) expression and systemic metabolism via apolipoprotein B–CREBH signaling

doi: 10.1016/j.jbc.2025.110977

Figure Lengend Snippet: CREBH knockdown rescues FGF21 expression and the metabolic phenotypes of LCK mice . A , schematic of the experimental timeline for adeno-associated virus (AAV) injection in Flox and LCK mice. B and C , representative immunoblots and quantification of hepatic CREBH protein levels after AAV-mediated knockdown. D – M , analyses performed after 7 weeks of AAV injection and subsequent euthanasia (male, n = 6 versus 6 versus 6): Hepatic mRNA levels of Creb3l3 , Fgf21 , Apoa4 , and Fsp27β ( D ). Representative immunoblots and quantification of hepatic FGF21 protein levels ( E and F ). Serum FGF21 levels ( G ). mRNA expression of thermogenic-related genes in epWAT ( H ) and scWAT ( I ). Representative H&E-stained sections of epWAT ( J ) and quantitative results ( K ). The weights of adipose tissues and the whole body ( L and M ). Data are presented as mean ± SD. p Values were determined using the unpaired two-tailed Student’s t test. ns indicates no significant difference ( p > 0.05). CREBH, cAMP-responsive element–binding protein H; epWAT, epididymal white adipose tissue; FGF21, fibroblast growth factor 21; LCK, liver-specific Rab2A knockout; scWAT, subcutaneous white adipose tissue.

Article Snippet: The following primary and secondary antibodies were used: goat anti FGF21 (R&D Systems; AF3057), mouse anti-Rab2A (Proteintech; 67501-1-Ig), mouse anti-tubulin (DUONENG-BIO; AB0178801), mouse anti-actin (Zen-Bioscience; 200068-8F10), rabbit anti-CREBH (Kerafast; EWS101), rabbit anti-GAPDH (Proteintech; 10494-1-AP), rabbit anti-APOB (Proteintech; 20578-1-AP), mouse anti-CHC (Santa Cruz; sc-12734), rabbit anti-GRP94 (Proteintech; 14700-1-AP), mouse anti-GRASP55 (Santa Cruz; sc-271840), HRP-conjugated goat anti-mouse IgG (Jackson ImmunoResearch; 115-035-003), HRP-conjugated goat anti-rabbit IgG (Jackson ImmunoResearch; 111-035-003), and HRP-conjugated rabbit anti-goat IgG (Jackson ImmunoResearch; 305-035-003).

Techniques: Knockdown, Expressing, Virus, Injection, Western Blot, Staining, Two Tailed Test, Binding Assay, Knock-Out

Journal: The Journal of Biological Chemistry

Article Title: Rab2A modulates liver fibroblast growth factor 21 (FGF21) expression and systemic metabolism via apolipoprotein B–CREBH signaling

doi: 10.1016/j.jbc.2025.110977

Figure Lengend Snippet: APOB knockdown rescues CREBH activation, FGF21 expression, and the metabolic phenotypes of LCK mice . A , schematic of the experimental timeline for adeno-associated virus (AAV) injection in Flox and LCK mice. B , representative immunoblots of hepatic APOB protein levels after AAV-mediated knockdown. C and D , representative immunoblots and quantification of hepatic CREBH cleavage levels after APOB knockdown. E – M , analyses performed after 8 weeks of AAV injection and subsequent euthanasia (male, n = 5 versus 5 versus 5): hepatic mRNA levels of Creb3l3 , Fgf21 , Apoa4 , Fsp27β , and Apoc2 ( E ). Representative immunoblots and quantification of hepatic FGF21 protein levels ( F and G ). Serum FGF21 levels ( H ). mRNA expression of thermogenic-related genes in scWAT ( I ). Representative H&E-stained sections of epWAT ( J ) and quantitative results ( K ). The weights of adipose tissues and the whole body ( L and M ). Data are presented as mean ± SD. p Values were determined using the unpaired two-tailed Student’s t test. ns indicates no significant difference ( p > 0.05). AAV, adeno-associated virus; APOB, apolipoprotein B; CREBH, cAMP-responsive element–binding protein H; epWAT, epididymal white adipose tissue; FGF21, fibroblast growth factor 21; LCK, liver-specific Rab2A knockout; scWAT, subcutaneous white adipose tissue.

Article Snippet: The following primary and secondary antibodies were used: goat anti FGF21 (R&D Systems; AF3057), mouse anti-Rab2A (Proteintech; 67501-1-Ig), mouse anti-tubulin (DUONENG-BIO; AB0178801), mouse anti-actin (Zen-Bioscience; 200068-8F10), rabbit anti-CREBH (Kerafast; EWS101), rabbit anti-GAPDH (Proteintech; 10494-1-AP), rabbit anti-APOB (Proteintech; 20578-1-AP), mouse anti-CHC (Santa Cruz; sc-12734), rabbit anti-GRP94 (Proteintech; 14700-1-AP), mouse anti-GRASP55 (Santa Cruz; sc-271840), HRP-conjugated goat anti-mouse IgG (Jackson ImmunoResearch; 115-035-003), HRP-conjugated goat anti-rabbit IgG (Jackson ImmunoResearch; 111-035-003), and HRP-conjugated rabbit anti-goat IgG (Jackson ImmunoResearch; 305-035-003).

Techniques: Knockdown, Activation Assay, Expressing, Virus, Injection, Western Blot, Staining, Two Tailed Test, Binding Assay, Knock-Out

Journal: The Journal of Biological Chemistry

Article Title: Rab2A modulates liver fibroblast growth factor 21 (FGF21) expression and systemic metabolism via apolipoprotein B–CREBH signaling

doi: 10.1016/j.jbc.2025.110977

Figure Lengend Snippet: Starvation-induced APOB trafficking defects activate the CREBH–FGF21 axis . Wildtype C57BL/6J mice were fasted for 16 h or not prior to analysis. A , hepatic triglyceride (TG) levels (n = 6 per group). B , VLDL–TG secretion rates (n = 5 per group). C , representative immunoblots of total hepatic APOB. D , quantification of APOB protein levels from ( C ). E , representative immunoblots of APOB distribution in the endoplasmic reticulum (ER) and Golgi apparatus fractions. F , quantification of APOB levels in subcellular fractions from ( E ). G , representative immunoblots of CREBH cleavage. H , quantification of cleaved CREBH from ( G ). I , hepatic mRNA levels of Fgf21 , Fsp27β , and Apoa4 (n = 4 per group). J , representative immunoblots of hepatic FGF21 proteins. K , quantification of FGF21 protein levels from ( J ). L , serum FGF21 levels (n = 5 per group). Data are presented as mean ± SD. p Values were determined using the unpaired two-tailed Student’s t test. APOB, apolipoprotein B; CREBH, cAMP-responsive element–binding protein H; FGF21, fibroblast growth factor 21; TG, triglyceride; VLDL, very low–density lipoprotein.

Article Snippet: The following primary and secondary antibodies were used: goat anti FGF21 (R&D Systems; AF3057), mouse anti-Rab2A (Proteintech; 67501-1-Ig), mouse anti-tubulin (DUONENG-BIO; AB0178801), mouse anti-actin (Zen-Bioscience; 200068-8F10), rabbit anti-CREBH (Kerafast; EWS101), rabbit anti-GAPDH (Proteintech; 10494-1-AP), rabbit anti-APOB (Proteintech; 20578-1-AP), mouse anti-CHC (Santa Cruz; sc-12734), rabbit anti-GRP94 (Proteintech; 14700-1-AP), mouse anti-GRASP55 (Santa Cruz; sc-271840), HRP-conjugated goat anti-mouse IgG (Jackson ImmunoResearch; 115-035-003), HRP-conjugated goat anti-rabbit IgG (Jackson ImmunoResearch; 111-035-003), and HRP-conjugated rabbit anti-goat IgG (Jackson ImmunoResearch; 305-035-003).

Techniques: Western Blot, Two Tailed Test, Binding Assay

Journal: The Journal of Biological Chemistry

Article Title: Rab2A modulates liver fibroblast growth factor 21 (FGF21) expression and systemic metabolism via apolipoprotein B–CREBH signaling

doi: 10.1016/j.jbc.2025.110977

Figure Lengend Snippet: Summary of results and work model . A graphical abstract was created to summarize the key findings of this study, outlining a multistep regulatory pathway. VLDL assembly and maturation (steps 1–3): newly assembled VLDL 2 particles are transported from the ER to the Golgi via COPII vesicles. At the Golgi, the Rab2A–HSD17B13 complex mediates lipid droplet–Golgi contacts, facilitating lipid transfer to support VLDL 1 maturation. CREBH activation and signaling (steps 4–6): full-length CREBH resides in the ER and is transported to the Golgi under specific conditions, where its proteolytic cleavage—partially dependent on APOB availability—generates the active N-terminal fragment. This fragment translocates to the nucleus and drives transcription of Fsp27β , Fgf21 , and Apoa4 . Systemic metabolic regulation (steps 7–9): secreted FGF21 promotes adipose tissue lipolysis; ApoA-IV enhances VLDL lipidation and expansion in the Golgi; and lipid droplet–localized FSP27β facilitates hepatic lipid storage. Pathophysiological perturbations: in Rab2A-deficient hepatocytes, disrupted lipid droplet–Golgi contacts reduce lipid flux to the Golgi, impairing VLDL maturation. This leads to APOB accumulation in the Golgi, which promotes CREBH cleavage and elevates FGF21 expression. During fasting, AMPK-mediated suppression of Rab2A activity, combined with partial inhibition of APOB ER-to-Golgi transport, further reduces VLDL secretion. These changes are sensed by CREBH, enhancing its activation and driving a transcriptional program that adaptively regulates systemic lipid homeostasis. Graphical abstract created with BioRender (Chen, 2025; https://BioRender.com/7gyj0x2 ). AMPK, AMP-activated protein kinase; APOB, apolipoprotein B; COPII, coat protein complex II; CREBH, cAMP-responsive element–binding protein H; ER, endoplasmic reticulum; VLDL, very low–density lipoprotein.

Article Snippet: The following primary and secondary antibodies were used: goat anti FGF21 (R&D Systems; AF3057), mouse anti-Rab2A (Proteintech; 67501-1-Ig), mouse anti-tubulin (DUONENG-BIO; AB0178801), mouse anti-actin (Zen-Bioscience; 200068-8F10), rabbit anti-CREBH (Kerafast; EWS101), rabbit anti-GAPDH (Proteintech; 10494-1-AP), rabbit anti-APOB (Proteintech; 20578-1-AP), mouse anti-CHC (Santa Cruz; sc-12734), rabbit anti-GRP94 (Proteintech; 14700-1-AP), mouse anti-GRASP55 (Santa Cruz; sc-271840), HRP-conjugated goat anti-mouse IgG (Jackson ImmunoResearch; 115-035-003), HRP-conjugated goat anti-rabbit IgG (Jackson ImmunoResearch; 111-035-003), and HRP-conjugated rabbit anti-goat IgG (Jackson ImmunoResearch; 305-035-003).

Techniques: Activation Assay, Expressing, Activity Assay, Inhibition, Binding Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: MOB1 deletion in murine mature adipocytes ameliorates obesity and diabetes.

doi: 10.1073/pnas.2424741122

Figure Lengend Snippet: Fig. 4. Increased beiging and β-oxidation of iWAT, and elevated energy expenditure, are induced by MOB1 deficiency. (A) mRNA levels of the indicated brown/ beige marker genes and beige regulator genes in iWAT of HFD-aMob1DKO mice (n = 4) after 20 wk of HFD feeding. mRNA levels were determined by microarray analysis. Data are expressed as a ratio compared to levels in iWAT of HFD-Cont mice (n = 4; 20 wk HFD feeding). *P < 0.05, **P < 0.01. (B, Top) Immunoblots to detect UCP1, PGC1α, β3-AR, FGF21, PPARα, and OPA1 proteins in iWAT from HFD-Cont and HFD-aMob1DKO mice (n = 9/group) after 20 wk of HFD feeding. HSP90, loading control. Bottom: Densitometric relative quantification of the indicated proteins in the blots in the top panels. *P < 0.05, **P < 0.01. (C) Serum norepinephrine levels in overnight-fasted HFD-Cont (n = 17) and HFD-aMob1DKO (n = 9) mice after 20 wk of HFD feeding. n.s., not significant. (D) mRNA levels of the indicated FAO-related genes in iWAT of HFD-aMob1DKO mice (n = 4) after 20 wk of HFD feeding. mRNA levels were determined by microarray analysis. Data are expressed as the ratio compared to levels in HFD-Cont mice (n = 4; 20 wk HFD feeding). *P < 0.05, **P < 0.01. (E–G) Monitoring and quantification of VO2 (E), VCO2 (F), and energy expenditure (EE) (G) during light (l) and dark (d) phases in HFD-Cont and HFD-aMob1DKO mice (n = 7/group) after HFD feeding for 20 wk. For part of the dark phase experiment, the temperature was reduced to 4 °C. In E, the arrow indicates the timing of administration of the β3-agonist CL316,243 (n = 5/group). *P < 0.05, **P < 0.01.

Article Snippet: HFD- aMob1DKO mice and HFD- Cont mice (17 wk old) received intraperitoneal administration of anti- FGF21 neutralizing antibody (R&D Systems, AF3057) (15 μg/mouse/dose) or IgG control (R&D Systems, AB- 108- C) (15 μg/mouse/dose) on experimental days 1, 4, 7, 10, 13, 16, and 19.

Techniques: Marker, Microarray, Western Blot, Control, Quantitative Proteomics

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: MOB1 deletion in murine mature adipocytes ameliorates obesity and diabetes.

doi: 10.1073/pnas.2424741122

Figure Lengend Snippet: Fig. 7. Activation of a YAP1-FGF21-OPA1 axis and FGF21 dependence of the MOB1-deficient mouse phenotype. (A, Left) Immunoblot to detect FGF21 and OPA1 proteins in iWAT from HFD-aMob1DKO and HFD-aMob1/Yap1TKO mice (n = 6/group) after 20 wk of HFD feeding. HSP90, loading control. Right: Densitometric relative quantification of the indicated proteins in the blots in the Left panels *P < 0.05, **P < 0.01. (B) qPCR to detect mRNA levels of the indicated genes in 3T3L1 cells overexpressing YAP5SA (Dox+) or not (Dox−). Data are expressed as the fold increase in mRNA expression in Dox+ cells over Dox− cells. Data are from four independent trials. *P < 0.05, **P < 0.01. (C, Left) Immunoblot to detect FGF21, OPA1, and YAP1 proteins in 3T3L1 cells overexpressing YAP5SA (Dox+) or not (Dox−) (n = 4/group). Exo, exogenous; endo, endogenous. HSP90, loading control. Right: Densitometric relative quantification of the indicated proteins in the blots in the Left panel. *P < 0.05, **P < 0.01. (D) Left: Immunoblot to detect OPA1 proteins in 3T3L1 cells overexpressing YAP5SA (Dox+) in the presence of either Scramble siRNA, Fgf21siRNA #1, or Fgf21siRNA #2 (n = 6/group). HSP90, loading control. Right: Densitometric relative quantification of OPA1 proteins in the blots in the Left panel. *P < 0.05. (E) Quantitative evaluation of ChIP assays used to detect binding of the YAP1 complex to TBS-containing DNA upstream of the Fgf21 gene in 3T3L1 cells overexpressing YAP5SA (Dox+) (n = 3/group). Immunoprecipitation by control IgG and DNA downstream of the Fgf21 gene lacking a TBS served as a negative control. DNA upstream of the Ctgf gene containing a TBS was used as a positive control. Data are relative enrichment values. *P < 0.05, **P < 0.01. (F) Quantitation of relative Fgf21-luciferase reporter activity in 293T cells overexpressing YAP5SA (“YAP5SA”) or not (“control”) when the luciferase vector contained Fgf21 promoter DNA (−289 to −1) plus WT TBS1 and TBS2; the same Fgf21 DNA (−289 to −1) plus mutated TBS1; the same Fgf21 DNA (−289 to −1) plus mutated TBS2; or control DNA downstream of Fgf21 (+1,813 ~+ 2,101) (n = 3/group). Please see SI Appendix, Fig. S11 A and B. *P < 0.05. (G) Changes in body weight of HFD-aMob1DKO (n = 7/group) and HFD-Cont (n = 6/group) male mice that were fed HFD for 20 wk and then injected i.p. on day 0 with either neutralizing anti-FGF21 antibody or normal IgG. Data are representative of two independent experiments. *P < 0.05. (H) Basal lipolysis as determined by free glycerol (Left) and free FAs (Right) spontaneously released from iWAT of the antibody-injected HFD-aMob1DKO (n = 7/group) and HFD-Cont (n = 6/group) mice in G). **P < 0.01. Data are representative of two independent experiments. (I) ROS production as determined by MFI of labeled H2DCFDA in iWAT of the antibody- injected HFD-aMob1DKO (n = 7/group) and HFD-Cont (n = 6/group) mice in G. *P < 0.05. Data are representative of two independent experiments.

Article Snippet: HFD- aMob1DKO mice and HFD- Cont mice (17 wk old) received intraperitoneal administration of anti- FGF21 neutralizing antibody (R&D Systems, AF3057) (15 μg/mouse/dose) or IgG control (R&D Systems, AB- 108- C) (15 μg/mouse/dose) on experimental days 1, 4, 7, 10, 13, 16, and 19.

Techniques: Activation Assay, Western Blot, Control, Quantitative Proteomics, Expressing, Binding Assay, Immunoprecipitation, Negative Control, Positive Control, Quantitation Assay, Luciferase, Activity Assay, Plasmid Preparation, Injection, Labeling