Journal: Molecular Metabolism

Article Title: ERRα fosters running endurance by driving myofiber aerobic transformation and fuel efficiency

doi: 10.1016/j.molmet.2023.101814

Figure Lengend Snippet: ERRα promotes oxidative muscle fibers and angiogenesis. (A) Representative photographs showing muscle appearances of ERRα 3SA mice and their littermate controls. Quad: quadriceps; Gas: gastrocnemius; Sol: soleus. (B,C) Representative Laminin stained (B) and succinate dehydrogenase (SDH) stained (C) cross-sections of quadriceps from ERRα 3SA mice and their littermate controls. Scale bar represents 50 μm. (D) Relative levels of mitochondrial DNA (mtDNA) to nuclear DNA (nDNA) of quadriceps from ERRα 3SA and WT littermates (n = 7–8). (E) Citrate synthase (CS) activity in quadriceps from ERRα 3SA and WT littermates (n = 6). (F) Immunoblots of mitochondrial Oxphos proteins in quadriceps from WT and ERRα 3SA mice. Each lane represents one mouse, n = 3. (G,H) mRNA (G, n = 4–5) and protein (H, each lane represents one mouse, n = 3) levels of genes encoding distinct myosin isoforms in quadriceps of ERRα 3SA and WT littermates. (I) Representative MyHC IIA and IIB immunofluorescence staining of cross-sections of quadriceps from ERRα 3SA mice and their littermate controls. MyHC IIX appears as unstained. Below, enlarged magnification for each muscle. Scale bar represents 500 μm. Quantification of fibers expressed as percentage of fibers is indicated for each MyHC. (J) mRNA levels of genes encoding distinct troponin isoforms in quadriceps of ERRα 3SA and WT littermates, n = 4–5. (K,L) mRNA (K, n = 4–5) and protein (L, each lane represents one mouse, n = 3) levels of myoglobin and angiogenesis genes in quadriceps of ERRα 3SA and WT littermates. (M) Quadriceps from ERRα 3SA and control mice were fluorescently stained for CD31. Nuclei were counterstained with DAPI. Scale bar represents 20 μm. Data are presented as means ± SEM, ∗p < 0.05, unpaired two-tailed Student's t test (D,E,G,I-K).

Article Snippet: The following antibodies were used: ERRα (1:1000 dilution; cat. no. ab76228; Abcam); phospho-ERRα (Ser19) (1:1000 dilution, homemade [ ]; Phospho-AMPKα (Thr172) (1:1000 dilution; cat. no. 2535S; Cell Signaling Technology); AMPKα (1:1000 dilution; cat. no. 2532S; Cell Signaling Technology); Vinculin (1:2000 dilution; cat. no. MAB3574; clone VIIF9; Sigma–Aldrich); Tubulin (1:5000 dilution; cat. no. CLT9002; clone DM1A; Cedarlane); PGC1α (1:200 dilution; cat. no. sc-13067; Santa Cruz Biotechnology); LDHA (1:1000 dilution; cat. no. 2012S; Cell Signaling Technology); LDHB (1:1000 dilution; cat. no. A7625; ABclonal); MCT1 (1:1000 dilution; cat. no. A3013; ABclonal); MCT4 (1:1000 dilution; cat. no. A10548; ABclonal); total OxPhos rodent antibody cocktail (1:2000 dilution; cat. no. ab110413 (MS604); Abcam); MyHC I (1:2000 dilution; cat. no. BA-D5; DSHB); MyHC IIA (1:2000 dilution; cat. no. SC-71; DSHB); MyHC IIB (1:2000 dilution; cat. no. BF-F3; DSHB); Myoglobin (1:1000 dilution; cat. no. A5471; ABclonal); VEGFA (1:1000 dilution; cat. no. A5708; ABclonal); FABP3 (1:1000 dilution; cat. no. A5312; ABclonal); ACADL (1:2000 dilution; cat. no. A1266; ABclonal); PDK4 (1:1000 dilution; cat. no. A13337; ABclonal); phospho-HSL (Ser563) (1:1000 dilution; cat. no. 4139; Cell Signaling Technology); phospho-HSL (Ser660) (1:1000 dilution; cat. no. 4126; Cell Signaling Technology); HSL (1:1000 dilution; cat. no. 41078; Cell Signaling Technology); ATGL (1:1000 dilution; cat. no. A6245; ABclonal); Perilipin-1 (1:1000 dilution; cat. no. 9349; Cell Signaling Technology); NDUFB3 (1:1000 dilution; cat. no. A14378; ABclonal); SDHB (1:5000 dilution; cat. no. ab14714; Abcam); UQCRQ (1:1000 dilution; cat. no. A9872; ABclonal); COX7A2 (1:1000 dilution; cat. no. A8406; ABclonal); ATP5K (1:1000 dilution; cat. no. A16769; ABclonal); Cytochrome c (1:1000 dilution; cat. no. A13430; ABclonal).

Techniques: Staining, Activity Assay, Western Blot, Immunofluorescence, Control, Two Tailed Test