Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: The double‐edged sword of endoplasmic reticulum stress in uremic sarcopenia through myogenesis perturbation

doi: 10.1002/jcsm.12288

Figure Lengend Snippet: IS treatment represses myoD expression and Akt phosphorylation during myoblast (MB) proliferation and myogenic differentiation (MD). (A and B) Protein expression of myoD in myoblsts treated with or without IS (1 mM) for 48 h is examined by western blot and quantified. (C and D) pPotein expression of myoD in differentiated myoblasts treated with or without IS (1 mM) at the indicated time points by western blot and quantified. (E–F) Akt expression and phosphorylation in myoblasts treated with or without IS (1 mM) for 48 h are determined by western blot and quantified. (G–H) Akt expression and phosphorylation during myoblasts differentiation treated with or without IS (1 mM) at the indicated time point are determined by western blot and quantified. Data represent means ± SEM for three independent experiments. * P < 0.05, ** P < 0.01, and *** P < 0.001, as compared with untreated control.

Article Snippet: Atrogin 1 (ab168372) from Abcam (Cambridge, MA); myoD (5.8A) (NB100‐56511) from Novus Biologicals (Littleton CO), and Myc (CSB‐MA000041M0m) from Cusabio (China).

Techniques: Expressing, Phospho-proteomics, Western Blot, Control

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: The double‐edged sword of endoplasmic reticulum stress in uremic sarcopenia through myogenesis perturbation

doi: 10.1002/jcsm.12288

Figure Lengend Snippet: Salubrinal, an eIF2α dephosphorylation inhibitor, deregulates C2C12 myogenic differentiation (MD). (A and B) Myoblasts (MB) were treated with or without 20 μM salubrinal for 48 h. MyoD, phospho‐eIF2α, and phospho‐AKT expressions are examined with western blot and quantified. (C and D) Myoblasts under 48 h differentiation are simultaneously exposed with or without 20 μM salubrinal. MyoG, MyHC, phospho‐AKT, and phospho‐ eIF2α expression level are determined with western blot and quantified. (E and F) Myoblasts are overexpressed with or without mouse S52D phosphomimetic eIF2α mutant for 48 h. The expression of eIF2α S52D was detected with anti‐myc antibody. MyoD and phospho‐AKT expressions are examined with western blot and quantified. (G and H) myoblasts under 48 h differentiation are simultaneously transfected with or without S52D mutant. The expression of eIF2α S52D was detected with anti‐myc antibody. MyoG, MyHC, and phospho‐AKT expression level are determined with western blot and quantified.

Article Snippet: Atrogin 1 (ab168372) from Abcam (Cambridge, MA); myoD (5.8A) (NB100‐56511) from Novus Biologicals (Littleton CO), and Myc (CSB‐MA000041M0m) from Cusabio (China).

Techniques: De-Phosphorylation Assay, Western Blot, Expressing, Mutagenesis, Transfection

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: The double‐edged sword of endoplasmic reticulum stress in uremic sarcopenia through myogenesis perturbation

doi: 10.1002/jcsm.12288

Figure Lengend Snippet: Model of IS‐induced defect in myoblast differentiation. The pro‐ and anti‐myogenesis roles of XBP1 and phospho‐eIF2α in different stages of myogenesis are pointed out. Notably, the ROS‐eIF2α axis is important for the antimyogenesis activity of IS. The red line indicates translational attenuation caused by phosphorylation of eIF2α probably results in decreased myoD expression. Overall, our research demonstrates the pivotal role of UPR signalling in age‐related muscle loss and sarcopenia.

Article Snippet: Atrogin 1 (ab168372) from Abcam (Cambridge, MA); myoD (5.8A) (NB100‐56511) from Novus Biologicals (Littleton CO), and Myc (CSB‐MA000041M0m) from Cusabio (China).

Techniques: Activity Assay, Phospho-proteomics, Expressing

Journal: Nature Communications

Article Title: MYOD-SKP2 axis boosts tumorigenesis in fusion negative rhabdomyosarcoma by preventing differentiation through p57 Kip2 targeting

doi: 10.1038/s41467-023-44130-0

Figure Lengend Snippet: a A model showing that in myoblasts and RMS cells that are both proliferating, H3K27ac (Ac) enrichment at SKP2 DNA regulatory regions allows MYOD, which is bound on these regions, to induce the expression of SKP2 thus supporting proliferation (left). Upon differentiation cues, reduction of H3K27ac enrichment results in a closed chromatin conformation, impairing MYOD ability to induce SKP2 transcription. These events promote differentiation, which is inhibited in RMS (right). b A model showing that in RMS cells, MYOD induces the expression of SKP2 , which in turn associates with the SCF/CRL1 complex to promote ubiquitylation and proteasomal degradation of p27 Kip1 and p57 Kip2 by direct interaction and p21 Cip1 in an indirect way, promoting oncogenic features (left). These SKP2 functions can be suppressed by SKP2 silencing (upper right) or by its functional inhibition by the SKP2 inhibitor SMIP004 or the NEDDylation inhibitor MLN4924 (lower right) to block RMS oncogenic properties. NAE NEDD8-activating enzyme, Ub Ubiquitin, N8 NEDD8. Created with BioRender.com.

Article Snippet: Antibodies against SKP2 (H-435, sc-7164, 1:500), p27 Kip1 (F-8, sc-1641, 1:500), MYOD (5–8 A, sc-32758, 1:200), pMYOD (Ser-200) (sc-101741, 1:200) and β- ACTIN (C4, sc-47778, 1:1000) were from Santa Cruz Biotechnology Inc., (Santa Cruz, CA, USA); p21 Cip1 (12D1, #2947, 1:1000), Nedd8 (19E3, #2754, 1:1000) and GAPDH (D16H11, #5174, 1:2000) were from Cell Signaling Technology, Inc., (Danvers, MA, USA); MYOG (F5D-c, 1:200) was from DSHB (University of Iowa, Iowa City, IA, USA); α-TUBULIN (NB100-680, 1:5000) was from Novus Biologicals (Littleton, CO, USA); VINCULIN (hVIN-1; V9131, 1:5000) was from Sigma (Sigma-Aldrich, St Louis, MO, USA); p57 Kip2 (#556346, 1:500) were from BD Biosciences (San Jose‘, CA, USA).

Techniques: Expressing, Functional Assay, Inhibition, Blocking Assay, Ubiquitin Proteomics