Journal: Nature Communications

Article Title: Inhibition of MYC by the SMARCB1 tumor suppressor

doi: 10.1038/s41467-019-10022-5

Figure Lengend Snippet: SNF5 inhibits DNA-binding by MYC. a Recombinant MYC:MAX or MAX:MAX complexes were incubated with increasing amounts of recombinant SNF5 (3-, 6-, 12-, 24-fold molar excess over MYC:MAX) and EMSA performed using a DNA probe carrying a wild-type E-box sequence. Note that MYC:MAX complexes also contain residual MAX:MAX dimers. Lane 1 shows that incubation with the MYC interactor WDR5 (12-fold molar excess) induces a supershift in the MYC:MAX complexes. Lane 13 is the top concentration of SNF5, without MYC or MAX proteins. b EMSA, as in a , except MYC:MAX complexes were incubated with either recombinant SNF5 (12-, 24-fold molar excess) or an SNF5 mutant lacking the imperfect repeats region (amino acids 176–309) that interact with MYC (∆RPT; 12-, 24-fold molar excess). Results were confirmed with two independent preparations of recombinant proteins. c SMARCB1 -null (KO) HEK293 cells were transduced to express FKBP12 (F36V)− HA-SNF5, treated with DMSO or 500 nM of dTAG-47 for the indicated times, lysates prepared, and the levels of HA-tagged degradable SNF5 determined by immunoblotting. Lysate from the KO cells is included as a control in lane 1. GAPDH is a loading control. d SMARCB1 -null (KO) HEK293 cells were transduced to express FKBP12 (F36V)− HA-SNF5 (+SNF5) and treated with DMSO or dTAG-47 (500 nM) for 2 h. ChIP assays were then performed. IgG control is shown for the dTAG-47-treated sample. Anti-MYC ChIPs are shown for the KO and the treated +SNF5 cells. Relative MYC binding is calculated as the signal at each of the indicated loci, relative to a non-MYC-bound locus (β-globin). n = 3 independent ChIP experiments. Error bars are standard error

Article Snippet: Each immunoprecipitation was performed on chromatin collected from 10 × 10 6 cells by dilution in ten volumes of FALB buffer (50 mM HEPES, pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton) using antibodies against MYC (N262, Santa Cruz Biotechnology, sc-764 or Cell Signaling, 9402; 3 μg) or normal rabbit IgG control (Cell Signaling, 2729 s; 3 μg).

Techniques: Binding Assay, Recombinant, Incubation, Sequencing, Concentration Assay, Mutagenesis, Western Blot, Control

Journal: Scientific reports

Article Title: The kisspeptin-1 receptor antagonist peptide-234 aggravates uremic cardiomyopathy in a rat model.

doi: 10.1038/s41598-023-41037-0

Figure Lengend Snippet: Figure 1. Experiment setup. Blood: blood sampling, BP: blood pressure, Urine: urine sampling, Echo: echocardiography, CKD: chronic kidney disease, PBS: phosphate-buffered saline, P234: KISS1R antagonist peptide-234, LV: left ventricle, Op: operation. Sham: sham-operated group, CKD: chronic kidney disease group, CKD + P234 D1: chronic kidney disease group treated with the lower dose (13 μg/day, dose 1) of KISS1R antagonist peptide-234, CKD + P234 D2: chronic kidney disease group treated with the higher dose (26 μg/day, dose 2) of KISS1R antagonist peptide-234.

Article Snippet: The time course and doses of P234 (Tocris Bioscience, Bristol, UK, catalog No.: 3881, molecular weight: 1295.42 g/mol; peptide sequence: Ac-DAla-Asn-Trp-Asn-Gly-Phe-Gly-D-Trp-Arg-Phe-NH2) were selected based on our preliminary data and previous studies26,44.

Techniques: Sampling, Saline

Journal: Scientific reports

Article Title: The kisspeptin-1 receptor antagonist peptide-234 aggravates uremic cardiomyopathy in a rat model.

doi: 10.1038/s41598-023-41037-0

Figure Lengend Snippet: Figure 2. The effects of the KISS1R antagonist peptide-234 on the development of CKD in 5/6 nephrectomized rats. (A) Serum urea concentration, (B) serum creatinine concentration, (C) creatinine clearance, (D) 24-h urinary protein excretion, (E) 24-h urinary creatinine excretion, and (F) 24-h urine volume at the endpoint. Values are presented as mean ± S.E.M., *p < 0.05 vs. sham group (n = 7–8, One-Way ANOVA, Holm-Sidak post hoc test), $p < 0.05 vs. the week 5 values in the same group (n = 7–8, Two-Way Repeated Measures ANOVA, Holm-Sidak post hoc test). Creatinine clearance was calculated according to the standard formula (urine creatinine concentration [μM] × urine volume for 24 h [mL])/(serum creatinine concentration [μM] × 24 × 60 min). ¥At the endpoint, urine volume and creatinine concentration were measured at week 12 and serum creatinine concentration at week 13. Sham: sham-operated group, CKD: chronic kidney disease group, CKD + P234 D1: chronic kidney disease group treated with the lower dose (13 μg/day, dose 1) of KISS1R antagonist peptide-234, CKD + P234 D2: chronic kidney disease group treated with the higher dose (26 μg/day, dose 2) of KISS1R antagonist peptide-234.

Article Snippet: The time course and doses of P234 (Tocris Bioscience, Bristol, UK, catalog No.: 3881, molecular weight: 1295.42 g/mol; peptide sequence: Ac-DAla-Asn-Trp-Asn-Gly-Phe-Gly-D-Trp-Arg-Phe-NH2) were selected based on our preliminary data and previous studies26,44.

Techniques: Concentration Assay

Journal: Scientific reports

Article Title: The kisspeptin-1 receptor antagonist peptide-234 aggravates uremic cardiomyopathy in a rat model.

doi: 10.1038/s41598-023-41037-0

Figure Lengend Snippet: Figure 3. The effects of the KISS1R antagonist peptide-234 on the echocardiographic parameters. (a) Representative M-mode images, (b) systolic posterior wall thickness (PWTs), (c) diastolic posterior wall thickness (PWTd), (d) Representative pulse wave and tissue Doppler images of mitral valve early flow velocity (e) and septal mitral annulus (e′) velocity, E) E/e′ ratio, (f) ejection fraction (EF). Values are presented as mean ± S.E.M., *p < 0.05 vs. sham, #p < 0.05 vs. CKD (n = 7–9, One-Way ANOVA, Holm-Sidak post hoc test). $p < 0.05 vs. week 5 values in the same group (n = 7–8, Two-Way Repeated-Measures ANOVA, Holm-Sidak post hoc test). Sham: sham-operated group, CKD: chronic kidney disease group, CKD + P234 D1: chronic kidney disease group treated with the lower dose (13 μg/day, dose 1) of KISS1R antagonist peptide-234, CKD + P234 D2: chronic kidney disease group treated with the higher dose (26 μg/day, dose 2) of KISS1R antagonist peptide-234.

Article Snippet: The time course and doses of P234 (Tocris Bioscience, Bristol, UK, catalog No.: 3881, molecular weight: 1295.42 g/mol; peptide sequence: Ac-DAla-Asn-Trp-Asn-Gly-Phe-Gly-D-Trp-Arg-Phe-NH2) were selected based on our preliminary data and previous studies26,44.

Techniques:

Journal: Scientific reports

Article Title: The kisspeptin-1 receptor antagonist peptide-234 aggravates uremic cardiomyopathy in a rat model.

doi: 10.1038/s41598-023-41037-0

Figure Lengend Snippet: Figure 4. The effects of the KISS1R antagonist peptide-234 on cardiomyocyte hypertrophy and interstitial fibrosis and molecular markers of heart failure at week 13. (a) Representative hematoxylin–eosin (HE)-stained slides at 100 × and 40 × magnifications and representative picrosirius red and fast green (PSFG)-stained slides at 20 × magnifcation (b) cardiomyocyte cross-sectional areas, (c) left ventricular collagen content, (d) A-type natriuretic peptide (Nppa), and (E) matrix metalloproteinase-9 (Mmp9) expressions in the left ventricles normalized to the ribosomal protein lateral stalk subunit P2 (Rplp2) gene expression. On the digital HE images, cardiomyocyte cross-sectional areas were measured in 100 selected cardiomyocytes on left ventricular sections cut on the same plane. The mean values of the collagen content of 10 representative PSFG-stained images were calculated and used for statistical evaluation in the case of each left ventricular slide. Scale bars represent 10 µm at the 100 × magnifed images, 20 µm at the 40 × magnifed images, and 50 µm at the 20 × magnifed images. Values are presented as mean ± S.E.M., *p < 0.05 vs. sham, #p < 0.05 vs. CKD (n = 7–8, one-way ANOVA, Holm-Sidak post hoc test). Sham: sham-operated group, CKD: chronic kidney disease group, CKD + P234 D1: chronic kidney disease group treated with the lower dose (13 μg/day, dose 1) of KISS1R antagonist peptide-234, CKD + P234 D2: chronic kidney disease group treated with the higher dose (26 μg/day, dose 2) of KISS1R antagonist peptide-234.

Article Snippet: The time course and doses of P234 (Tocris Bioscience, Bristol, UK, catalog No.: 3881, molecular weight: 1295.42 g/mol; peptide sequence: Ac-DAla-Asn-Trp-Asn-Gly-Phe-Gly-D-Trp-Arg-Phe-NH2) were selected based on our preliminary data and previous studies26,44.

Techniques: Staining, Gene Expression

Journal: Scientific reports

Article Title: The kisspeptin-1 receptor antagonist peptide-234 aggravates uremic cardiomyopathy in a rat model.

doi: 10.1038/s41598-023-41037-0

Figure Lengend Snippet: Figure 5. The effects of the KISS1R antagonist peptide-234 on the left ventricular expression of genes associated with inflammation and fibrosis. Relative gene expression of (a) interleukin-1 (Il1), (b), interleukin-6 (Il6), (c) tumor necrosis factor-α (Tnf), (d) connective tissue growth factor (Ctgf), (e) transforming growth factor-β (Tgfb), (f) collagen type 1 alpha 1 chain (Col1a1) and (g), collagen type 3 alpha 1 chain (Col3a1) normalized to the ribosomal protein lateral stalk subunit P2 (Rplp2) gene expression. Values are presented as mean ± S.E.M., *p < 0.05 vs. sham, #p < 0.05 vs. CKD (n = 7–8, One-Way ANOVA, Holm-Sidak post hoc test). Sham: sham- operated group, CKD: chronic kidney disease group, CKD + P234 D1: chronic kidney disease group treated with the lower dose (13 μg/day, dose 1) of KISS1R antagonist peptide-234, CKD + P234 D2: chronic kidney disease group treated with the higher dose (26 μg/day, dose 2) of KISS1R antagonist peptide-234.

Article Snippet: The time course and doses of P234 (Tocris Bioscience, Bristol, UK, catalog No.: 3881, molecular weight: 1295.42 g/mol; peptide sequence: Ac-DAla-Asn-Trp-Asn-Gly-Phe-Gly-D-Trp-Arg-Phe-NH2) were selected based on our preliminary data and previous studies26,44.

Techniques: Expressing, Gene Expression

Journal: Scientific reports

Article Title: The kisspeptin-1 receptor antagonist peptide-234 aggravates uremic cardiomyopathy in a rat model.

doi: 10.1038/s41598-023-41037-0

Figure Lengend Snippet: Figure 6. The effects of the KISS1R antagonist peptide-234 on the protein levels of KISS1R and ERK1/2 at week 13. Left ventricular protein levels and cropped representative Western blot imagines of (a) Kisspeptin receptor-1 (KISS1R, 40–140 kDa), (b) total ERK1 (44 kDa), (c) phospho-ERK1 (pERK1, 44 kDa), (d) pERK1/ERK1 ratio and (e) total ERK 2 (42 kDa), (f) phospho-ERK2 (pERK2, 42 kDa), (g) pERK2/ERK2 ratio. Values are presented as mean ± S.E.M., *p < 0.05 vs. sham (n = 7, One-Way ANOVA, Holm-Sidak post hoc test). Sham: sham-operated group, CKD: chronic kidney disease group, CKD + P234 D1: chronic kidney disease group treated with the lower dose (13 μg/day, dose 1) of KISS1R antagonist peptide-234, CKD + P234 D2: chronic kidney disease group treated with the higher dose (26 μg/day, dose 2) of KISS1R antagonist peptide-234. Images were captured with the Odyssey CLx machine and exported with Image Studio 5.2.5 software. The full-length Ponceau-stained membranes and the corresponding Western blot images are presented in the Supplementary Material (Figs. S3– S5).

Article Snippet: The time course and doses of P234 (Tocris Bioscience, Bristol, UK, catalog No.: 3881, molecular weight: 1295.42 g/mol; peptide sequence: Ac-DAla-Asn-Trp-Asn-Gly-Phe-Gly-D-Trp-Arg-Phe-NH2) were selected based on our preliminary data and previous studies26,44.

Techniques: Western Blot, Software, Staining

Journal: Scientific reports

Article Title: The kisspeptin-1 receptor antagonist peptide-234 aggravates uremic cardiomyopathy in a rat model.

doi: 10.1038/s41598-023-41037-0

Figure Lengend Snippet: Figure 7. The effects of the KISS1R antagonist peptide-234 on apoptosis-associated gene expressions and protein levels in the left ventricles at week 13. Relative gene expression of (a) BCL2-associated X apoptosis regulator (Bax), (b) B-Cell CLL/lymphoma 2 apoptosis regulator (Bcl2), (c) Bax/Bcl2 ratio, and (d) caspase7 (Casp7) normalized to the ribosomal protein lateral stalk subunit P2 (Rplp2) gene expression. Left ventricular protein levels and cropped representative imagines of (e) BAX (20 kDa), (f) BCL2 (26 kDa), (g) BAX/BCL2 ratio, and (h) CASP 7 (35 kDa). Values are presented as mean ± S.E.M., *p < 0.05 vs. sham, #p < 0.05 vs. CKD vehicle group (n = 7–8 for RT-qPCR and n = 7 for Western blot measurements, One-Way ANOVA, Holm-Sidak post hoc test). Sham: sham-operated group, CKD: chronic kidney disease group, CKD + P234 D1: chronic kidney disease group treated with the lower dose (13 μg/day, dose 1) of KISS1R antagonist peptide-234, CKD + P234 D2: chronic kidney disease group treated with the higher dose (26 μg/day, dose 2) of KISS1R antagonist peptide-234. Images were captured with the Odyssey CLx machine and exported with Image Studio 5.2.5 software. The full-length Ponceau-stained membranes and the corresponding Western blot images are presented in the Supplementary Material (Figs. S6–S8).

Article Snippet: The time course and doses of P234 (Tocris Bioscience, Bristol, UK, catalog No.: 3881, molecular weight: 1295.42 g/mol; peptide sequence: Ac-DAla-Asn-Trp-Asn-Gly-Phe-Gly-D-Trp-Arg-Phe-NH2) were selected based on our preliminary data and previous studies26,44.

Techniques: Gene Expression, Quantitative RT-PCR, Western Blot, Software, Staining

Journal: Cell reports

Article Title: Regulation of MYC by CARD14 in human epithelium is a determinant of epidermal homeostasis and disease

doi: 10.1016/j.celrep.2024.114589

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: ERK (rabbit) (WB (1:1000)) , Cell Signaling Technologies, Danvers, MA , Cat# 4695; RRID: AB_390779.

Techniques: FLAG-tag, Plasmid Preparation, Recombinant, Modification, Transfection, Staining, Protease Inhibitor, Blocking Assay, Western Blot, Stripping, XF Assay, Reporter Assay, Activity Assay, Bicinchoninic Acid Protein Assay, Sequencing, Cloning, Mutagenesis, Software, Membrane