Journal: bioRxiv

Article Title: Sexual identity of enterocytes regulates rapamycin-mediated intestinal homeostasis and lifespan extension

doi: 10.1101/2021.10.22.465415

Figure Lengend Snippet: a, Feminisation of male guts by expression of tra F in ECs reduced the number of Lysotracker-stained puncta in the gut, and it restored the response to rapamycin treatment. (n = 7 intestines per condition; n = 2-3 pictures per intestine, data points represent the average value per intestine; linear mixed model, interaction p<0.05; post-hoc test, NS p>0.05, **p<0.01). b, Masculinisation of female guts by knock-down of tra F in ECs increased the number of Lysotracker-stained puncta in the gut, and abolished the response to rapamycin treatment. (n = 7 intestines per condition; n = 2-3 pictures per intestine, data points represent the average value per intestine; linear mixed model, interaction p<0.05; post-hoc test, NS p>0.05, **p<0.01). c, Expression of histones H3 and H4 in the gut of feminised males was lower than in males, and rapamycin treatment increased it to the level in males (n = 3-4 biological replicates of 10 intestines per replicate, two-way ANOVA, H3 and H4, interaction p<0.05; post-hoc test, NS p>0.05, *p<0.05, **p<0.01). d, Expression of Bchs in the gut of feminised males was lower than in males, and rapamycin treatment increased it to the level in males (n = 4 biological replicates of 10 intestines per replicate, two-way ANOVA, H3 and H4, interaction p<0.05; post-hoc test, NS p>0.05, *p<0.05). e, Expression of histones H3 and H4 in the gut of masculinised females was higher than in females, and rapamycin treatment did not increase it further (n = 4 biological replicates of 10 intestines per replicate, two-way ANOVA, interaction p>0.05; post-hoc test, NS p>0.05, *p<0.05, **p<0.01).

Article Snippet: Following antibodies were used: Atg8a (Péter Nagy’s lab, 1:5000), Phospho-Drosophila p70 S6 Kinase (Thr398) (Cell Signaling #9209, 1:1000), total S6K (self-made, 1:1000), Histone H3 (Abcam #ab1791, 1:10000) and Histone H4 (Active motif #39269, 1:3000).

Techniques: Expressing, Staining, Knockdown

Journal: Oncotarget

Article Title: Identification of a novel gene fusion in ALT positive osteosarcoma

doi: 10.18632/oncotarget.26029

Figure Lengend Snippet: ( A ) Western blot of SJSA1 and G292 cells for DAXX, ATRX, H3.3 or α-tubulin. Chimeric DAXX-KIFC3 protein runs at a higher molecular weight in G292 cells compared to wild type DAXX in SJSA1 cells. ( B ) Western blot for DAXX in SJSA1 and G292 cells following 48 hour reverse transfection with 100 nM siRNA targeting either DAXX (left panel) or KIFC3 (right panel). α-tubulin was used as a loading control. ( C ) Western blot for DAXX, GAPDH and Histone H4 following cellular fractionation. Nuclear fractions (NUC), cytoplasmic fractions (CYTO) or whole cell extracts (WCE) of SJSA1 and G292. GAPDH and Histone H4 were used to demonstrate successful separation of cytoplasmic and nuclear fractions. ( D ) Immunoprecipitation of ATRX followed by ATRX, DAXX, and H3.3 detection by Western blot in SJSA1 and G292.

Article Snippet: ATRX (Cell Signaling, D1N2E), ATRX (Santa Cruz H-300), DAXX (Cell Signaling 25C12), FLAG-M2 (Sigma F1804), GAPDH (Santa Cruz, 0411), Histone H3.3 (Abcam, EPR17899), Histone H4 (Active Motif, 39269), PML (Santa Cruz H-238), PML (Santa Cruz PG-M3), α-Tubulin (Cell Signaling, 11H10), Purified Rabbit IgG (Bethyl Laboratories P120-101), Alexa Fluor 488 conjugated Donkey Anti-Rabbit IgG (Jackson ImmunoResearch), Cy3 conjugated Donkey anti-Rabbit (Jackson ImmunoResearch), Peroxidase conjugated Goat Anti-Mouse IgG (Jackson ImmunoResearch), Peroxidase conjugated Goat Anti-Rabbit IgG (Jackson ImmunoResearch).

Techniques: Western Blot, Molecular Weight, Transfection, Control, Cell Fractionation, Immunoprecipitation

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Histone deacetylase 8 is deregulated in urothelial cancer but not a target for efficient treatment

doi: 10.1186/s13046-014-0059-8

Figure Lengend Snippet: Western blot analysis of histone H3 and H4 acetylation in urothelial cancer cell lines after HDAC8 knockdown and HDAC8 inhibitor treatment. Amounts of acetylated and total histone H3 and H4 were analyzed by western blotting. The modifications are determined in the urothelial cancer cell lines RT-112, VM-CUB1, SW-1710, 639-V and UM-UC-3 after (A) siRNA mediated HDAC 8 knockdown (72 h) in comparison to untreated and irrelevant control and (B) pharmacological HDAC8 treatment (compound 2, compound 5, compound 6; IC 50 , 72 h). DMSO served as a solvent control.

Article Snippet: The detection of acetylated and non-acetylated histones was performed with primary antibodies against acetylated histone H3 (1:2,000, #39139, Active Motif, La Hulpe, Belgium), total histone H3 (1:1,000, #3638, Cell Signaling Technology, Inc., Danvers, MA), acetylated histone H4 (1:1,000, #39243, Active Motif, La Hulpe, Belgium) and total histone H4 (1:500, #39269, Active Motif, La Hulpe, Belgium).

Techniques: Western Blot, Knockdown, Comparison, Control, Solvent