Journal: Cell reports

Article Title: A P53-Independent DNA Damage Response Suppresses Oncogenic Proliferation and Genome Instability

doi: 10.1016/j.celrep.2020.01.020

Figure Lengend Snippet: (A) Mre11 suppresses oncogene-induced γH2AX foci formation in both p53-proficient and p53-deficient pMMECs. Bar graphs show quantification of the percent of nuclei containing ≥5 γH2AXfoci in the different genotypes. Representative images (right) of the nuclei containing γH2AX foci are shown. White bar indicates 5 μm. (B) Bar graphs depicting the fold change in tail DNA percent for both alkaline (left) and neutral (right) COMET assays in pMMECs with the genotypes shown post-Cre-sgRNA transduction. Representative images of alkaline and neutral COMETs in R26Cas9+sgControl, R26Cas9/Myc+sgControl, and R26Cas9/Myc+sgMre11 pMMECs are shown. Data are represented as mean ± SEM. (C) Mre11 suppresses oncogene-induced R-loop formation independently of Trp53. Scatterplot shows a quantification of the nuclear S9.6foci in pMMECs from the genetic backgrounds shown after transduction with Cre-sgControl versus Cre-sgMre11. Representative images (right) of the nuclei containing S9.6 foci are shown. White bar indicates 5 μm. (D) S9.6 (R-loop) foci after RNase H1 overexpression in R26CasaRb1fl/flTrp53fl/fl pMMECs transduced with Cre-sgControl or Cre-sgMre11. (E and F) Additionally, RNase H1 overexpression counteracts the increase in (E) γH2AX and (F) p-RPA2 foci seen in Mre11 hypomorphic R26CasaRb1fl/flTrp53fl/fl pMMECs. ***p < 0.001; ****p < 0.0001. p values are calculated using a two-tailed Mann-Whitney test. See also Figure S4.

Article Snippet: Rabbit Anti-phosphorylated Histone H2AX (g-H2AX) Polyclonal Antibody (1:500 for IF) , Trevigen , Cat# 4418-APC-100.

Techniques: Transduction, Over Expression, Two Tailed Test, MANN-WHITNEY

Journal: Cell reports

Article Title: A P53-Independent DNA Damage Response Suppresses Oncogenic Proliferation and Genome Instability

doi: 10.1016/j.celrep.2020.01.020

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit Anti-phosphorylated Histone H2AX (g-H2AX) Polyclonal Antibody (1:500 for IF) , Trevigen , Cat# 4418-APC-100.

Techniques: Virus, Recombinant, Modification, Adhesive, Single Cell Gel Electrophoresis, TA Cloning, Sequencing, Generated, Software

Journal: Cancer Cell International

Article Title: Promotion of tumor development in prostate cancer by progerin

doi: 10.1186/1475-2867-10-47

Figure Lengend Snippet: Impaired DNA repairs in PC-3 cells with ectopic progerin expression . PC-3 cells were transfected with different lamin A expression constructs or vector control. The cells were treated with CPT (0.1 μM) for 12 hours and 48 hours later, the cells were stained with either γ-H2AX (A) or Phospho-Chk2 (Thr68) (B) antibody. Representative images of cells were taken with a fluorescence microscope at 400 × magnification. (C) PC-3 cells were transfected with different either progerin-pEGFP or LA-pEGFP as control. The transfected cells were treated with CPT (0.1 μM) with the indicated times. The total cell lysates were probed with either γ-H2AX or Phospho-Chk2 (Thr68) antibody. (D) Representative photomicrographs of the comet assay showing the DNA migration pattern. Representative images of cells were taken with a fluorescence microscope at 600 × magnification. Relative lengths of the DNA tail and Tail moments are presented as below. The bars represent the mean ± SE (n = 20). *P < 0.01; ** P < 0.001, compared with PBS control at the corresponding time. (E) The cells were treated with CPT (0.5 μM) for 12 hours and 48 hours, the cell viability was measured by trypan blue exclusion based cell staining.

Article Snippet: GFP antibody was purchased from Clontech; Lamin A/C (sc-20681) and emerin antibodies were purchased from Santa Cruz biotechnology; anti-phosphorylated histone-H2AX polyclonal antibody was from Trevigen.

Techniques: Expressing, Transfection, Construct, Plasmid Preparation, Control, Staining, Fluorescence, Microscopy, Single Cell Gel Electrophoresis, Migration