Journal: bioRxiv

Article Title: Reciprocal regulation of p21 and Chk1 controls the Cyclin D1-RB pathway to mediate senescence onset after DNA damage-induced G2 arrest

doi: 10.1101/2021.08.17.452482

Figure Lengend Snippet: A. Representative immunofluorescence images (n=2) showing co-localization of γH2AX and 53BP1 foci in non-treated (NT, cell in prophase) and HDF exposed to ICRF-193 for indicated times. Right panel shows magnified area to appreciate differences in size/shape of gH2AX foci in cells exposed to ICRF-193 for 16 hr (up) and 72 hr (down). Bar, 10 μM. B. Representative immunofluorescence images (n=3) showing γH2AX foci in WT and E6- expressing HDF exposed to ICRF-193 for indicated times. Asterisk (*) denotes cell in prophase. Bar, 10 μM. C. Quantification of 53BP1 foci in WT and E6-expressing HDF exposed to ICRF-193 (IC) for indicated times. NT, non-treated cells. Pooled cells (>200) from three independent experiments. Box plot whiskers indicate 10-90 % boundary. D. Quantification of γH2AX signal intensity in the nuclei of WT and E6-expressing HDF exposed to ICRF-193 at indicated timepoints (percent of NT cells). Pooled cells (>100) from three independent experiments. Box plot whiskers indicate 10-90 % boundary. NT, non- treated cells. E. Immunofluorescence images showing co-expression and co-localization of γH2AX and 53BP1 foci in non-treated (NT) early passage HDF (EP-NT), HDF exposed to ICRF-193 (IC16h) and in several representative senescent HDF (PD 84). Arrows indicate γH2AX/53BP1 foci in senescent cells. Bar, 10 μM. F. Proposed roles for p53/p21 and CycD1-RB modules in suppressing Ckh1 and ATM/ATR signalling during G2 arrest-G2 exit switch. P values were calculated with two-sided Student’s t test; ***P ≤ 0.001, ****P ≤ 0.0001.

Article Snippet: Following primary antibodies were used: cyclin E1 (HE12, SCBT sc-247; 1:100), cyclin D1 (CST 2926; Abcam ab16663), cyclin A (Novocastra 6E6 and SCBT sc751), cyclin B1 (SCBT, sc-752; 1:100), p21 (CST 2946, 2947), H2A.X phospho-S139 (MerckMillipore clone JBW301, 05- 636), 53BP1 (Novus biologicals NB100-304); Ki-67 (Abcam ab16667 and BDTL 610968).

Techniques: Immunofluorescence, Expressing

Journal: bioRxiv

Article Title: Dose-dependent effects of histone methyltransferase NSD2 on site-specific double-strand break repair

doi: 10.1101/2023.10.18.562991

Figure Lengend Snippet: (a) Immunofluorescence staining of GM0637 cells at 30 minutes and 2 hours after UVA micro-irradiation, using anti-NSD2 and anti-53BP1 antibodies. NSD2, 53BP1, and DNA (DAPI) are shown in green, red, and blue, respectively, and merged. Arrowheads indicate the micro-irradiation sites. Scale bars: 10 μm. (b) FRAP analysis to monitor NSD2 dynamics at damage sites. EGFP-NSD2-expressing GM10637 cells were first micro-irradiated (yellow boxes), and then, photobleached (red boxes). The EGFP signal was examined before and at the indicated times after micro-irradiation. Scale bars: 5 μm.

Article Snippet: The cells were incubated anti-NSD2 antibody (Abcam, ab75359) and anti-53BP1 antibody (Novus, NB100-304) in PBS containing BSA at 37°C for 30 min.

Techniques: Immunofluorescence, Staining, Irradiation, Expressing

Journal: Molecular Cell

Article Title: Ribonucleotide Reductase Requires Subunit Switching in Hypoxia to Maintain DNA Replication

doi: 10.1016/j.molcel.2017.03.005

Figure Lengend Snippet:

Article Snippet: Rabbit polyclonal anti-53BP1 , Novus Biologicals , Cat# NB100-904.

Techniques: Transduction, Recombinant, Protease Inhibitor, SYBR Green Assay, Mutagenesis, Purification, Gel Extraction, Imaging, Sequencing, Negative Control, Real-time Polymerase Chain Reaction, Software, Expressing