Journal: eLife

Article Title: TopBP1 biomolecular condensates as a new therapeutic target in advanced-stage colorectal cancer

doi: 10.7554/eLife.106196

Figure Lengend Snippet: ( A ) Quantification of the percentage of phosphorylated Chk2 (pChk2) (T68), and pATM (S1981)-positive cells in HCT116 colorectal cancer (CRC) cells incubated with FOLFIRI (5-FU: 6 µM and SN-38: 50 nM) and increasing concentrations of AZD2858 (from 250 to 1000 nM) for 2 hr and ( B ) of pChk1 and γH2AX-positive cells after incubation for 20 hr. Data were obtained by immunofluorescence analysis using the ‘Expression analysis’ application of the Celigo Imaging Cytometer (Nexcelom). The combination of FOLFIRI and AZD2858 is shaded in pink. The experiment was replicated three times, and each point represents a biological replicate. The statistical significance was determined by linear modeling analysis interrogating the effect of each drug separately and of their interaction (p-values given in the inset).

Article Snippet: The used antibodies were against Chk1 phosphorylated at S345 (Cell Signaling Technology, #2348), Chk1 (Santa Cruz Biotechnology, #sc8408), Chk2 phosphorylated at T68 (Cell Signaling Technology, #2661S), Chk2 (Milipore, #05-649), TopBP1 (Euromedex, #A300-111A or Santa Cruz Biotechnology, #sc-271043), ATM phosphorylated at S1981 (Rockland, #200-301-400), α-tubulin (Sigma, #T5168), RPA phosphorylated at S33 (Abcam, #ab2118877), RPA (Abcam, #AB2175), GSK-3β phosphorylated at S9 (Cell Signaling Technology, #9336), GSK-3β (Cell Signaling Technology, #9832), PARP1 (Santa Cruz Biotechnology, #sc-8007), cleaved caspase-3 (Cell Signaling Technology, #9661).

Techniques: Incubation, Immunofluorescence, Expressing, Imaging, Cytometry