ercc excision repair 1 ercc1  (Cell Signaling Technology Inc)

Journal: Cancer Science

Article Title: dUTPase inhibition confers susceptibility to a thymidylate synthase inhibitor in DNA‐repair‐defective human cancer cells

doi: 10.1111/cas.14718

Figure Lengend Snippet: Knockdown of BER or HR pathway genes sensitizes human cancer cells to the combination of TAS‐114 with FdUrd. A, Sensitivity profiles for TAS‐114 (3 μmol/L)‐plus‐FdUrd co‐treatment in HeLa cells treated with siRNAs against DNA damage repair genes or nonsense siRNA (negative control, NC). Drugs were added 24 h after cell seeding. Thymidine (30 μmol/L) was added 24 h after drug addition, and the cells were incubated for 48 h; except BRCA2 knockdown cells, which were incubated for 96 h. IC 50 values were calculated, and the sensitivity score for each knockdown cell line was determined relative to negative control cells. B, Viability curve of each knockdown cell line after TAS‐114 (3 μmol/L)‐plus‐FdUrd co‐treatment. Data are means ± SD ( n = 3). NC, negative control; HR, homologous recombination; MMR, mismatch repair; NER, nucleotide excision repair; BER, base excision repair; BRCA, breast cancer gene; MLH1, mutL homolog 1; ERCC1, ERCC Excision Repair 1; POLB, DNA polymerase beta; UNG, uracil‐DNA glycosylase

Article Snippet: Negative control siRNA (Negative control #1) and siRNAs against the following genes were purchased from Thermo Fisher Scientific Inc: UNG (catalog no. s14678), BRCA1 (s459), BRCA2 (s2085), MutL Homology 1 ( MLH1 , s224047), and ERCC Excision Repair 1 ( ERCC1 , s4785). siRNA against DNA polymerase β ( POLB , catalog no. D005164‐01‐0002) was purchased from Dharmacon.

Techniques: Negative Control, Incubation, Homologous Recombination

Journal: Frontiers in Oncology

Article Title: Hsa_circ_0001946 Inhibits Lung Cancer Progression and Mediates Cisplatin Sensitivity in Non-small Cell Lung Cancer via the Nucleotide Excision Repair Signaling Pathway

doi: 10.3389/fonc.2019.00508

Figure Lengend Snippet: hsa_circ_0001946 mediated cisplatin resistance via the NER signaling pathway (A) . The IC 50 values of cisplatin in A549 and A549/DDP cell lines (B) . The IC 50 value of cisplatin after transfection of A549 cells with hsa_circ_0001946 siRNA. The hsa_circ_0001946 group exhibited a higher IC 50 value of cisplatin than the negative control group (C) . Cell proliferation was evaluated by EDU assay after siRNA transfection and treatment with 15 μM cisplatin (D) . Apoptosis of cells was assessed by Hoechst assay after siRNA transfection and treatment with 15 μM cisplatin (E) . Representative flow cytometry results showing that the effects of hsa_circ_0001946 on cisplatin decreased cell apoptosis in the A549 cell line (F) . Cell viability was also evaluated by dye exclusion assay using flow cytometry after siRNA transfection and treatment with 15 μM cisplatin (G) . HCR analysis of UV damaged pCMV plasmid (H) . The damaged pCMV plasmids were then transfected into scrambled control or hsa_circ_0001946 silenced cells followed by analysis of luciferase activity. HCR results showed increased DNA damage repair in siRNA transfection group (I) . The expression of the NER pathway-associated proteins, XPA, XPC, Rad23B, RPA14, RPA32, RPA70, and ERCC1 was detected by western blotting after siRNA transfection (All data are presented as the mean ± SEM, * P < 0.05, ** P < 0.01, *** P < 0.001).

Article Snippet: After 2 h of blocking, PVDF membranes were incubated with antibodies against β-actin (Sigma-Aldrich), AKT, P-AKT, Bcl-2, Bax, caspase 3, cleaved caspase 3, p53, EGFR, replication protein A 32 (RPA32) (Wanleibio, Shenyang, China), xeroderma pigmentosum complementation group A (XPA), xeroderma pigmentosum complementation group C (XPC), RPA70, radiation sensitive 23 homolog B (Rad23B) (Proteintech, Rosemont, IL), RPA14 (Omnimabs, Alhambra, CA), RPA70 (Abcam, Cambridge, UK), and ERCC excision repair 1 (ERCC1) (Cell Signaling Technology, Boston, USA) overnight at 4°C.

Techniques: Transfection, Negative Control, EdU Assay, Flow Cytometry, Exclusion Assay, Plasmid Preparation, Control, Luciferase, Activity Assay, Expressing, Western Blot