Journal: Mutation research

Article Title: Common fragile sites in colon cancer cell lines: role of mismatch repair, RAD51 and poly(ADP-ribose) polymerase-1.

doi: 10.1016/j.mrfmmm.2011.04.006

Figure Lengend Snippet: Fig. 5. Clastogenic effects of transient silencing of the HR component RAD51 in MMR− or MMR+ cells untreated or exposed to APC. (A) Analysis of RAD51 expression. Real-time

Article Snippet: Immunofluorescence staining, using a monoclonal antibody directed against RAD51 (3C10, Santa Cruz Biotechnology, CA, USA), was previously described [14].

Techniques: Expressing

Journal: Journal of Cellular and Molecular Medicine

Article Title: MET inhibition enhances PARP inhibitor efficacy in castration‐resistant prostate cancer by suppressing the ATM/ATR and PI3K/AKT pathways

doi: 10.1111/jcmm.17037

Figure Lengend Snippet: γH2AX staining and ATM/ATR pathway downregulation following MET suppression and enhancing sensitivity to olaparib. (A) Representative immunofluorescent staining of γH2AX (red) and DAPI (blue) in DU145 and PC3 cells in the presence of Cri ((MET inhibitor crizotinib; 4 μM) and Ola (PARP inhibitor olaparib; 64 μM), Cri or Ola monotherapy, or dimethyl sulphoxide for 3 days. Scale bar, 50 μm. More than five foci per nucleus were considered as positive cells. (B) Western blot analysis of γH2AX expression in DU145 and PC3 cells. (C‐D) Western blot analysis of p‐ATM/ATM, p‐ATR/ATR, and RAD51 expression in DU145 and PC3 cells. Statistically significant differences were assessed by Student's t test in three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001. ns = no statistical difference

Article Snippet: Antibodies against GAPDH (6004–1‐Ig), BCL‐2 (60178–1‐Ig), ATM (27156–1‐AP), ATR (19787–1‐AP), IgG (H + L) (SA00013‐4), and RAD51 (14961–1‐AP) were purchased from Proteintech (Wuhan, China).

Techniques: Staining, Western Blot, Expressing

Journal: Journal of Cellular and Molecular Medicine

Article Title: MET inhibition enhances PARP inhibitor efficacy in castration‐resistant prostate cancer by suppressing the ATM/ATR and PI3K/AKT pathways

doi: 10.1111/jcmm.17037

Figure Lengend Snippet: Olaparib and crizotinib synergistically inhibit the growth of subcutaneous tumours in vivo . Once DU145 subcutaneous tumour reached 50 mm 3 , mice were injected intraperitoneally with the olaparib (Ola, 40 mg/kg) and crizotinib (Cri, 5 mg/kg), either alone or in combination for 4 weeks (5 day per week). (A) Brief experiment process diagram. (B–D) Curves of tumour volume growth, representative gross images of tumour sizes, and tumour weight after treating with different groups as indicated. (E‐F) The representative immunohistochemical analysis of the expression of Ki67, RAD51, cleaved caspase 3, γH2AX, p‐PI3K, p‐AKT, p‐ATR and p‐ATM proteins in DU145 xenografted tumour cells after treating with different groups as indicated. Statistically significant differences were assessed by one‐way analysis of variance and Student's t test. * p < 0.05, ** p < 0.01, *** p < 0.001. ns = no statistical difference

Article Snippet: Antibodies against GAPDH (6004–1‐Ig), BCL‐2 (60178–1‐Ig), ATM (27156–1‐AP), ATR (19787–1‐AP), IgG (H + L) (SA00013‐4), and RAD51 (14961–1‐AP) were purchased from Proteintech (Wuhan, China).

Techniques: In Vivo, Injection, Immunohistochemical staining, Expressing

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: XPO1 Inhibition Using Selinexor Synergizes With Chemotherapy in Acute Myeloid Leukemia (AML) by Targeting DNA Repair and Restoring Topoisomerase IIα to the Nucleus

doi: 10.1158/1078-0432.CCR-15-2885

Figure Lengend Snippet: (A) Expression levels of Chk1, MSH2, Rad51, MLH1, PMS2 and MSH6 were measured by quantitative PCR from total mRNA extracted from AML cell lines 6 hrs after selinexor treatment. The average relative expression and standard deviation of two independent experiments is shown. Selinexor treated versus untreated, * p<0.05. (B) Immunoblots of whole proteins from MOLM-13 and MV4-11 cell lines after treatment with DMSO or selinexor at the indicated doses and time points. Increased γH2A.X concurrently with increased caspase 3 cleavage are apoptosis indicators. One representative experiment of two is shown. Total mRNA (C) and protein expression (D) of Chk1 and Rad51 measured by real-time PCR and Western Blotting after DMSO or selinexor treatment in primary AML blasts. Quantification of RNA expression was done by quantitative PCR from whole RNA patient samples treated with selinexor for 10 and 24 hrs and protein expression was analyzed by immunoblots of whole protein extracts treated with selinexor for 24 and 48 hrs. Selinexor treated versus untreated, * p<0.05.

Article Snippet: Taqman Gene Assays and Antibodies All the real time PCR Taqman gene assays were purchased from Life Technologies (MSH2: Hs00953523_m1; MLH1: Hs00179866_m1; MSH6: Hs00264721_m1; PMS2: HS00241053_m1; Rad51: Hs00153418_m1; Chk1: Hs00967506_m1).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Standard Deviation, Western Blot, RNA Expression

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: XPO1 Inhibition Using Selinexor Synergizes With Chemotherapy in Acute Myeloid Leukemia (AML) by Targeting DNA Repair and Restoring Topoisomerase IIα to the Nucleus

doi: 10.1158/1078-0432.CCR-15-2885

Figure Lengend Snippet: (A) c-Myc protein expression in AML cell lines MV4-11 and MOLM-13 treated with selinexor for 24 hrs. One representative western blot of three experiments is shown. (B) Chromatin immunoprecipitation (ChIP) assays of c-Myc on the Rad51 and Chk1 promoter regions in MV4-11 cells after treatment with DMSO or selinexor for 24 hrs.

Article Snippet: Taqman Gene Assays and Antibodies All the real time PCR Taqman gene assays were purchased from Life Technologies (MSH2: Hs00953523_m1; MLH1: Hs00179866_m1; MSH6: Hs00264721_m1; PMS2: HS00241053_m1; Rad51: Hs00153418_m1; Chk1: Hs00967506_m1).

Techniques: Expressing, Western Blot, Chromatin Immunoprecipitation

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Expression of Rad51 in HIV-1 infected primary human microglial cells. (A) Quantitation of the changes in Rad51 levels was performed by densitometric analysis of the scanned X-ray films. The results were normalized relative to expression of Grb2 and are presented as a histogram. (B) p24 levels in the supernatant of the infected microglial cells during the course of HIV-1 infection as measured by ELISA and is shown over time in days post-infection.

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Expressing, Infection, Quantitation Assay, Enzyme-linked Immunosorbent Assay

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Effect of Rad51 or siRNA-Rad51 or p65 on HIV-1 LTR activity. (A) Schematic representation of the structural organization of the HIV-1 LTR enhancer region and basal/core promoter and the positions of the deletion mutants that were used in this study. (B) Primary human microglial cells were transfected with 100 ng of the HIV-1 LTR deletion mutants (−167/+66, −94/+66, −80/+66, −40/+66) fused to the luciferase reporter gene either alone or in combination with 300 ng of plasmids expressing Rad51 and/or p65 subunit of NFκB. The numbers represent the fold change in the HIV-1 promoter activity mediated by Rad51 and/or p65/NFκB. (C) The effect of silencing of p65 subunit of NFκB gene expression on HIV-1 LTR activity was assayed when by transfection the HIV-1 LTR plasmid (−167/+66) alone or in combination with 100 nM of p65 siRNA (sip65), non-targeting siRNA (ntsiRNA) or plasmid-expressing Rad51 with or without p65 siRNA. (E) Effect of silencing of Rad51 expression was also assayed by transfection of the HIV-1 LTR with 100 nM of Rad51 siRNA (siRad51), non-targeting siRNA (ntsiRNA) or plasmid-expressing p65/NFκB with or without Rad51 siRNA. The amounts of DNA in each transfection mixture were normalized with pcDNA. Luciferase activity was determined 48 hours after transfection. (D and E) The levels of expression of p65/NFκB and Rad51 were analyzed by Western blots for the lysates from the experiments shown in (C and E), respectively. Grb2 served as a loading control.

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Activity Assay, Transfection, Luciferase, Expressing, Gene Expression, Plasmid Preparation, Western Blot, Control

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Colocalization of Rad51 and p65/NFκB. Microglia were transfected with plasmid expressing p65/NFκB together with plasmids expressing either GFP-Rad51 or GFP. Cells transfected with GFP-Rad51 show bright green staining that is cytoplasmic, predominantly perinuclear and punctuate staining in the nucleus (D). Green staining in GFP-transfected control cells was uniformly distributed in nuclei and cytoplasm (A). By immunofluorescence we detected p65/NFκB protein in both nuclei and cytoplasm of infected cells (B and E rhodamine). Superimposition of both fluorochromes shows colocalization of both proteins in the perinuclear region and in the nuclei of cells (G). DAPI was used for labeling nuclear DNA (C and F).

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Transfection, Plasmid Preparation, Expressing, Staining, Control, Immunofluorescence, Infection, Labeling

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Physical interaction between Rad51 and p65/NFκB. (A) Interaction between p65/NFκB and Rad51 was evaluated by immunoprecipitation (IP)/western blot analysis. Total cell lysate prepared from U-87MG cells was immunoprecipitated using anti-Rad51 antibody (lane 2). Nonimmune mouse serum (NMS) was used for the negative control for the IP reaction (lane 1). After IP, protein complexes were washed, separated by SDS-PAGE followed by Western blot analysis using anti-p65 antibody. Fifty micrograms of the total cell lysate protein extract was included on the gel as a positive control (lane 3). (B) Protein extracts from U-87MG cells were incubated with GST-Rad51 or its deletion mutants (1 µM) in a GST pull-down assay for identification of the region of Rad51 that binds to p65/NFκB. GST was used as a negative control for the binding reaction (lane 1). NFκB/p65 was assessed by Western blot. Fifty micrograms of total cell protein extract was used as a positive control for protein expression (lane 7). (C) The integrity of GST and the GST-Rad51 fusion proteins used in the assay was demonstrated directly by SDS-PAGE followed by Coomassie blue staining. (D) Schematic representation of the organization of Rad51, the deletion mutants and their relative binding to p65/NFκB.

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Immunoprecipitation, Western Blot, Negative Control, SDS Page, Positive Control, Incubation, Pull Down Assay, Binding Assay, Expressing, Staining

Journal: Cell Cycle

Article Title: Activation of HIV-1 LTR by Rad51 in microglial cells

doi: 10.4161/cc.9.18.12930

Figure Lengend Snippet: Effect of Rad51 on the interaction of p65/NFκB with κB DNA sequence element of HIV-1 LTR measured by gel shift in nuclear extracts from U87 MG cells. (A) Approximately 100,000 cpm of synthetic [γ32P]-labeled double-stranded DNA oligonucleotide probe corresponding to the HIV-1 LTR κB site was incubated with 10 µg of nuclear extracts prepared from U87 MG cells transfected with 100 nmol siRNA for Rad51 (lane 3) or 200 nmol siRad51 (lane 4), 200 nmol non-target siRNA (lane 5), pcDNA6A-RAD51 (1–339) (lane 6), with p65/NFκB (lanes 11–14) and control pcDNA (lane 2). Labeled probe was also incubated with nuclear extracts prepared from pcDNA-transfected U87 MG cells in the presence of a specific DNA competitor (cold probe, lane 7), non-specific competitor (an unrelated dsDNA) (lanes 8 and 12), anti-p65 antibody (lanes 9 and 13) and normal mouse serum (NMS) (lanes 10 and 14). (B) Western blots were performed to determine the levels of expression of Rad51 with Lamin A as a loading control (lower part). (C) To identify the region of Rad51 involved in activation of the HIV-1 LTR, primary human microglial cells were transfected with 100 ng of LTR-luciferase plasmid (−167/+66) either alone or in combination with 300 ng of plasmids expressing Rad51: full length (1–339), Rad51 (1–160), Rad51 (1–80), Rad51 (1–40) and Rad51 (80–339). The amount of DNA in each transfection mixture was normalized with pCDNA6A. Luciferase activity was determined 48 hours after transfection. (D) Expression of Rad51 and deletion mutants was verified by western blot analysis of cell lysates prepared from the transfected primary human microglial cells using antibody against MYC-tagged Rad51.

Article Snippet: Rad51 siRNA was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA).

Techniques: Sequencing, Gel Shift, Labeling, Incubation, Transfection, Control, Western Blot, Expressing, Activation Assay, Luciferase, Plasmid Preparation, Activity Assay

Journal: Cancer Cell International

Article Title: RAD51 is a poor prognostic marker and a potential therapeutic target for oral squamous cell carcinoma

doi: 10.1186/s12935-023-03071-w

Figure Lengend Snippet: RAD51 expression is higher in oral cancer tissues and associated with poor patient prognosis. ( A ) Immunohistochemical stain of RAD51 (100x). Representative photo for RAD51 expression in oral non-cancerous tissues. ( B ) Representative photo for low RAD51 expression in oral cancer tissues. ( C ) Representative photo for high RAD51 expression in oral cancer tissues. ( D ) Quantitative result for RAD51 expression in oral non-cancerous and cancer tissues. ( E ) Progression free survival (PFS) in OSCC patients stratified by RAD51 expression and high RAD51 expression had a trend of shorter PFS. ( F ) PFS in oral squamous cell carcinoma (OSCC) patients receiving adjuvant chemotherapy stratified by RAD51 expression and high RAD51 expression had a significant shorter PFS. ( G ) PFS in OSCC patients receiving adjuvant radiotherapy stratified by RAD51 expression and high RAD51 expression had a significant shorter PFS. ( H ) Overall survival (OS) in OSCC patients stratified by RAD51 expression and high RAD51 expression had a worse OS. ( I ) OS in OSCC patients receiving adjuvant chemotherapy stratified by RAD51 expression and high RAD51 expression had a significant worse OS. ( J ) OS in OSCC patients receiving adjuvant radiotherapy stratified by RAD51 expression and high RAD51 expression had a borderline worse OS. NS, not significant, PCI, positive stained-cell index

Article Snippet: The pReceiver Lv105 lentiviral vector expressing the human RAD51 gene from GeneCopoeia (Rockville, MD, USA) was used to overexpress RAD51 in OSCC cells, and an empty pReceiver Lv105 lentiviral vector (GeneCopoeia, Rockville, MD, USA) was used for the negative control.

Techniques: Expressing, Immunohistochemical staining, Staining, Adjuvant

Journal: Cancer Cell International

Article Title: RAD51 is a poor prognostic marker and a potential therapeutic target for oral squamous cell carcinoma

doi: 10.1186/s12935-023-03071-w

Figure Lengend Snippet: RAD51 was an independent risk factor for PFS and OS in OSCC patients receiving chemotherapy

Article Snippet: The pReceiver Lv105 lentiviral vector expressing the human RAD51 gene from GeneCopoeia (Rockville, MD, USA) was used to overexpress RAD51 in OSCC cells, and an empty pReceiver Lv105 lentiviral vector (GeneCopoeia, Rockville, MD, USA) was used for the negative control.

Techniques: Expressing

Journal: Cancer Cell International

Article Title: RAD51 is a poor prognostic marker and a potential therapeutic target for oral squamous cell carcinoma

doi: 10.1186/s12935-023-03071-w

Figure Lengend Snippet: RAD51 protein was a poor prognostic factor for PFS in OSCC patients receiving adjuvant radiotherapy

Article Snippet: The pReceiver Lv105 lentiviral vector expressing the human RAD51 gene from GeneCopoeia (Rockville, MD, USA) was used to overexpress RAD51 in OSCC cells, and an empty pReceiver Lv105 lentiviral vector (GeneCopoeia, Rockville, MD, USA) was used for the negative control.

Techniques: Adjuvant, Expressing

Journal: Cancer Cell International

Article Title: RAD51 is a poor prognostic marker and a potential therapeutic target for oral squamous cell carcinoma

doi: 10.1186/s12935-023-03071-w

Figure Lengend Snippet: RAD51 overexpressing oral cancer cells survived better with less apoptotic cells after irradiation and cisplatin. ( A ) Colony formation assay in human oral cavity squamous cell carcinoma cell line, OECM1 EV (control) and OECM1 RAD51 OE (overexpression) cells after different doses of irradiation, and OECM1 RAD51 OE cells had more colonies after irradiation, ( B ) The quantification of colony formation showed that OECM1 RAD51 OE cells had better survival after different doses of irradiation than OECM1 EV cells, ( C ) XTT assay in OECM1 EV and OECM1 RAD51 OE cells at 72 h after irradiation showed that OECM1 RAD51 OE cells had higher cell viability after irradiation, ( D ) OECM1 EV and OECM1 RAD51 OE cells were stained with a combination of annexin V and PI and analyzed by FACS. Cells positive for annexin V and PI staining were counted as apoptotic cells. The bar chart describes the percentage distribution of apoptotic cells. OECM1 RAD51 OE cells have less apoptotic cells at 72 h after 10 Gy irradiation than OECM1 EV cells, ( E ) XTT assay in OECM1 EV and OECM1 RAD51 OE cells at 48 h after treatment with difference doses of cisplatin and RAD51 OE cells survived better after cisplatin treatment, ( F ) XTT assay in OECM1 EV and OECM1 RAD51 OE cells at 72 h after cisplatin treatment and RAD51 OE cells survived better after cisplatin treatment, ( G ) OECM1 EV and OECM1 RAD51 OE cells were stained with a combination of annexin V and PI and analyzed by FACS. OECM1 RAD51 OE cells have less apoptotic cells at 72 h after treatment with 5 µg/ml cisplatin Gy, gray; *, P < 0.05; **, P < 0.01; ***, P < 0.001

Article Snippet: The pReceiver Lv105 lentiviral vector expressing the human RAD51 gene from GeneCopoeia (Rockville, MD, USA) was used to overexpress RAD51 in OSCC cells, and an empty pReceiver Lv105 lentiviral vector (GeneCopoeia, Rockville, MD, USA) was used for the negative control.

Techniques: Irradiation, Colony Assay, Control, Over Expression, XTT Assay, Staining

Journal: Cancer Cell International

Article Title: RAD51 is a poor prognostic marker and a potential therapeutic target for oral squamous cell carcinoma

doi: 10.1186/s12935-023-03071-w

Figure Lengend Snippet: B02 significantly enhanced cytotoxicity of cisplatin in both OECM1 EV cells and RAD51 OE cells. ( A ) XTT assay in OECM1 EV and OECM1 RAD51 OE cells at 48 h after B02 treatment. B02 decreased cell viability both in control and RAD51 OE cells, ( B ) XTT assay in OECM1 EV and OECM1 RAD51 OE cells at 72 h after B02 treatment, ( C ) XTT assay in OECM1 EV cells at 48 h after cisplatin treatment with/without B02 and B02 enhanced cytotoxicity of cisplatin, ( D ) XTT assay in OECM1 EV cells at 72 h after cisplatin treatment with/without B02 and B02 enhanced cytotoxicity of cisplatin, ( E ) XTT assay in OECM1 RAD51 OE cells at 48 h after cisplatin treatment with/without B02 and B02 enhanced cytotoxicity of cisplatin, ( F ) XTT assay in OECM1 RAD51 OE cells at 72 h after cisplatin treatment with/without B02 and B02 enhanced cytotoxicity of cisplatin OECM1 EV, control; OECM1 RAD51 OE, overexpression; *, P < 0.05; **, P < 0.01; ***, P < 0.001

Article Snippet: The pReceiver Lv105 lentiviral vector expressing the human RAD51 gene from GeneCopoeia (Rockville, MD, USA) was used to overexpress RAD51 in OSCC cells, and an empty pReceiver Lv105 lentiviral vector (GeneCopoeia, Rockville, MD, USA) was used for the negative control.

Techniques: XTT Assay, Control, Over Expression

Journal: Cancer Cell International

Article Title: RAD51 is a poor prognostic marker and a potential therapeutic target for oral squamous cell carcinoma

doi: 10.1186/s12935-023-03071-w

Figure Lengend Snippet: More tumorspheres were formed after RAD51 overexpression, however, no difference in expressions of stemness marker. Tumorsphere formation in ( A ) OECM1 EV (control) and ( B ) OECM1 RAD51 OE (overexpression) cells; ( C ) The bar chart describes the number of spheres formed in OECM1 EV and OECM1 RAD51 OE cells. More tumorspheres were formed in OECM1 RAD51 OE cells, ( D ) CD44 expression in OECM1 EV and OECM1 RAD51 OE cells, ( E ) CD133 expression in OECM1 EV and OECM1 RAD51 OE cells, ( F ) Nanog expression in OECM1 EV and OECM1 RAD51 OE cells, ( G ) Oct-4 expression in OECM1EV and OECM1 RAD51 OE cells, ( H ) SOX2 expression in OECM1 EV and OECM1 RAD51 OE cells, ( I ) RAD51 expression in OECM1 EV and OECM1 RAD51 OE cells NS, not significant; *, P < 0.05

Article Snippet: The pReceiver Lv105 lentiviral vector expressing the human RAD51 gene from GeneCopoeia (Rockville, MD, USA) was used to overexpress RAD51 in OSCC cells, and an empty pReceiver Lv105 lentiviral vector (GeneCopoeia, Rockville, MD, USA) was used for the negative control.

Techniques: Over Expression, Marker, Control, Expressing

Journal: PLOS One

Article Title: Identification of novel antiviral host factors by functional gene expression analysis using in vitro HBV infection assay systems

doi: 10.1371/journal.pone.0314581

Figure Lengend Snippet: Primers used in this study. Primers used in custom TaqMan® Array Fast plate.

Article Snippet: 78 , RAD51 , Human , FAM , Hs00947967_m1 , Applied Biosystems.

Techniques: