anti parp2  (Proteintech)

Journal: Nucleic Acids Research

Article Title: XRCC1 mediates PARP1- and PAR-dependent recruitment of PARP2 to DNA damage sites

doi: 10.1093/nar/gkaf086

Figure Lengend Snippet: PARP1 and PAR enrich PARP2 near the breaks. ( A ) Representative images of micro-irradiation-induced GFP–PARP2 and RFP–XRCC1 foci in PARP2 KO or PARP1/2 DKO RPE-1 cells. The arrowheads indicate the micro-irradiation sites. ( B ) The relative intensity kinetics of GFP–PARP2. ( C ) The maximal relative intensity of GFP–PARP2 foci. ( D ) The time it took for the micro-irradiation-induced GFP–PARP2 foci to reach maximal intensity was plotted. Mann–Whitney test was used to calculate P -values. ( E ) The maximal relative intensity of RFP–XRCC1. (B, C, and E) The bars represent means and the standard errors of mean (SEM) from two consistent repeats with >17 cells imaged on the Ti microscope. ( F ) Representative images of micro-irradiation-induced GFP–PARP2 RA and RFP–XRCC1 foci in PARP2 KO RPE-1 cells ± 1 μM niraparib for 1 h. The arrowheads indicate the micro-irradiation sites. ( G ) The relative intensity kinetics of GFP–PARP2 RA foci. ( H and I ) The maximal relative foci intensity of (H) GFP–PARP2 RA and (I) RFP–XRCC1. (G–I) The bars represent means and SEM from one of three consistent repeats with >7 cells per repeat imaged on the Ti2 microscope. The two-tailed unpaired Student’s t -test was used to calculate the P -values; *** P < .001.

Article Snippet: Anti-PARP2 antibody (Active Motif, Cat. 39044) was used at 1:2000.

Techniques: Irradiation, MANN-WHITNEY, Microscopy, Two Tailed Test

Journal: Nucleic Acids Research

Article Title: XRCC1 mediates PARP1- and PAR-dependent recruitment of PARP2 to DNA damage sites

doi: 10.1093/nar/gkaf086

Figure Lengend Snippet: XRCC1 is required for PAR-dependent enrichment of PARP2 near the breaks. ( A ) Western blot analysis of WT, PARP1 KO, XRCC1 KO ,and PARP1/XRCC1 DKO RPE-1 cells. ( B ) Representative images of micro-irradiation-induced GFP–PARP2 foci in WT, PARP1 KO, XRCC1 KO ,and PARP1/XRCC1 DKO RPE-1 cells. The arrowhead indicates micro-irradiation sites. ( C ) The maximal relative intensity of GFP–PARP2. ( D ) The relative intensity kinetics of GFP–PARP2 foci. (C and D) The bars represent means and SEMs from two consistent repeats with >8 cells imaged each on the Ti microscope. ( E ) Western blotting shows the MMS-induced PARylation using SDS or Triton lysis buffer. ( F ) Representative images of GFP–PARP2 RA or PARP2 EA and RFP–XRCC1 in XRCC1 KO RPE-1 cells. ( G ) The maximal relative intensity of GFP–PARP2 EA foci ± RFP–XRCC1. ( H ) The relative intensity kinetics of GFP–PARP2 EA . ( I ) The maximal relative intensity of GFP-PARP2 RA ± RFP–XRCC1. ( J ) The relative intensity kinetics of GFP–PARP2 RA . The bars represent means and SEM. Panels (G) and (H) represent one of two consistent repeats, with >7 cells imaged on a Ti2 microscope, and panels (I) and (J) represent two consistent repeats, with >16 cells imaged on a Ti microscope. The bars and error bars represent means and SEM from one out of three representative experiments with n > 8 cells per experiment. The two-tailed unpaired Student’s t -test was used to calculate the P -values. ns, P > .05; * P < .05; ** P < .01; *** P < .001.

Article Snippet: Anti-PARP2 antibody (Active Motif, Cat. 39044) was used at 1:2000.

Techniques: Western Blot, Irradiation, Microscopy, Lysis, Two Tailed Test

Journal: Nucleic Acids Research

Article Title: XRCC1 mediates PARP1- and PAR-dependent recruitment of PARP2 to DNA damage sites

doi: 10.1093/nar/gkaf086

Figure Lengend Snippet: XRCC1-dependent enrichment of PARP2 requires the CAT domain of PARP2. ( A ) The schematic of PARP2 domains and truncated PARP2 with aa marked. ( B ) Representative images of micro-irradiation-induced GFP–PARP2 and RFP–XRCC1, GFP–PARP2 ΔWGR , and RFP–XRCC1, and GFP–PARP2 ΔCAT and RFP–XRCC1 foci in PARP2 KO RPE-1 cells. ( C ) The relative intensity kinetics of GFP–PARP2, GFP–PARP2 ΔWGR , and GFP–PARP2 ΔCAT . ( D ) The maximal relative intensity of GFP–PARP2, GFP–PARP2 ΔWGR , and GFP–PARP2 ΔCAT . ( E ) The maximal relative intensity of RFP–XRCC1. Panels (C–E) represent one of two consistent repeats with >7 cells imaged on a Ti microscope. ( F ) Representative images of GFP–PARP2 CAT alone and GFP–PARP2 CAT and RFP–XRCC1 in XRCC1 KO RPE-1 cells. ( G ) The relative intensity kinetics and ( H ) the maximal relative intensity of GFP–PARP2 CAT w/ and w/o RFP–XRCC1. Panels (F-H) were acquired using the Ti2 microscope. The bars represent the means and the SEM. The two-tailed unpaired Student’s t -test was used to calculate the P -values. ns, P > .05; * P < .05; *** P < .001.

Article Snippet: Anti-PARP2 antibody (Active Motif, Cat. 39044) was used at 1:2000.

Techniques: Irradiation, Microscopy, Two Tailed Test

Journal: Nucleic Acids Research

Article Title: XRCC1 mediates PARP1- and PAR-dependent recruitment of PARP2 to DNA damage sites

doi: 10.1093/nar/gkaf086

Figure Lengend Snippet: The BRCT2 domain of XRCC1 recruits PARP2. ( A ) The schematic of XRCC1 domains and their interaction partners PARP1/2, PolB, Lig3, PNKP, APTX, and APLF with aa marked for human XRCC1. The sequence information of truncated XRCC1 mutants is labeled. ( B ) Representative live cell images of micro-irradiation-induced GFP–PARP2 alone, or GFP–PARP2 with RFP–XRCC1 WT , RFP–XRCC1 BRCT1 , RFP–XRCC1 NT , RFP–XRCC1 CT , and RFP–XRCC1 CTΔLinker . ( C ) The relative intensity kinetics and ( D ) the maximal relative intensity of GFP–PARP2 foci ± various RFP–XRCC1 fragments. ( E ) The maximal relative intensity of various RFP–XRCC1 fragments. The bars and error bars represent the means and SEM from one out of two independent experiments with n > 7 cells per experiment collected at the Ti microscope. The two-tailed unpaired Student’s t -test was used to calculate the P -values. ns, P > .05; * P < .05; ** P < .01; *** P < .001.

Article Snippet: Anti-PARP2 antibody (Active Motif, Cat. 39044) was used at 1:2000.

Techniques: Sequencing, Labeling, Irradiation, Microscopy, Two Tailed Test

Journal: Nucleic Acids Research

Article Title: XRCC1 mediates PARP1- and PAR-dependent recruitment of PARP2 to DNA damage sites

doi: 10.1093/nar/gkaf086

Figure Lengend Snippet: The BRCT2 of XRCC1 binds to PARP2 in vivo and in vitro . ( A ) Co-IP of ectopically expressed GFP–PARP2 and RFP–XRCC1. CAT: PARP2 catalytic domain, including HD and ART. EA and RA both inactivate mutations of PARP2. ( B ) Co-IP of ectopically expressed GFP–PARP2 and endogenous XRCC1. ( C ) Representative western blotting and ( D ) quantification GST-pull down between PARP2 and BRCT1 or BRCT2 domain of XRCC1. ( E ) Representative gel and ( F ) Quantification of in vitro GST-pull down between PARP2 and XRCC1 BRCT2 domain. The bars represent the means and SEM. The two-tailed unpaired Student’s t -test was used to calculate the P -values. ns, P > .05; ** P < .01; *** P < .001.

Article Snippet: Anti-PARP2 antibody (Active Motif, Cat. 39044) was used at 1:2000.

Techniques: In Vivo, In Vitro, Co-Immunoprecipitation Assay, Western Blot, Two Tailed Test

Journal: Nucleic Acids Research

Article Title: XRCC1 mediates PARP1- and PAR-dependent recruitment of PARP2 to DNA damage sites

doi: 10.1093/nar/gkaf086

Figure Lengend Snippet: The XRCC1 and PARP2 facilitate PBZ domain recruitment. ( A ) Representative images of micro-irradiation-induced RFP–BRCT1 foci in WT, PARP1 KO, PARP2 KO, and XRCC1 KO RPE-1 cells. The arrowheads indicate micro-irradiation sites. ( B ) The relative intensity kinetics of RFP–XRCC1–BRCT1. ( C ) The maximal relative intensity of RFP–XRCC1–BRCT1. ( D ) Representative images of micro-irradiation-induced RFP–APLF–PBZ foci in WT, PARP1 KO, PARP2 KO, and XRCC1 KO RPE-1 cells. ( E ) The maximal relative intensity of RFP–PBZ (APLF). ( F ) The relative intensity kinetics of RFP–PBZ (APLF). The arrowheads indicate micro-irradiation sites. The bars represent the means and SEM from at least two consistent repeats with n > 16 cells imaged on the Ti2 microscope. The two-tailed unpaired Student’s t -test was used to calculate the P -values. ns, P > .05; ** P < .01; *** P < .001.

Article Snippet: Anti-PARP2 antibody (Active Motif, Cat. 39044) was used at 1:2000.

Techniques: Irradiation, Microscopy, Two Tailed Test

Journal: Nucleic Acids Research

Article Title: XRCC1 mediates PARP1- and PAR-dependent recruitment of PARP2 to DNA damage sites

doi: 10.1093/nar/gkaf086

Figure Lengend Snippet: The phosphorylation binding moiety on the XRCC1–BRCT2 domain is necessary for PARP2 enrichment. ( A ) Alphafold 3 model of the interaction between XRCC1–BRCT2 and PARP2–CAT. The key residues D575–Y576 of XRCC1 and K251 for PARP2 are shown. ( B ) Representative images of micro-irradiation-induced GFP–PARP2 foci in XRCC1 KO cells ± BRCT2 domains. ( C ) The relative intensity kinetics of PARP2. ( D ) The maximal relative intensity of GFP–PARP2 when cotransfected with various XRCC1 mutants. ( E ) Representative images of micro-irradiation-induced GFP–LIG3 foci in XRCC1 KO cells ± BRCT2 domains. ( F ) The maximal relative intensity of GFP–LIG3 when cotransfected with various XRCC1 mutants. ( G ) The working model deplicates the recruitment of PARP2–CAT by XRCC1. The two-tailed unpaired Student’s t -test was used to calculate the P -values. ns, P > .05; * P < .05; ** P < .01; *** P < .001.

Article Snippet: Anti-PARP2 antibody (Active Motif, Cat. 39044) was used at 1:2000.

Techniques: Phospho-proteomics, Binding Assay, Irradiation, Two Tailed Test