parp1 protein  (MedChemExpress)

Journal: The EMBO Journal

Article Title: S-phase PARylation of microprotein RSMC enhances the function of Sororin in sister chromatid cohesion

doi: 10.1038/s44318-025-00641-8

Figure Lengend Snippet: ( A ) The interaction between Flag-PARP1 and Myc-RSMC is shown by CoIP. HEK293T cells were transfected with indicated plasmids for 48 h. Cell lysates were immunoprecipitated with anti-Flag M2 agarose followed by immunoblotting with antibodies against Flag or Myc. An asterisk indicates a nonspecific signal. ( B , C ) In situ interaction between RSMC and PARP1 detected by PLA. RSMC-3Flag cells were synchronized in early (0 h post double-thymidine release) or mid-S (5 h) phases. 10 μM olaparib was added 1 h before release. Fixed cells were subjected to PLA using anti-Flag and anti-PARP1 antibodies. Scale bar: 10 μm. ( C ) Quantification of PLA foci number. Data from three independent assays ( n > 800 cells) are shown as mean ± SEM. Statistical significance was calculated using one-way ANOVA with Tukey’s post hoc test. In G1/S, Olaparib(−) vs Olaparib(+) **** P = 2.7041e-5; In Mid S, Olaparib(−) vs Olaparib(+) **** P = 7.1195e-9; G1/S Olaparib(−) vs Mid S Olaparib(−) **** P = 7.1195e-9. ( D ) PARP1 PARylates RSMC but not RSMC-16A in vitro. In vitro PARylation assays were performed using purified GST-RSMC WT or 16 A in the presence or absence of recombinant NAD + . PARylated RSMC was detected using an anti-PAR antibody. The specificity of RSMC’s PARylation signal was validated as shown in Fig. . ( E ) RSMC is PARylated in vivo. HEK293T cells expressing GFP-tagged vector, RSMC or RSMC-16A were lysed and subjected to GFP IP, followed by immunoblotting with anti-PAN PAR and anti-GFP antibodies. ( F , G ) PARylation enhances RSMC’s interaction with Sororin. In vitro, PARylation of RSMC was conducted in the absence or presence of NAD + . After removing PARP1 and NAD + , Sororin was incubated with immobilized RSMC. Input (20%) and unbound supernatants (20%) were subjected to CBB staining. The bead-bound proteins were checked by immunoblotting (3% to detect GST-bound RSMC, and 30% each to check RSMC PARylation and its associated Sororin). SN means supernatants. ( G ) Quantification of the pulldown fraction of Sororin by PARylated RMSC (lane 3) or non-PARylated RSMC (lane 4). Mean ± SEM from three independent assays were shown. Statistical significance was calculated using Student’s t test. * P = 0.0119. ( H ) Sororin-12A is defective in cohesion. Cells expressing Flag-tagged vector, Sororin WT, or Sororin-12A were transfected with Sororin siRNA. Cohesion defects were quantified. Immunoblots of Flag, Sororin, and Tubulin validated Sororin expression. Data from three independent assays are shown as mean ± SEM. Significance determined by one-way ANOVA with Tukey’s post hoc test. siSororin(−) vs siSororin(+) **** P = 1.0334e-6, siSororin(−) vs siSororin(+)+12 A *** P = 0.0002, siSororin(−) vs siSororin(+)+Sororin P = 0.7104, siSororin(+) vs siSororin(+)+12 A *** P = 0.0002, siSororin(−) vs siSororin(+)+Sororin **** P = 1.7747e-6, siSororin(+)+12 A vs siSororin(+)+Sororin *** P = 0.0006. ( I , J ) The interaction between RSMC and Sororin increased during the S phase. ( I ) In situ interaction between RSMC and Sororin was detected by PLA. RSMC-3Flag cells were synchronized in early S (0 h post double-thymidine release) or mid-S (5 h) phases and treated with olaparib. Fixed cells were then subjected to PLA assay with anti-Flag and anti-Sororin antibodies. Scale bar: 10 μm. ( J ) PLA foci numbers were quantified from three independent assays ( n > 900 cells). Mean ± SEM is shown. Statistical significance was calculated using one-way ANOVA with Tukey’s post hoc test. In G1/S, Olaparib(−) vs Olaparib(+) **** P = 4.2095e-8; In Mid S, Olaparib(−) vs Olaparib(+) **** P = 4.2095e-8; G1/S Olaparib(−) vs Mid S Olaparib(−) **** P = 4.2095e-8. .

Article Snippet: For the in vitro PARylation assay, purified GST-RSMC or GST-RSMC-16A was bound to Glutathione Sepharose 4B beads and incubated with the PARP1 enzyme (Sino Biological, 11040-H08B) for 1 h at 37 °C in reaction buffer (50 mM Tris–HCl pH 8.0, 150 mM NaCl, 10 mM MgCl 2 , 10% glycerol, 1 mM DTT) supplemented with or without 200 μM NAD + , followed with three times wash with reaction buffer supplied with 300 mM NaCl.

Techniques: Transfection, Immunoprecipitation, Western Blot, In Situ, In Vitro, Purification, Recombinant, In Vivo, Expressing, Plasmid Preparation, Incubation, Staining

Journal: The EMBO Journal

Article Title: S-phase PARylation of microprotein RSMC enhances the function of Sororin in sister chromatid cohesion

doi: 10.1038/s44318-025-00641-8

Figure Lengend Snippet: ( A ) siPARP1 does not affect sister chromatid cohesion. Mitotic cells were collected after 48 h PARP1 RNAi and subjected to chromosome spread analyses. Sister chromatid cohesion defects were quantified from three independent assays. Immunoblots of PARP1 and GAPDH were shown below each column to confirm knockdown efficiency. Data represent mean ± SEM. The statistical significance was calculated via Student’s t test. P = 0.3984. ( B ) PARP activity is required for cohesion. PARP1 KO HeLa cells were transfected with PARP2 or control siRNA with or without 10 μM olaparib treatment. Mitotic cells were analyzed by chromosome spreads after 48 h. More than 300 mitotic cells were scored from four independent assays. Immunoblots of PARP1, PARP2 and GAPDH were shown below each column to confirm knockout and knockdown efficiency. Mean ± SEM are shown. The statistical significance was calculated via one-way ANOVA followed by Tukey’s post hoc test. P value of WT vs others (from left to right), ** P = 0.0065, * P = 0.0231, ** P = 0.0031; P value of Olaparib(+) vs others (from left to right), P = 0.8862, P = 0.9699; PARP1 KO+siPARP2 vs PARP1 KO+siPARP2+Olaparib P = 0.6589. ( C ) PARPs function redundantly with ESCO1/2 in cohesion. HeLa cells transfected with ESCO1/2 or control siRNA treated ± 10 μM olaparib. Mitotic cells were analyzed by chromosome spreads after 48 h. Over 300 mitotic cells were scored from three independent assays. Mean ± SEM are shown. The statistical analyses were performed by One-way ANOVA followed by Tukey’s post hoc test. P value of WT vs others (from left to right), **** P = 4.3539e-6, P = 0.8503, **** P = 2.3505e-6, **** P = 4.0305e-6, P = 0.3061, P = 0.4265, ****P = 5.6290e-11, *P = 0.0142, ****P = 7.6187e-7; Olaparib(+) vs Olaparib(+)+Flag-Sororin ****P = 7.2240e-5, Olaparib(+) vs Olaparib(+)+Flag-Sororin-12A P > 0.9999, Olaparib(+) vs Olaparib(+)+siE1&2 ****P = 3.0100e-6; Olaparib(+)+Flag-Sororin vs Olaparib(+)+Flag-Sororin-12A ****P = 3.6410e-5; siE1&2 vs siE1&2+Flag-Sororin **P = 0.0028, siE1&2 vs siE1&2+Flag-Sororin-12A **P = 0.0017, siE1&2 vs siE1&2+ Olaparib(+) ****P = 3.2485e-6; siE1&2+Flag-Sororin vs siE1&2+Flag-Sororin-12A P > 0.9999; siE1&2+Olaparib(+) vs siE1&2+Olaparib(+)+Flag-Sororin ****P = 6.6151e-9, siE1&2+Olaparib(+) vs siE1&2+Olaparib(+)+Flag-Sororin-12A ****P = 1.9114e-5; siE1&2+Olaparib(+)+Flag-Sororin vs siE1&2+Olaparib(+)+Flag-Sororin **P = 0.0017. ( D , E ) Quantification of Wapl-PDS5B interactions by competitive microscale thermophoresis (MST). Cy5-labeled PDS5B (34 nM constant concentration) was titrated with 2-fold serially diluted Wapl (starting at 116 nM). ( D ) Binding curves and ( E ) calculated dissociation constants (Kd) under: control (black), 17 nM Sororin-12A (orange) and 17 nM each Sororin-12A and RSMC (yellow). Data represent mean ± SEM from three independent experiments. Statistical significance was determined by one-way ANOVA followed by Tukey’s post hoc test. PDS5B-Wapl vs. PDS5B-Wapl+Sororin-12A ** P = 0.0071, PDS5B-Wapl vs. PDS5B-Wapl+Sororin-12A + RSMC ** P = 0.0026, PDS5B-Wapl+Sororin-12A vs. PDS5B-Wapl+Sororin-12A + RSMC P = 0.5637. .

Article Snippet: For the in vitro PARylation assay, purified GST-RSMC or GST-RSMC-16A was bound to Glutathione Sepharose 4B beads and incubated with the PARP1 enzyme (Sino Biological, 11040-H08B) for 1 h at 37 °C in reaction buffer (50 mM Tris–HCl pH 8.0, 150 mM NaCl, 10 mM MgCl 2 , 10% glycerol, 1 mM DTT) supplemented with or without 200 μM NAD + , followed with three times wash with reaction buffer supplied with 300 mM NaCl.

Techniques: Western Blot, Knockdown, Activity Assay, Transfection, Control, Knock-Out, Microscale Thermophoresis, Labeling, Concentration Assay, Binding Assay

Journal: The EMBO Journal

Article Title: S-phase PARylation of microprotein RSMC enhances the function of Sororin in sister chromatid cohesion

doi: 10.1038/s44318-025-00641-8

Figure Lengend Snippet: ( A ) The in vitro PARylation assay was performed using purified GST or GST-RSMC and Flag-PARP1 in the absence or presence of olaparib (20 μM) in the reaction buffer at 37 °C for 1 h. Immunoblotting was carried out with an anti-PAR monoclonal antibody. ( B ) RSMC is PARylated by PARP1 at SIMs. The in vitro PARylation assay was performed using purified GST-RSMC or GST-RSMCΔSIM and PARP1 in the reaction buffer at 37 °C for 1 h. Immunoblotting was carried out with an anti-PAR monoclonal antibody. ( C ) PARylation enhances RSMC’s binding to Sororin. In vitro PARylation of RSMC was conducted as above in the absence or presence of NAD + . After the removal of PARP1 and NAD + , Sororin was incubated with immobilized RSMC. The resins were then washed three times, resuspended with 1×loading buffer and subjected to immunoblotting analyses with the indicated antibodies. ( D ) Diagram of Sororin with three PAR-binding motifs containing 12 residues and Sororin binds to PAR chains in vitro via 3 PAR-binding motifs. PAR (20 nM) was incubated with Sororin or Sororin-12A immobilized on Ni 2+ and subjected to dot blot assays. ( E ) Sororin-12A is defective in binding RSMC in vitro. Immobilized GST-RSMC or GST (control) on glutathione beads was incubated with purified 6×His-Flag-Sororin WT or 12 A mutant. After washing, bound proteins were eluted and analyzed by SDS-PAGE followed by CBB staining. ( F ) Sororin-12A weakens its association with RSMC in cells. Cells were transfected with GFP, GFP-RSMC, Flag-Sororin WT, or Flag-Sororin-12A. Lysates were subjected to GFP-IP before immunoblotting with anti-Flag and anti-GFP antibodies. ( G ) Sororin-12A hardly binds PARylated RSMC in vitro. Immobilized RSMC was PARylated in the presence of NAD + , then PARP1 and NAD + were washed away. Afterwards, PARylated RSMC was incubated with either Sororin WT or 12 A mutant. Bound proteins were analyzed by immunoblotting using anti-Sororin, anti-GST and anti-PAR antibodies. ( H ) Sororin-12A is defective in cohesion. Flag-tagged vector, Sororin WT, or Sororin-12A was transfected into HEK293T WT or RSMC +/− cells. Cohesion defects were quantified. Data are presented as means ± SEM from three independent assays. The statistical significance was calculated via one-way ANOVA followed by Tukey’s post hoc test. P value of RSMC +/+ vs others (from left to right), **** P = 1.2454e-5, P = 0.4180, **** P = 5.0813e-6; P value of RSMC +/− vs others (from left to right), **** P = 3.8307e-5, P = 0.4946; RSMC +/− +Flag-Sororin vs RSMC +/− +Flag-Sororin-12A **** P = 1.3754e-5. .

Article Snippet: For the in vitro PARylation assay, purified GST-RSMC or GST-RSMC-16A was bound to Glutathione Sepharose 4B beads and incubated with the PARP1 enzyme (Sino Biological, 11040-H08B) for 1 h at 37 °C in reaction buffer (50 mM Tris–HCl pH 8.0, 150 mM NaCl, 10 mM MgCl 2 , 10% glycerol, 1 mM DTT) supplemented with or without 200 μM NAD + , followed with three times wash with reaction buffer supplied with 300 mM NaCl.

Techniques: In Vitro, Purification, Western Blot, Binding Assay, Incubation, Dot Blot, Control, Mutagenesis, SDS Page, Staining, Transfection, Plasmid Preparation

Journal: The EMBO Journal

Article Title: S-phase PARylation of microprotein RSMC enhances the function of Sororin in sister chromatid cohesion

doi: 10.1038/s44318-025-00641-8

Figure Lengend Snippet: ( A – D ) Inhibition of PARP1/2 (olaparib) or DNA replication (emetine, EME) impedes Sororin recruitment on chromatin. ( A ) Chromatin fractionation was performed as described in Fig. . ( B ) Quantitative data are presented as means ± SEM from five independent repeats. P value of RSMC +/+ vs others (from left to right), **** P = 3.2014e-6, **** P = 1.4982e-6, **** P = 2.3967e-5, **** P = 4.2127e-6, **** P = 5.3302e-5; RSMC +/− vs RSMC +/− (+PARPi) P = 0.9993, RSMC +/+ ( + EME) vs RSMC +/+ ( + EME&PARPi) P = 0.8598. ( C ) Immunofluorescence analysis was performed as described in Fig. . ( D ) Quantitative data were from three independent assays. One-way ANOVA with Tukey’s post hoc test was used for the comparisons. P value from left to right, from bottom to top: **** P = 3.5116e-8, **** P = 3.5116e-8, **** P = 3.5116e-8, **** P = 3.5116e-8, **** P = 3.5116e-8, P = 0.0578, p > 0.9999, P = 0.9465, P = 0.4497, P = 0.3004. ( E , F ) RSMC-16A fails to rescue cohesion defects caused by PARP inhibition. Cells transfected with Flag-RSMC-16A were treated with or without 10 μM olaparib for 48 h. Mitotic cells were collected for chromosome spreads. Sister chromatid cohesion defects were quantified in >300 mitotic cells across three independent biological replicates. Data represent mean ± SEM. Statistical significance was determined by one-way ANOVA with Tukey’s post hoc test. P value of Olaparib(−) vs others (from left to right): **** P = 4.0419e-7, P = 0.3914, **** P = 1.3348e-6; Olaparib(+) vs others (from left to right): **** P = 9.2022e-7, P = 0.1477; Olaparib(+)+Flag-RSMC vs Olaparib(+)+Flag-RSMC-16A **** P = 3.1850e-6. ( G , H ) PARP1 cooperates with ESCO1/2 to recruit Sororin. ( G ) Chromatin fractionation of HeLa cells treated with ESCO1/2 RNAi ± olaparib. Cells were harvested 5 h after release from the double-thymidine block. Immunoblots of whole-cell extracts (left) and chromatin fractions (right) are shown. ( H ) Quantification of chromatin-bound Sororin normalized to histone H3. Data represent mean ± SEM from four independent experiments. Statistical significance was determined by one-way ANOVA with Tukey’s post hoc test. siNC(−PARPi) vs siNC(+PARPi) **** P = 1.1595e-5, siNC(−PARPi) vs siE1&2(−PARPi) **** P = 6.4937e-7, siNC(−PARPi) vs siE1&2(+PARPi) **** P = 3.0904e-8, siNC(+PARPi) vs siE1&2(-PARPi) P = 0.0917, siNC(+PARPi) vs siE1&2(+PARPi) *** P = 0.003, siE1&2(−PARPi) vs siE1&2(+PARPi) * P = 0.0224. .

Article Snippet: For the in vitro PARylation assay, purified GST-RSMC or GST-RSMC-16A was bound to Glutathione Sepharose 4B beads and incubated with the PARP1 enzyme (Sino Biological, 11040-H08B) for 1 h at 37 °C in reaction buffer (50 mM Tris–HCl pH 8.0, 150 mM NaCl, 10 mM MgCl 2 , 10% glycerol, 1 mM DTT) supplemented with or without 200 μM NAD + , followed with three times wash with reaction buffer supplied with 300 mM NaCl.

Techniques: Inhibition, Fractionation, Immunofluorescence, Transfection, Blocking Assay, Western Blot

Journal: The EMBO Journal

Article Title: S-phase PARylation of microprotein RSMC enhances the function of Sororin in sister chromatid cohesion

doi: 10.1038/s44318-025-00641-8

Figure Lengend Snippet: ( A , B ) Wapl KD can fully rescue the cohesion defects caused by RSMC depletion. Cells were transfected with siRNA specifically targeted to RSMC and Wapl. Chromosome spreads analyses were performed. Bar = 10 μm. Data are presented as mean ± SEM from three independent assays. The statistical significance was calculated via one-way ANOVA with Tukey’s post hoc test. siNC vs siRSMC **** P = 9.0586e-5, siNC vs siWapl P = 0.8788, siNC vs siRSMC+siWapl P = 0.7031, siRSMC vs siRSMC+siWapl **** P = 3.8718e-5, siWapl vs siRSMC+siWapl P = 0.9836. ( C , D ) Quantification of binding affinity between fluorescently labeled PDS5B and Wapl by competitive microscale thermophoresis (MST). PDS5B was labeled with Cy5 and kept at a constant concentration at 34 nM. Wapl concentrations were serially diluted in a twofold gradient, starting from the highest concentration of 116 nM. The binding curve ( C ) and the calculated dissociation constant (Kd) ( D ) for Wapl-PDS5B interactions are shown with or without the following proteins: none (black), RSMC (17 nM, orange), Sororin (17 nM, purple), Sororin and RSMC (17 nM each, red), Sororin and RSMC-16A (17 nM each, green), Sororin and RSMCΔSIM (17 nM each, cyan). Data are presented as mean ± SEM from three independent assays. Statistical significance was evaluated using one-way ANOVA with Tukey’s post hoc test followed by Tukey’s post hoc test. P value of PDS5B-Wapl vs others (from left to right), P = 0.9984,*** P = 0.004, **** P = 8.5085e-8, *** P = 1.5291e-4, **** P = 6.1405e-5; P value of PDS5B-Wapl+RSMC vs others (from left to right), *** P = 0.006, **** P = 1.1771e-7, *** P = 0.0003, *** P = 0.0001; PDS5B-Wapl+Sororin vs PDS5B-Wapl+Sororin+RSMC **** P = 5.7977e-5; PDS5B-Wapl+Sororin+RSMC vs PDS5B-Wapl+Sororin+RSMC-16A *** P = 0.0001; PDS5B-Wapl+Sororin+RSMC vs PDS5B-Wapl+Sororin+RSMC ΔSIM *** P = 0.0003; PDS5B-Wapl+Sororin+RSMC-16A vs PDS5B-Wapl+Sororin+RSMC-ΔSIM P = 0.9804. ( E ) Quantification of binding affinity between PDS5B and Sororin by MST. MST was performed as described above. Sororin concentrations were serially diluted in a twofold gradient, starting from the highest concentration of 40 μM. Data are presented as mean ± SEM from four independent assays. ( F ) A parallel Sororin recruitment model on the fork. During the normal S phase, in front of the fork, cohesin encircles chromatin dynamically due to the anti-cohesion activity of PDS5-Wapl. On the moving fork, PARP1 is activated by nicks and flaps in replication intermediates (unligated Okazaki fragments). Then, PARP1 targets an alternative ORF-encoded microprotein named RSMC in this study. PARylation of RSMC reinforces RSMC–Sororin interaction, which contributes to the recruitment of Sororin together with the well-established ESCO1/2-catalyzed SMC3 acetylation pathway. In addition, RSMC can stimulate the anti-Wapl activity of Sororin. In brief, two fork-associated activities, PARP1-mediated RSMC PARylation and ESCO1/2-mediated SMC3 acetylation, cooperatively recruit Sororin to establish sister chromatid cohesion in human cells. ESCO1/2 and their associated fork components including MCM, PCNA and CRL4 MMS22L are omitted for brevity. .

Article Snippet: For the in vitro PARylation assay, purified GST-RSMC or GST-RSMC-16A was bound to Glutathione Sepharose 4B beads and incubated with the PARP1 enzyme (Sino Biological, 11040-H08B) for 1 h at 37 °C in reaction buffer (50 mM Tris–HCl pH 8.0, 150 mM NaCl, 10 mM MgCl 2 , 10% glycerol, 1 mM DTT) supplemented with or without 200 μM NAD + , followed with three times wash with reaction buffer supplied with 300 mM NaCl.

Techniques: Transfection, Binding Assay, Labeling, Microscale Thermophoresis, Concentration Assay, Activity Assay

Journal: Materials Today Bio

Article Title: Macrophage membrane coating enhances the therapeutic effects of Houttuynia cordata exosome-like nanovesicles against triple-negative breast cancer

doi: 10.1016/j.mtbio.2025.102604

Figure Lengend Snippet: MCELNs and CELNs elicited apoptosis in TNBC cells. (A)–(B) Flow cytometry detection of apoptosis in MDA-MB-231 cells treated with MCELNs and CELNs for 24 h. (C)–(D) Antagonism of MPA on cell apoptosis induced by MCELNs. (E)–(L) Western blot measured the expression levels of full length PARP1, cleaved PARP1, and cleaved caspase 3 following the treatment of MDA-MB-231 cells with MCELNs for 24 h ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: The main materials used in this study included P-CHK1, P-ATR, CDK4, CDC25C, P21, γ-H2AX (Abclone), cleaved PARP1, full length PARP1, cleaved caspase 3, BSA (BioFroxx), BCA Protein Concentration Assay Kit (Biyotime Shanghai), SDS-PAGE protein loading buffer (BOSTER, Wuhan), Three-color prestained protein standards (Abclone), Cell Cycle Assay Kit (RedFluorescence) (Biyotime Shanghai), enhanced RIPA lysate (BOSTER, Wuhan), RPMI 1640 medium(Gibco, USA), PKH67 (MedChemExpress, USA), PEG8000 (Chron Chemicals), 5-Fluorouracil (MedChemExpress, USA), and Camptothecin (MedChemExpress, USA).

Techniques: Flow Cytometry, Western Blot, Expressing

Journal: Materials Today Bio

Article Title: Macrophage membrane coating enhances the therapeutic effects of Houttuynia cordata exosome-like nanovesicles against triple-negative breast cancer

doi: 10.1016/j.mtbio.2025.102604

Figure Lengend Snippet: MCELNs and CELNs elicited apoptosis in TNBC cells. (A)–(B) Flow cytometry detection of apoptosis in MDA-MB-231 cells treated with MCELNs and CELNs for 24 h. (C)–(D) Antagonism of MPA on cell apoptosis induced by MCELNs. (E)–(L) Western blot measured the expression levels of full length PARP1, cleaved PARP1, and cleaved caspase 3 following the treatment of MDA-MB-231 cells with MCELNs for 24 h ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: The main materials used in this study included P-CHK1, P-ATR, CDK4, CDC25C, P21, γ-H2AX (Abclone), cleaved PARP1, full length PARP1, cleaved caspase 3, BSA (BioFroxx), BCA Protein Concentration Assay Kit (Biyotime Shanghai), SDS-PAGE protein loading buffer (BOSTER, Wuhan), Three-color prestained protein standards (Abclone), Cell Cycle Assay Kit (RedFluorescence) (Biyotime Shanghai), enhanced RIPA lysate (BOSTER, Wuhan), RPMI 1640 medium(Gibco, USA), PKH67 (MedChemExpress, USA), PEG8000 (Chron Chemicals), 5-Fluorouracil (MedChemExpress, USA), and Camptothecin (MedChemExpress, USA).

Techniques: Flow Cytometry, Western Blot, Expressing

Journal: Advanced Science

Article Title: Novel Cyano‐Artemisinin Dimer ZQJ29 Targets PARP1 to Induce Ferroptosis in Pancreatic Cancer Treatment

doi: 10.1002/advs.202501935

Figure Lengend Snippet: ZQJ29 inhibits PARP1 activity through direct binding with PARP1. A) PANC‐1 and KP4 cells were treated with varying concentrations of ZQJ29 for 24 h. B,C) Statistical analysis of Figure . D) PANC‐1 and KP4 cells were treated with 5 µM of ZQJ29 for specified periods. E,F) Statistical analysis of Figure . G) Molecular docking model illustrating the binding interaction between ZQJ29 and PARP1. H) Immunofluorescence staining of PARP1 (red) and nuclear DAPI staining (blue) in PANC‐1 and KP4 cells after 24 h treatment with ZQJ29 (0, 1, 2.5, and 5 µ m ). Scale bar: 200 µm. I) Thermal stability analysis of PARP1‐ZQJ29 interaction using CETSA across a temperature gradient (45‐70 °C). (J–K) Statistical analysis of Figure . L) PARP1 stability at 60 °C under treatment with different ZQJ29 concentrations. M,N) Statistical analysis of Figure . O) SPR assay. P) Stability of PARP1 treated with varying pronase/protein ratios. Q,R) Statistical analysis of Figure . S) PARP1 stability under different ZQJ29 concentrations (1:3000). T,U) Statistical analysis of Figure . The data was shown as mean value ± SD. * p < 0.05; ** p < 0.01; *** p < 0.001; ns, not significant.

Article Snippet: Recombinant human PARP1 protein (Sino Biological, 11040‐H08B) was immobilized on an activated carboxymethylated 5 (CM5) sensor chip using the amine coupling method.

Techniques: Activity Assay, Binding Assay, Immunofluorescence, Staining, SPR Assay

Journal: Advanced Science

Article Title: Novel Cyano‐Artemisinin Dimer ZQJ29 Targets PARP1 to Induce Ferroptosis in Pancreatic Cancer Treatment

doi: 10.1002/advs.202501935

Figure Lengend Snippet: ZQJ29‐induced ferroptosis is PARP1‐dependent. A) PPI network analysis. B) Protein expression in mouse tumor tissues. C) Statistical analysis of Figure . D) Heatmap of protein expression from proteomic data. E) Protein expression in PANC‐1 and KP4 cells treated with varying concentrations of ZQJ29. F,G) Statistical analysis of Figure . H–J) Protein expression in PANC‐1 and KP4 cells treated with ZQJ29 alone or in combination with PARP1 inhibitor (Olaparib), SLC7A11 inhibitor (Erastin), or GPX4 inhibitor (ML‐210). K–P) Statistical analysis of Figure . Q) Expression of PARP1, TP53, SLC7A11, and GPX4 in PANC‐1 and KP4 cells transfected with different PARP1 siRNAs. R,S) Statistical analysis of Figure Q. T,U) Cell viability assessed by CCK‐8 assay in PANC‐1 and KP4 cells transfected with control‐siRNA or PARP1‐siRNA, followed by treatment with ZQJ29 (1 µ m ) for 24 h. The data was shown as mean value ± SD. * p < 0.05; ** p < 0.01; *** p < 0.001; ns, not significant.

Article Snippet: Recombinant human PARP1 protein (Sino Biological, 11040‐H08B) was immobilized on an activated carboxymethylated 5 (CM5) sensor chip using the amine coupling method.

Techniques: Expressing, Transfection, CCK-8 Assay, Control

Journal: Advanced Science

Article Title: Novel Cyano‐Artemisinin Dimer ZQJ29 Targets PARP1 to Induce Ferroptosis in Pancreatic Cancer Treatment

doi: 10.1002/advs.202501935

Figure Lengend Snippet: The schematic illustration for ZQJ29 targeted PARP1 to activate ferroptosis for anti‐pancreatic cancer.

Article Snippet: Recombinant human PARP1 protein (Sino Biological, 11040‐H08B) was immobilized on an activated carboxymethylated 5 (CM5) sensor chip using the amine coupling method.

Techniques: