Journal: Genes & Diseases

Article Title: IRP1/ARID3A complex promotes pancreatic cancer chemoresistance by suppressing CYGB-related ferroptosis

doi: 10.1016/j.gendis.2025.101866

Figure Lengend Snippet: IRP1 promotes pancreatic cancer GEM-resistant in vivo and in vitro. (A) IC50 curves of GEM plus erastin/hemin in IRP1-knockdown Patu-8988 cell line. (B) IC50 curves of GEM plus erastin/hemin in IRP1-overexpressing PANC-1 cell line. (C) Patu-8988 cells stably transfected with the empty vector were subcutaneously inoculated in nude mice. Afterward, the mice were subjected to injection with GEM plus PBS, GEM plus erastin, or GEM plus hemin. (D) Tumor growth curves of the three groups of nude mice (∗∗∗ P < 0.001). (E) Patu-8988 cells stably transfected with the IRP1-overexpressing lentivirus were subcutaneously inoculated in nude mice. Afterward, the mice were subjected to injection with GEM plus PBS, GEM plus erastin, or GEM plus hemin. (F) Tumor growth curves of the three groups of nude mice (ns: P > 0.05). (G, H) The expression of IRP1 was detected in tumor tissue sections from the xenografts using immunohistochemistry. (I) The results of the intersection for co-immunoprecipitation and liquid chromatography-mass spectrometry against IRP1 antibody, gene set enrichment analysis (GSEA) databases (chromatin binding, co-regulatory transcription factors), and PRO-DIA (data-independent acquisition). GEM, gemcitabine; PBS, phosphate-buffered saline.

Article Snippet: Stable knockdown or overexpression lentiviruses were provided by Shanghai Genechem Co., Ltd., China.

Techniques: In Vivo, In Vitro, Knockdown, Stable Transfection, Transfection, Plasmid Preparation, Injection, Expressing, Immunohistochemistry, Immunoprecipitation, Liquid Chromatography, Mass Spectrometry, Binding Assay, Data-independent acquisition, Saline

Journal: Genes & Diseases

Article Title: IRP1/ARID3A complex promotes pancreatic cancer chemoresistance by suppressing CYGB-related ferroptosis

doi: 10.1016/j.gendis.2025.101866

Figure Lengend Snippet: holo-IRP1/ARID3A axis promotes pancreatic cancer cells resistant to ferroptosis and chemotherapy. (A) DCFH-DA staining (green) was used to detect ROS generation in PANC-1 cells with empty vector, overexpression plasmids targeting IRP1, or overexpression plasmids targeting IRP1 and RNAi targeting ARID3A transfected. The nuclei were stained blue with Hochest33342. (B) DCFH-DA staining (green) was used to detect ROS generation in Patu-8988 cells with empty vector, overexpression plasmids targeting IRP1, or overexpression plasmids targeting IRP1 and RNAi targeting ARID3A transfected. The nuclei were stained blue with Hochest33342. (C) The oxygen consumption rates of PANC-1 cells were showed as a bar chart (∗∗∗ P < 0.001). (D) The oxygen consumption rates of Patu-8988 cells were shown as a bar chart (∗∗∗ P < 0.001, ∗∗ P < 0.01). (E) MDA levels were detected in PANC-1 cells with empty vector, overexpression plasmids targeting IRP1, or overexpression plasmids targeting IRP1 and RNAi targeting ARID3A transfected (∗∗∗ P < 0.001, ∗∗∗∗ P < 0.0001). (F) MDA levels were detected in Patu-8988 cells with empty vector, overexpression plasmids targeting IRP1, or overexpression plasmids targeting IRP1 and RNAi targeting ARID3A transfected (∗∗∗ P < 0.001, ∗∗∗∗ P < 0.0001). (G) IC50 curves of GEM plus hemin in PANC-1 cells with empty vector, overexpression plasmids targeting IRP1, or overexpression plasmids targeting IRP1 and RNAi targeting ARID3A transfected. (H) IC50 curves of GEM plus hemin in Patu-8988 cells with empty vector, overexpression plasmids targeting IRP1, or overexpression plasmids targeting IRP1 and RNAi targeting ARID3A transfected. (I) Cellular localization of IRP1 and ARID3A in PANC-1 cells transfected with vector, IRP1 WT , or IRP1 C437S lentivirus. (J) Cellular localization of IRP1 and ARID3A in Patu-8988 cells transfected with vector, IRP1 WT , or IRP1 C437S lentivirus. MDA, malondialdehyde; GEM, gemcitabine; DCFH-DA, 2′,7′-dichlorofluorescin diacetate; ROS, reactive oxygen species.

Article Snippet: Stable knockdown or overexpression lentiviruses were provided by Shanghai Genechem Co., Ltd., China.

Techniques: Staining, Plasmid Preparation, Over Expression, Transfection

Journal: Genes & Diseases

Article Title: IRP1/ARID3A complex promotes pancreatic cancer chemoresistance by suppressing CYGB-related ferroptosis

doi: 10.1016/j.gendis.2025.101866

Figure Lengend Snippet: IRP1 C437S weakens the function of holo-IRP1 in pancreatic cancer cells. (A) IC50 curves of GEM plus hemin in PANC-1 cells transfected with vector, IRP1 WT , IRP1 C437S , IRP1 C437S plus ARID3A overexpression plasmid, ARID3A overexpression plasmid, and IRP1 WT plus ARID3A overexpression plasmid. (B) IC50 curves of GEM plus hemin in Patu-8988 cells transfected with vector, IRP1 WT , IRP1 C437S , IRP1 C437S plus ARID3A overexpression plasmid, ARID3A overexpression plasmid, and IRP1 WT plus ARID3A overexpression plasmid. (C) MDA levels of PANC-1 cells transfected with vector, IRP1 WT , IRP1 C437S , IRP1 C437S plus ARID3A overexpression plasmid, ARID3A overexpression plasmid, and IRP1 WT plus ARID3A overexpression plasmid (∗∗∗ P < 0.001). (D) MDA levels of Patu-8988 cells transfected with vector, IRP1 WT , IRP1 C437S , IRP1 C437S plus ARID3A overexpression plasmid, ARID3A overexpression plasmid, and IRP1 WT plus ARID3A overexpression plasmid (∗∗ P < 0.01, ∗∗∗ P < 0.001). (E) DCFH-DA staining (green) was used to detect ROS generation in PANC-1 cells transfected with vector, IRP1 WT , IRP1 C437S , IRP1 C437S plus ARID3A overexpression plasmid, ARID3A overexpression plasmid, and IRP1 WT plus ARID3A overexpression plasmid. (F) DCFH-DA staining (green) was used to detect ROS generation in Patu-8988 cells transfected with vector, IRP1 WT , IRP1 C437S , IRP1 C437S plus ARID3A overexpression plasmid, ARID3A overexpression plasmid, and IRP1 WT plus ARID3A overexpression plasmid. (G) The oxygen consumption rates of PANC-1 cells were shown as a bar chart (∗∗ P < 0.01, ∗∗∗ P < 0.001). (H) The oxygen consumption rates of Patu-8988 cells were shown as a bar chart (∗∗ P < 0.01, ∗∗∗ P < 0.001). MDA, malondialdehyde; GEM, gemcitabine; DCFH-DA, 2′,7′-dichlorofluorescin diacetate; ROS, reactive oxygen species.

Article Snippet: Stable knockdown or overexpression lentiviruses were provided by Shanghai Genechem Co., Ltd., China.

Techniques: Transfection, Plasmid Preparation, Over Expression, Staining

Journal: Genes & Diseases

Article Title: IRP1/ARID3A complex promotes pancreatic cancer chemoresistance by suppressing CYGB-related ferroptosis

doi: 10.1016/j.gendis.2025.101866

Figure Lengend Snippet: IRP1 and ARID3A reduce the chromatin accessibility of the CYGB promoter region and inhibit CYGB expression. (A) The intersection of down-regulated peaks in ATAC sequencing, up-regulated genes in RNA sequencing, and the Ferroptosis driver database. (B) The binding peak of ARID3A and chromatin accessibility regulated by IRP1 in the promoter region of CYGB were visualized by an integrative genomics viewer (IGV). (C) CYGB mRNA expression level in ARID3A-knockdown Patu-8988 cells was detected by quantitative reverse transcription PCR (∗∗ P < 0.01, ∗∗∗ P < 0.001). (D) CYGB mRNA expression level in ARID3A-overexpressing PANC-1 cells was detected by quantitative reverse transcription PCR (∗ P < 0.05, ∗∗∗ P < 0.001). (E) ARID3A, IRP1, and CYGB protein expression levels were detected in PANC-1 and Patu-8898 cells by Western blotting to verify the relationship between ARID3A, IRP1, and CYGB under the condition of iron overload. (F) CYGB expression level was detected in Patu-8898 cells by Western blotting to verify the relationship between ARID3A, IRP1, and CYGB under the condition of iron overload (∗∗∗ P < 0.001). (G) CYGB expression level was detected in PANC-1 cells by Western blotting to verify the relationship between ARID3A, IRP1, and CYGB under the condition of iron overload (∗ P < 0.05, ∗∗∗ P < 0.001). (H, I) Chromatin immunoprecipitation assay performed with ARID3A antibody followed by detection of CYGB promoter through quantitative reverse transcription PCR in Patu-8988 and PANC-1 cells (normal condition, erastin added, hemin added; ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001). (J) DCFH-DA staining (green) was used to detect ROS generation in Patu-8988 cells transfected with control, siRNAs targeting CYGB. (K) The oxygen consumption rates of Patu-8988 cells were shown as a bar chart (∗∗∗ P < 0.001). (L) DCFH-DA staining (green) was used to detect ROS generation in PANC-1 cells transfected with the vector, overexpression plasmid targeting CYGB. (M) The oxygen consumption rates of PANC-1 cells were shown as a bar chart (∗∗∗ P < 0.001). (N) MDA levels were detected in Patu-8988 cells transfected with control, siRNAs targeting CYGB (∗∗∗ P < 0.001). (O) MDA levels were detected in PANC-1 cells transfected with the vector, overexpression plasmid targeting CYGB (∗∗∗ P < 0.001). MDA, malondialdehyde; DCFH-DA, 2′,7′-dichlorofluorescin diacetate; ROS, reactive oxygen species.

Article Snippet: Stable knockdown or overexpression lentiviruses were provided by Shanghai Genechem Co., Ltd., China.

Techniques: Expressing, Sequencing, RNA Sequencing, Binding Assay, Knockdown, Reverse Transcription, Western Blot, Chromatin Immunoprecipitation, Staining, Transfection, Control, Plasmid Preparation, Over Expression