Journal: Molecular Biology of the Cell

Article Title: Interplay of septin amphipathic helices in sensing membrane-curvature and filament bundling

doi: 10.1091/mbc.E20-05-0303

Figure Lengend Snippet: Genetic analyses of Shs1 and Cdc12 AH domains. (A) Viability of cdc12-6 cells expressing indicated SHS1 alleles expressed from the endogenous locus (with 3xHA tag, unless indicated with GFP tag, which lacks 3xHA) based on tetrad dissections at permissive temperature (24°C). Cells from genotypes labeled in blue appeared normal, without obvious septin defects. Genotypes in orange were sick, with partially or fully penetrant septin defects. Genotypes labeled in red were inviable. (B) DIC images of cdc12-6 shs1 mutants (see A) at 24°C. Scale bar, 5 μm. (C) Heterozygous diploids expressing the indicated Shs1 protein fused to GFP from the SHS1 locus. Scale bar, 5 μm. (D) Scatter plot quantifying cdc12-6-SpoVM septin complex adsorption onto different membrane curvatures. Black bars represent the mean. Error bars are the SD for more than 30 measured beads at each curvature across three replicates. Adsorption of cdc12-6-SpoVM complexes was significantly greater on a membrane curvature of 2 μm −1 than on curvatures of 0.67 and 0.4 μm −1 ; ** ( p < 0.01 and p < 0.0001, respectively). ns, adsorption was not significantly different. (E) Western blot comparing expression of indicated Cdc12 chimeras fused to 3xHA epitope in heterozygous diploids.

Article Snippet: A polyclonal rabbit α-HA epitope antibody preparation was used at 1:2000 dilution (Rockland Immunochemicals).

Techniques: Expressing, Labeling, Adsorption, Membrane, Western Blot

Journal: bioRxiv

Article Title: Progranulin deficiency induces lipid droplet accumulation in microglia via a STAT3-GPAT3 axis

doi: 10.64898/2026.01.17.700052

Figure Lengend Snippet: a. Volcano plot of differentially expressed genes in Grn -/- BV2 microglial cells. b. GO enrichment analysis of differentially expressed genes (DEGs) from RNA-seq in Grn -/- BV2 microglia. c. Analysis of GPAT3 mRNA levels in Grn -/- BV2 microglial cells (n=3). d. Western blot analysis of GPAT3 and statistical results of gray value quantification in WT and Grn -/- BV2 microglial cells (n=3). e. Analysis of Gpat3 mRNA levels Grn -/- mouse brains at different ages (n=3). f. Representative GPAT3 fluorescence in thalamus of Grn -/- mice at different ages and fluorescence quantification of GPAT3 (n=3). Scale bar, 20 μm. g. Representative BODIPY staining of GPAT3-overexpressing BV2 microglial cells and fluorescence quantification of BODIPY(n=3). Scale bar, 20 μm. h. Representative BODIPY staining of Grn -/- BV2 cells treated with Gpat3 siRNA or scramble siRNA negative control (NC) and fluorescence quantification of BODIPY signals (n=3). Scale bar, 20 μm. i. Representative BODIPY staining of Grn -/- BV2 cells treated with FSG67 or DMSO and fluorescence quantification of BODIPY signals (n=3). Scale bar, 20 μm. j. Western blot analysis of FLAG-tagged PGRN protein in Grn -/- BV2 microglial cells after Grn restoration. k. Analysis of Grn mRNA levels in Grn -/- BV2 microglial cells after Grn restoration (n=6). l-m. Representative GPAT3 fluorescence and BODIPY staining of Grn -/- BV2 microglial cells following PGRN restoration. Scale bar, 20 μm.

Article Snippet: Antibodies used in this experiment are: PGRN (R&D, AF2557, 1:1,000), LAMP1 (Abcam, ab208943, 1:1,000), GPAT3 (Proteintech, 20603-1-AP, 1:2,000), IBA1 (Wako, 019-19741, 1:2,000), GFAP (Cell Signaling Technology, 12389, 1:1,000).

Techniques: RNA Sequencing, Western Blot, Fluorescence, Staining, Negative Control

Journal: bioRxiv

Article Title: Progranulin deficiency induces lipid droplet accumulation in microglia via a STAT3-GPAT3 axis

doi: 10.64898/2026.01.17.700052

Figure Lengend Snippet: a. Venn diagram showing the overlapping of DEGs in four sets of transcriptomic comparison: WT vs. KO2 (Qiantang platform, 1844 DEGs), WT vs. KO2 (Novogene platform, 6770 DEGs), WT vs. KO8 (Qiantang platform, 2127 DEGs), and WT vs. KO8 (Novogene platform, 6042 DEGs). The unique and common DEGs among these comparisons are displayed. Gpat3 is found in the intersection of the four groups. b. Four paired Venn diagrams illustrating the overlap of each individual dataset from with the previously established LD signature genes. The left circle corresponds to the DEG set from , and the right circle corresponds to the list of LD-related marker genes. The overlapping sections highlight shared genes between the DEGs in Grn -/- BV2 microglial cells and the established LD-related genes.

Article Snippet: Antibodies used in this experiment are: PGRN (R&D, AF2557, 1:1,000), LAMP1 (Abcam, ab208943, 1:1,000), GPAT3 (Proteintech, 20603-1-AP, 1:2,000), IBA1 (Wako, 019-19741, 1:2,000), GFAP (Cell Signaling Technology, 12389, 1:1,000).

Techniques: Comparison, Marker

Journal: bioRxiv

Article Title: Progranulin deficiency induces lipid droplet accumulation in microglia via a STAT3-GPAT3 axis

doi: 10.64898/2026.01.17.700052

Figure Lengend Snippet: a. Analysis of Gpat3 mRNA levels in Grn -/- BV2 microglial cells and Grn -/- BV2 cells treated with Gpat3 siRNA1/2/3 (n=3). b. Western blot analysis of GPAT3 in control Grn -/- BV2 cells and Gpat3 siRNA1-treated Grn -/- BV2 cells (n=3). c. Representative PGRN fluorescence a of Grn -/- BV2 microglial cells and Grn -/- Grn OE BV2 cells. Scale bar, 20 μm. d. Analysis of Plin2 and Plin3 mRNA levels in Grn -/- BV2 microglial cells and Grn -/- Grn OE BV2 cells (n=3). e. Analysis of enzymes for triglyceride degradation mRNA levels in Grn -/- BV2 microglial cells and Grn -/- Grn OE BV2 cells (n=3).

Article Snippet: Antibodies used in this experiment are: PGRN (R&D, AF2557, 1:1,000), LAMP1 (Abcam, ab208943, 1:1,000), GPAT3 (Proteintech, 20603-1-AP, 1:2,000), IBA1 (Wako, 019-19741, 1:2,000), GFAP (Cell Signaling Technology, 12389, 1:1,000).

Techniques: Western Blot, Control, Fluorescence

Journal: bioRxiv

Article Title: Progranulin deficiency induces lipid droplet accumulation in microglia via a STAT3-GPAT3 axis

doi: 10.64898/2026.01.17.700052

Figure Lengend Snippet: a. Gene se enrichment analysis (GSEA) of IL-6 signaling in Grn -/- BV2 microglial cells. b. Analysis of IL-6 mRNA levels in Grn -/- BV2 microglial cells and Grn -restored Grn -/- BV 2 microglial cells (n=6). c. Analysis of IL-6 mRNA levels in Grn -/- mouse brains at different ages (n=3). d. Western blot analysis of p-STAT3(tyr705) and statistical results of gray value quantification. in Grn -/- BV2 microglial cells and Grn -restored Grn -/- BV2 microglial cells (n=4). e. Western blot analysis of p-STAT3(tyr705) and p-STAT3(ser727) and statistical results of gray value quantification 9-month-old Grn -/- mice (n=4). f. Western blot analysis of p-STAT3(tyr705) p-STAT3(ser727) and statistical results of gray value quantification in Grn -/- BV2 microglial cells and Grn -/- BV2 microglial cells treated with Stattic (n=3). g. Analysis of GPAT3 mRNA levels in Grn -/- BV2 microglial cells and Grn -/- BV2 microglial cells treated with Stattic (n=6). h. Western blot analysis of GPAT3 and statistical results of gray value quantification in Grn -/- BV2 microglial cells and Grn -/- BV2 microglial cells treated with Stattic (n=3). i. Representative BODIPY staining of Grn -/- BV2 microglial cells and Grn -/- BV2 microglial cells treated with Stattic and fluorescence quantification of BODIPY (n=3). Scale bar, 20 μm.

Article Snippet: Antibodies used in this experiment are: PGRN (R&D, AF2557, 1:1,000), LAMP1 (Abcam, ab208943, 1:1,000), GPAT3 (Proteintech, 20603-1-AP, 1:2,000), IBA1 (Wako, 019-19741, 1:2,000), GFAP (Cell Signaling Technology, 12389, 1:1,000).

Techniques: Western Blot, Staining, Fluorescence

Journal: bioRxiv

Article Title: Progranulin deficiency induces lipid droplet accumulation in microglia via a STAT3-GPAT3 axis

doi: 10.64898/2026.01.17.700052

Figure Lengend Snippet: a. Cell viability assay of mouse neuronal N2a cells treated with conditioned media from either BV 2 microglia cells or Grn -/- BV2 microglial cells (n = 5). b. Quantitative analysis of intracellular lactate dehydrogenase (LDH) levels in N2a cells treated as in (A) (n = 5). c. Reactive oxygen species (ROS) levels in N2a cells treated as in (A) (n = 5). d. Cell viability assay of mouse neuronal N2a cells treated with conditioned media from either Grn -/- microglial cells or Grn -rescued Grn -/- microglia (n = 5). e. Intracellular reactive oxygen species (ROS) levels in mouse neurons treated as in (D) (n = 5). f. N2a cell viability after treatment with conditioned media from either Grn -/- microglial cells or siRNA- Gpat3 treated Grn -/- microglial cells (n = 4). g. N2a cell viability after treatment with conditioned media from either Grn -/- microglial cells or FSG67-treated Grn -/- microglial cells (n = 3).

Article Snippet: Antibodies used in this experiment are: PGRN (R&D, AF2557, 1:1,000), LAMP1 (Abcam, ab208943, 1:1,000), GPAT3 (Proteintech, 20603-1-AP, 1:2,000), IBA1 (Wako, 019-19741, 1:2,000), GFAP (Cell Signaling Technology, 12389, 1:1,000).

Techniques: Viability Assay

Journal: bioRxiv

Article Title: Progranulin deficiency induces lipid droplet accumulation in microglia via a STAT3-GPAT3 axis

doi: 10.64898/2026.01.17.700052

Figure Lengend Snippet: a. Representative GPAT3 fluorescence in thalamus of Grn -/- mice injected with AAV-CTRL or AAV- Grn and fluorescence quantification of GPAT3 signals (n = 3). Scale bar, 20 μm. b. Representative LipidSpot staining in thalamus of Grn -/- mice injected with AAV-CTRL or AAV- Grn and fluorescence quantification of LipidSpot (n = 3). Scale bar, 20 μm. c. Representative IBA1 fluorescence in thalamus of Grn -/- mice injected with AAV-CTRL or AAV- Grn and fluorescence quantification of IBA1 signals (n = 3). Scale bar, 20 μm. d. Representative GFAP fluorescence in thalamus of Grn -/- mice injected with AAV-CTRL or AAV- Grn and fluorescence quantification of GFAP signals (n = 3). Scale bar, 20 μm. e. Representative BODIPY and IBA1 fluorescence staining in thalamus of Grn -/- mice treated with FSG67 or vehicle and fluorescence quantification of BODIPY in IBA + cells (n = 3). Scale bar, 20 μm. f. Recognition working memory assessed by the novel object recognition test in WT and Grn -/- mice treated with FSG67 or vehicle (n = 5 for each group). g. Anxiety behavior assessed by the light-dark transition test in WT and Grn -/- mice treated with FSG67 or vehicle (n = 5 for each group). h. Anxiety behavior assessed by the open field test in WT and Grn -/- mice treated with FSG67 or vehicle (n = 5 for each group). i. Representative open-field trajectory heatmaps for the groups in (SuperMaze software). j. Recognition working memory assessed by the novel object recognition test in Grn -/- mice injected with AAV-CTRL or AAV- Grn (n = 7 for each group). k. Anxiety behavior assessed by the light-dark transition test in Grn -/- mice injected with AAV-CTRL or AAV- Grn (n = 7 for each group). l. Anxiety behavior assessed by the open field test in Grn -/- mice injected with AAV-CTRL or AAV- Grn (n = 7 for each group). m. Representative open-field trajectory heatmaps for the groups in (AnyMaze software).

Article Snippet: Antibodies used in this experiment are: PGRN (R&D, AF2557, 1:1,000), LAMP1 (Abcam, ab208943, 1:1,000), GPAT3 (Proteintech, 20603-1-AP, 1:2,000), IBA1 (Wako, 019-19741, 1:2,000), GFAP (Cell Signaling Technology, 12389, 1:1,000).

Techniques: Fluorescence, Injection, Staining, Software

Journal: Journal for Immunotherapy of Cancer

Article Title: AGPAT3 reshapes tumor cell vulnerability to IFNγ-mediated ferroptosis and enhances immunotherapy efficacy through lipid remodeling

doi: 10.1136/jitc-2025-013305

Figure Lengend Snippet: IFN-γ alters the ether lipid metabolism via the IRF1-AGPAT3 axis. ( A ) Schematic summarizing the PUFA-ePLs biosynthesis pathway and their contribution to ferroptosis susceptibility. ( B ) Heatmap of different groups on the expression of specific genes in A375 cells and H1299 cells. ( C ) Representative western blot for AGPS, AGPAT3 and GNPAT in A375 cells and H1299 cells after IFN-γ incubation, IFN-γ: 60 mg/mL. ( D ) Cell viability of different groups treated with different concentrations of RSL3/RSL3+Fer-1 for 24 hours after IFN-γ incubation for 48 hours (n=3), IFN-γ: 100 mg/mL, Fer-1: 1 µM. ( E ) Relative ROS level of different groups treated with RSL3/RSL3+Fer-1 for 3 hours after IFN-γ incubation for 48 hours (n=3), IFN-γ: 60 mg/mL, RSL3: 500 nM, Fer-1: 1 µM. ( F ) Heatmap of different groups on the expression of IFN-γ downstream transcription factors in A375 cells and H1299 cells. ( G–H ) IRF1 binding sites at the AGPAT3 promoter region in lung cancer cell lines from GSE186168 ChIP-seq data. ( I–J ) The peak of IRF1 binding at the Agpat3 promoter region becomes higher after IFN-γ incubation from GSE201881 ( I ) and GSE141606 ( J ) ChIP-seq data. ( K ) IRF1 binding sites at the AGPAT3 promoter region in A375 from CUT&Tag data. ChIP-seq, chromatin Immunoprecipitation sequencing; CUT&Tag, Cleavage Under Targets and Tagmentation; IFN, interferon; PUFA-ePLs, polyunsaturated ether phospholipids; ROS, reactive oxygen species.

Article Snippet: The following antibodies were used: β-actin (1:5000, Proteintech, #66009-1-Ig, China), AGPS (1:1000, Abcam, #TA321589S, USA), AGPAT3 (1:1000, Proteintech, #25723-1-AP, China), GNPAT (1:1000, #ab184186, #14931-1-AP, China), IRF1 (1:1000, CST, #8478, USA), FSP1 (1:400, Proteintech, #20886-1-AP, China), and GPX4 (1:200, Abcam, #ab125066, USA).

Techniques: Expressing, Western Blot, Incubation, Binding Assay, ChIP-sequencing

Journal: Journal for Immunotherapy of Cancer

Article Title: AGPAT3 reshapes tumor cell vulnerability to IFNγ-mediated ferroptosis and enhances immunotherapy efficacy through lipid remodeling

doi: 10.1136/jitc-2025-013305

Figure Lengend Snippet: IFN-γ alters the ether lipid metabolism via the IRF1-AGPAT3 axis. ( A–B ) Changes in ratio of ether lipids and eater lipids in A375 and H1299 cells in NC, sh-AGPAT3 and sh-AGPAT3+IFN-γ group, IFN-γ: 60 mg/mL. ( C–D ) Changes in ratio of different ether lipids in A375 and H1299 cells in NC, sh-AGPAT3 and sh-AGPAT3+IFN-γ group, IFN-γ: 60 mg/mL. ( E–F ) Heatmap of ether-PC and ether-PE in A375 ( E ) and H1299 cells ( F ) in different groups, IFN-γ: 60 mg/mL. IFN, interferon; NC, normal control; PC, phosphatidylcholine; PE, phosphatidylethanolamine.

Article Snippet: The following antibodies were used: β-actin (1:5000, Proteintech, #66009-1-Ig, China), AGPS (1:1000, Abcam, #TA321589S, USA), AGPAT3 (1:1000, Proteintech, #25723-1-AP, China), GNPAT (1:1000, #ab184186, #14931-1-AP, China), IRF1 (1:1000, CST, #8478, USA), FSP1 (1:400, Proteintech, #20886-1-AP, China), and GPX4 (1:200, Abcam, #ab125066, USA).

Techniques: Control

Journal: Journal for Immunotherapy of Cancer

Article Title: AGPAT3 reshapes tumor cell vulnerability to IFNγ-mediated ferroptosis and enhances immunotherapy efficacy through lipid remodeling

doi: 10.1136/jitc-2025-013305

Figure Lengend Snippet: Tumor AGPAT3 affects the efficacy of ICI. ( A ) Schematic diagram of in vivo experiments for IFN-γ enhancing anti-PD-1 response in mouse tumor models. ( B–C ) Average tumor growth kinetics ( B ) and tumor weights ( C ) of LLC and B16 in C1-C5 groups under different treatments (n=5). ( D–E ) The proportion of CD8 + IFN-γ + cells and CD8 + TIM3 + in T cells from LLC tumor. ( F–G ) The proportion of CD8 + IFN-γ + cells ( F ) and CD8 + TIM3 + ( G ) in T cells from B16 tumor. ( H–I ) Relative infiltration of M1 macrophages and M2 macrophages and M1/M2 ratio in the TIME of each group from LLC tumor ( H ) (n=5) and B16 tumor ( I ) (n=5). FACS, fluorescenceactivated cell sorting; IFN, interferon; LLC, lewis lung carcinoma; PBS, phosphate-buffered saline; PD-1, programmed cell death protein-1; TIME, tumor immune microenvironment; .

Article Snippet: The following antibodies were used: β-actin (1:5000, Proteintech, #66009-1-Ig, China), AGPS (1:1000, Abcam, #TA321589S, USA), AGPAT3 (1:1000, Proteintech, #25723-1-AP, China), GNPAT (1:1000, #ab184186, #14931-1-AP, China), IRF1 (1:1000, CST, #8478, USA), FSP1 (1:400, Proteintech, #20886-1-AP, China), and GPX4 (1:200, Abcam, #ab125066, USA).

Techniques: In Vivo, FACS, Saline

Journal: Journal for Immunotherapy of Cancer

Article Title: AGPAT3 reshapes tumor cell vulnerability to IFNγ-mediated ferroptosis and enhances immunotherapy efficacy through lipid remodeling

doi: 10.1136/jitc-2025-013305

Figure Lengend Snippet: The clinical and immunological roles of AGPAT3 across cancer types. ( A ) Correlation between AGPAT3 expression and FD.score in TCGA pan-cancer. ( B ) Correlation between AGPAT3 expression and ferroptosis-related gene in TCGA-LUSC and TCGA-SKCM. ( C ) Correlation between AGPAT3 expression and CD8 + Teff score in HNSC, LUSC and UVM of TCGA database. ( D ) Correlation between AGPAT3 expression and IFNG in ICI cohorts. ( E ) Survival analysis of AGPAT3 expression in the Snyder2017 cohort. ( F ) Comparison of AGPAT3 expression between responder and non-responder in GSE165278 and IMvigor210. ESCA, esophageal carcinoma; FD.score, ferroptosis-driver signature score; GBM, glioblastoma; HNSC, head and neck squamous carcinoma; ICI, immune checkpoint inhibitor; IFN, interferon; KIRC, kidney renal clear cell carcinoma; KIRP, kidney renal papillary cell carcinoma; LGG, lower grade glioma; LIHC, liver hepatocellular carcinoma; LUSC, lung squamous cell carcinoma; NR, non-responders; OV, ovarian serous cystadenocarcinoma; PCPG, pheochromocytoma and paraganglioma; PRAD, prostate adenocarcinoma; R, responders; SARC, sarcoma; SKCM, skin cutaneous melanoma; STAD, stomach adenocarcinoma; TCGA, The Cancer Genome Atlas; TGCT, testicular germ cell tumors; THCA, thyroid carcinoma; UCEC, uterine corpus endometrial carcinoma; UVM, uveal melanoma.

Article Snippet: The following antibodies were used: β-actin (1:5000, Proteintech, #66009-1-Ig, China), AGPS (1:1000, Abcam, #TA321589S, USA), AGPAT3 (1:1000, Proteintech, #25723-1-AP, China), GNPAT (1:1000, #ab184186, #14931-1-AP, China), IRF1 (1:1000, CST, #8478, USA), FSP1 (1:400, Proteintech, #20886-1-AP, China), and GPX4 (1:200, Abcam, #ab125066, USA).

Techniques: Expressing, Comparison