Journal: bioRxiv

Article Title: PBRM1-VHL cooperation rewires lipid and iron metabolism to promote ferroptosis resistance in clear cell renal cell carcinoma

doi: 10.64898/2026.05.02.722429

Figure Lengend Snippet: (A) CUT&RUN genome browser tracks at the FTL locus (chr19:49,467,374–49,471,955) in Caki-2 cells. Tracks show H3K4me3, IgG, PBRM1, and PHF10 (PBAF-specific complex subunit) occupancy. Gene structure is shown below. (B) CUT&RUN genome browser tracks at the SLC40A1 locus (chr2:190,424,572–190,446,813) in Caki-2 cells. Tracks are displayed as in (A). (C) Immunoblot of ferritin heavy chain (FTH1) in Caki-2 cells expressing Empty vector, VHL, PBRM1, or PBRM1+VHL. PCNA serves as a loading control. (D) Immunoblot of transferrin receptor (TFRC) in Caki-2 cells expressing Empty vector, VHL, PBRM1, or PBRM1+VHL. PCNA serves as a loading control. (E) Representative flow cytometry histograms (YL1-A channel) and quantification of FerroOrange mean fluorescence intensity (MFI) in Caki-2 reconstituted cells treated with RSL3 (0.33 µM, 8 h). MFI is normalized to Empty vector. (F) Total cellular iron content (Fe²⁺ + Fe³⁺) in Caki-2 reconstituted cells under RSL3 treatment (0.33 µM, 8 h), quantified by colorimetric iron assay and normalized to Empty vector. (E-F) Data are presented as mean ± SD with individual replicates shown ( n = 3 biologically independent replicates). Statistical significance was assessed by one-way ANOVA with Dunnett’s multiple-comparisons test using Empty vector as the reference group. * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001; **** P ≤ 0.0001.

Article Snippet: Libraries were prepared using the CUTANATM CUT&RUN Library Prep Kit (EpiCypher, Durham, NC, USA; 14-1001 and 14-1002) and quality-checked by Qubit and Agilent TapeStation at Purdue Genomics Core Facility (Purdue University, West Lafayette, IN, USA).

Techniques: Western Blot, Expressing, Plasmid Preparation, Control, Flow Cytometry, Fluorescence, Iron Assay

Journal: bioRxiv

Article Title: PBRM1-VHL cooperation rewires lipid and iron metabolism to promote ferroptosis resistance in clear cell renal cell carcinoma

doi: 10.64898/2026.05.02.722429

Figure Lengend Snippet: (A) Box plots of expression-AUC Z-scores for the 13 ferroptosis resistance candidate genes across DepMap cancer cell lines. For each gene, the Pearson correlation between basal expression and ferroptosis inducer AUC was Z-scored across all compounds in the CTRP library. Individual points represent correlations for RSL3 (red circles), ML162 (blue triangles), ML210 (purple squares), and erastin (green diamonds). Dotted lines indicate the threshold of Z = ±0.5 (adjusted P < 0.05). Positive Z-scores indicate that higher gene expression correlates with ferroptosis resistance across cell lines. (B) Box plots of expression-AUC Z-scores for the six ferroptosis sensitizer candidate genes, formatted as in (A). Negative Z-scores indicate that lower gene expression correlates with ferroptosis resistance across cell lines. (C) Box plots of CRISPR-AUC Z-scores for the 13 resistance candidate genes. For each gene, Pearson correlations between CRISPR gene effect scores (Chronos) and GPX4 inhibitor AUC (RSL3, ML162, ML210) were Z-scored across all genes. Individual points represent correlations for each drug as in (A). Dotted lines indicate the threshold of Z = ±0.3 (adjusted P < 0.05). Negative Z-scores indicate that gene loss is disproportionately lethal in ferroptosis-resistant cell lines, reflecting functional dependency of the resistant state on those genes. (D) Box plots of CRISPR-AUC Z-scores for the six sensitizer candidate genes, formatted as in (B). Positive Z-scores indicate that gene loss correlates with increased ferroptosis sensitivity, consistent with a role in sustaining the ferroptosis-susceptible state. (E) Scatter plot of expression-AUC correlation (r, x-axis) versus CRISPR dependency-AUC correlation (r, y-axis) for candidate genes relative to canonical ferroptosis regulators. Points are colored by category: anchor genes (green), resistance candidates (red), and sensitizer candidates (blue). GPX4 and ACSL4 serve as positive and negative reference anchor points for expected resistance and sensitivity signals, respectively. (F) Cell viability of Caki-2 reconstituted cells following co-treatment with RSL3 (0.33 µM, 18 h) and the SCD inhibitor CAY10566 (SCDi; 0.5 µM). Viability is normalized to DMSO-treated controls within each genotype. data are mean ± SD ( n = 3 biologically independent replicates). (G) CUT&RUN genome browser tracks at the SCD locus (chr10:102,105,143–102,126,035) in Caki-2 cells. Tracks show H3K4me3, IgG, PBRM1, and PHF10 (PBAF-specific complex subunit) occupancy. Gene structure is shown below. (H) CUT&RUN genome browser tracks at the SREBF1 locus (chr17:17,714,907–17,714,724) in Caki-2 cells. Tracks are displayed as in (G). (I) Species-level heatmap of the top remodeled phosphatidylcholine (PC) lipid species across individual biological replicates for each genotype (Empty, PBRM1, VHL, PBRM1+VHL). Each row represents one lipid species annotated at sum-composition level. Rows are colored by unsaturation category: green indicates low-unsaturation species (≤1 double bond; resistance-associated) and red indicates high-unsaturation species (≥2 double bonds; sensitivity-associated). Color scale represents log2 abundance relative to Empty vector mean within each batch. (J) Species-level heatmap of the top remodeled phosphatidylethanolamine (PE) lipid species across individual biological replicates, formatted as in (G). (K) TAG low-to-high unsaturation ratio (double bonds ≤ 1 versus ≥ 2) across Empty vector, PBRM1, VHL, and PBRM1+VHL cells. Data are from lipidomics batch 2 ( n = 4 per genotype, except VHL, n = 3; see legend and Methods). Individual replicates are shown with mean ± SEM. Statistical significance was assessed by unpaired two-sided Welch’s t -test relative to Empty vector. * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001; **** P ≤ 0.0001.

Article Snippet: Libraries were prepared using the CUTANATM CUT&RUN Library Prep Kit (EpiCypher, Durham, NC, USA; 14-1001 and 14-1002) and quality-checked by Qubit and Agilent TapeStation at Purdue Genomics Core Facility (Purdue University, West Lafayette, IN, USA).

Techniques: Expressing, Gene Expression, CRISPR, Functional Assay, Plasmid Preparation

Journal: Precision Clinical Medicine

Article Title: Agrimol B inhibits pancreatic ductal adenocarcinoma by induction of lethal mitophagy through decreasing mitochondrial transcription termination factor 3

doi: 10.1093/pcmedi/pbag009

Figure Lengend Snippet: Agrimol B induces mitochondrial damage in PDAC cells. (A) Results of label-free quantitative proteomics after Agrimol B treatment for 24 h. (B) Western blot analysis of HADHA, TIM23, and SOD2 in PANC-1 and AsPC-1 cells. (C) Flow cytometric analysis of Fluo-4 AM accumulation in cells treated with or without 45 μmol/l Agrimol B. (D) Representative images of Fluo-4 AM accumulation in PANC-1 and AsPC-1 cells treated with or without 45 μmol/l Agrimol B for 24 h. Scale bars, 10 μm. (E) Flow cytometric analysis of mitochondrial ROS accumulation in cells treated with or without 45 μmol/l Agrimol B. (F) Representative images of mitochondrial morphology stained with Annexin V-FITC and MitoTracker Red CMXRos in PANC-1 and AsPC-1 cells treated with or without 45 μmol/l Agrimol B for 24 h. Scale bars, 10 μm. (G) Quantitative RT-PCR analysis of mtDNA copies. (H) ATP levels in PANC-1 and AsPC-1 cells treated with or without 45 μmol/l Agrimol B for 24 h. (I) Mitochondrial morphology was observed via transmission electron microscopy after treatment with or without Agrimol B for 24 h. Scale bars, 500 nm.

Article Snippet: After different transfections, PANC-1 and AsPC-1 cells were seeded in 6-well plates and maintained for 24 h. DNA was extracted according to the manufacturer’s instructions (NucleoSpin Tissue) and mtDNA was detected using the Human Mitochondrial DNA Monitoring Primer Set (Takara).

Techniques: Quantitative Proteomics, Western Blot, Staining, Quantitative RT-PCR, Transmission Assay, Electron Microscopy

Journal: Infectious Diseases of Poverty

Article Title: The ribosomal RNA synthesis ratio biomarker in Mycobacterium ulcerans for drug activity evaluation

doi: 10.1186/s40249-026-01449-2

Figure Lengend Snippet: Multiplexed Mul ddPCR. A Mul ddPCR is species-specific. Quadruple positive droplets were obtained with high concentrations of Mul cDNA, whereas quadruple negative droplets were obtained with cDNA from Mtb or Msm . B Mul cDNA samples need to be diluted to allow target quantification. Several tenfold dilutions of the Mul cDNA were required to obtain different positive and negative droplet populations, allowing for the quantification of the different targets. Mtb M. tuberculosis , Mul M. ulcerans , Msm M. smegmatis , ETS1 external transcribed spacer 1, ITS1 internal transcribed spacer 1, cDNA complementary DNA, FAM 6-Carboxyfluorescein, HEX Hexachlorofluorescein, Cy5 Cyanine-5, ROX Rhodamine X, C- negative control

Article Snippet: Bio-Rad Laboratories synthesized the ETS1, 23S, and 16S primers-probe sets, and the ITS1 primers-probe set was synthesized by CerTest (CerTest Biotec, S.L., San Mateo de Gállego, España) (Table ).

Techniques: Negative Control

Journal: Infectious Diseases of Poverty

Article Title: The ribosomal RNA synthesis ratio biomarker in Mycobacterium ulcerans for drug activity evaluation

doi: 10.1186/s40249-026-01449-2

Figure Lengend Snippet: Comparison of four diverse RS-ratio determinations in Mycobacterium ulcerans exposed to rifampicin, clarithromycin, and amoxicillin/clavulanate combinations . Comparison of the diverse RS-ratio determinations using the different possible pairs of targets ETS1/23S, ITS1/23S, ETS1/16S, and ITS1/16S in Mul samples. Values were calculated as: copies of ETS1 or ITS1 divided by copies of 23S or 16S and multiplied by 10 4 . RIF was used at 0.025 µg/ml (1/2 × MIC), CLA at 0.0625 µg/ml (1 × MIC), and AMX at 0.125 µg/ml (1 × MIC in the presence of CLV). CLV was added at a fixed 5 µg/ml concentration. RS-ratio values correspond to a single biological replicate, where data points and error bars represent the mean and standard deviation of technical replicates (2–3) using different cDNA dilutions. RIF rifampicin, CLA clarithromycin, AMX/CLV amoxicillin/clavulanate, ETS1 external transcribed spacer 1, ITS1 internal transcribed spacer 1, RS-ratio rRNA synthesis ratio, MIC Minimum inhibitory concentration

Article Snippet: Bio-Rad Laboratories synthesized the ETS1, 23S, and 16S primers-probe sets, and the ITS1 primers-probe set was synthesized by CerTest (CerTest Biotec, S.L., San Mateo de Gállego, España) (Table ).

Techniques: Comparison, Concentration Assay, Standard Deviation