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$a , Schematic of chemically induced strategy for recruitment of the BAF complex (via SMARCA2/4) to <t>BCL6.</t> The BAF complex at BCL6 loci leads to eviction of nucleosomes and BCL6, increased chromatin accessibility, and de-repression of pro-apoptotic BCL6 target genes. b , Overview of chemoproteomic competition assay for SMARCA ligand characterization. SMARCA ligand with linker was immobilized on NHS-activated Sepharose beads and outcompeted with the SMARCA ligand (top). Each data point in the volcano plot (bottom) represents a dose-response score for a protein. The p-value indicates the significance of a dose-response according to the CurveCurator statistics pipeline, and the log2 fold change indicates the effect size of the dose-dependent depletion. Binding curves for highlighted BAF complex members in orange are shown in Extended Data Figure 1a. Protein with missing/imputed values in their dose-response curves were filtered out. Hits: Log 2 FC ≥ |1| and -log 10 P value ≥ 2 (dotted lines). c , Chemical structures of BCL6-SMARCA bifunctional compounds, TRIP1, TRIP1-neg1 (SMARCA non-binding), and TRIP1-neg2 (BCL6 non-binding). d , CellTiter-Glo cell viability assays in diffuse large B cell lymphoma (DLBCL) cell lines. KARPAS-422 (top) and SU-DHL4 (bottom) cells were treated with a dilution series of TRIP1 or negative controls for 72 hours. Viability with compound treatment is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. e , BCL6 transcriptional reporter de-repression. KARPAS-422 cells expressing a BCL6 transcriptional reporter (top) were treated with a dilution series of TRIP1 or negative controls for 24 hours. Reporter activity is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. f , CaspaseGlo 3/7 apoptosis detection. KARPAS-422 cells were treated with a dilution series of compounds for 16 hours. Caspase 3/7 activity is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. g , SMARCA4 target engagement. Monoclonal HEK293T cells expressing endogenously tagged SMARCA4-HiBiT were pre-treated with a dilution series of TRIP1 or the SMARCA ligand for 2 hours, followed by co-treatment with 0.1 µM ACBI1 for 8 hours. SMARCA4-HiBiT levels are shown as a remaining fraction relative to DMSO vehicle control (no ACBI1 treatment). D max represents the fraction of SMARCA4 remaining when ACBI1 is co-treated with DMSO. Gray bar indicates early cytotoxic/pro-apoptotic concentrations of TRIP1. Data represent mean ± SD, n = 6 independent replicates.$
Bcl6, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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bcl6 - by Bioz Stars, 2026-07

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1) Product Images from "Leveraging the BAF chromatin remodeling complex for targeted transcriptional rewiring in cancer"

Article Title: Leveraging the BAF chromatin remodeling complex for targeted transcriptional rewiring in cancer

Journal: bioRxiv

doi: 10.64898/2026.03.30.715217

$a , Schematic of chemically induced strategy for recruitment of the BAF complex (via SMARCA2/4) to BCL6. The BAF complex at BCL6 loci leads to eviction of nucleosomes and BCL6, increased chromatin accessibility, and de-repression of pro-apoptotic BCL6 target genes. b , Overview of chemoproteomic competition assay for SMARCA ligand characterization. SMARCA ligand with linker was immobilized on NHS-activated Sepharose beads and outcompeted with the SMARCA ligand (top). Each data point in the volcano plot (bottom) represents a dose-response score for a protein. The p-value indicates the significance of a dose-response according to the CurveCurator statistics pipeline, and the log2 fold change indicates the effect size of the dose-dependent depletion. Binding curves for highlighted BAF complex members in orange are shown in Extended Data Figure 1a. Protein with missing/imputed values in their dose-response curves were filtered out. Hits: Log 2 FC ≥ |1| and -log 10 P value ≥ 2 (dotted lines). c , Chemical structures of BCL6-SMARCA bifunctional compounds, TRIP1, TRIP1-neg1 (SMARCA non-binding), and TRIP1-neg2 (BCL6 non-binding). d , CellTiter-Glo cell viability assays in diffuse large B cell lymphoma (DLBCL) cell lines. KARPAS-422 (top) and SU-DHL4 (bottom) cells were treated with a dilution series of TRIP1 or negative controls for 72 hours. Viability with compound treatment is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. e , BCL6 transcriptional reporter de-repression. KARPAS-422 cells expressing a BCL6 transcriptional reporter (top) were treated with a dilution series of TRIP1 or negative controls for 24 hours. Reporter activity is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. f , CaspaseGlo 3/7 apoptosis detection. KARPAS-422 cells were treated with a dilution series of compounds for 16 hours. Caspase 3/7 activity is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. g , SMARCA4 target engagement. Monoclonal HEK293T cells expressing endogenously tagged SMARCA4-HiBiT were pre-treated with a dilution series of TRIP1 or the SMARCA ligand for 2 hours, followed by co-treatment with 0.1 µM ACBI1 for 8 hours. SMARCA4-HiBiT levels are shown as a remaining fraction relative to DMSO vehicle control (no ACBI1 treatment). D max represents the fraction of SMARCA4 remaining when ACBI1 is co-treated with DMSO. Gray bar indicates early cytotoxic/pro-apoptotic concentrations of TRIP1. Data represent mean ± SD, n = 6 independent replicates.$
Figure Legend Snippet: a , Schematic of chemically induced strategy for recruitment of the BAF complex (via SMARCA2/4) to BCL6. The BAF complex at BCL6 loci leads to eviction of nucleosomes and BCL6, increased chromatin accessibility, and de-repression of pro-apoptotic BCL6 target genes. b , Overview of chemoproteomic competition assay for SMARCA ligand characterization. SMARCA ligand with linker was immobilized on NHS-activated Sepharose beads and outcompeted with the SMARCA ligand (top). Each data point in the volcano plot (bottom) represents a dose-response score for a protein. The p-value indicates the significance of a dose-response according to the CurveCurator statistics pipeline, and the log2 fold change indicates the effect size of the dose-dependent depletion. Binding curves for highlighted BAF complex members in orange are shown in Extended Data Figure 1a. Protein with missing/imputed values in their dose-response curves were filtered out. Hits: Log 2 FC ≥ |1| and -log 10 P value ≥ 2 (dotted lines). c , Chemical structures of BCL6-SMARCA bifunctional compounds, TRIP1, TRIP1-neg1 (SMARCA non-binding), and TRIP1-neg2 (BCL6 non-binding). d , CellTiter-Glo cell viability assays in diffuse large B cell lymphoma (DLBCL) cell lines. KARPAS-422 (top) and SU-DHL4 (bottom) cells were treated with a dilution series of TRIP1 or negative controls for 72 hours. Viability with compound treatment is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. e , BCL6 transcriptional reporter de-repression. KARPAS-422 cells expressing a BCL6 transcriptional reporter (top) were treated with a dilution series of TRIP1 or negative controls for 24 hours. Reporter activity is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. f , CaspaseGlo 3/7 apoptosis detection. KARPAS-422 cells were treated with a dilution series of compounds for 16 hours. Caspase 3/7 activity is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. g , SMARCA4 target engagement. Monoclonal HEK293T cells expressing endogenously tagged SMARCA4-HiBiT were pre-treated with a dilution series of TRIP1 or the SMARCA ligand for 2 hours, followed by co-treatment with 0.1 µM ACBI1 for 8 hours. SMARCA4-HiBiT levels are shown as a remaining fraction relative to DMSO vehicle control (no ACBI1 treatment). D max represents the fraction of SMARCA4 remaining when ACBI1 is co-treated with DMSO. Gray bar indicates early cytotoxic/pro-apoptotic concentrations of TRIP1. Data represent mean ± SD, n = 6 independent replicates.

Techniques Used: Competitive Binding Assay, Binding Assay, Control, Expressing, Activity Assay, Drug discovery

a , In vitro TR-FRET ternary complex formation assay. The interaction between biotinylated SMARCA4 bromodomain (100 nM) and FITC-labeled BCL6 BTB domain (500 nM) was measured with increasing concentrations of TRIP1. The biotinylated SMARCA4 bromodomain was captured with streptavidin-terbium cryptate donor (SA-Tb, 2 nM), and TR-FRET signal was monitored upon complex formation. The resulting TR-FRET ratio was normalized to the DMSO vehicle control; data represent mean ± SD, n = 2 independent replicates. b , TR-FRET competition assay. The interaction between biotinylated SMARCA2/4 bromodomains (100 nM) and FITC-labeled BCL6 BTB domain (500 nM) was measured in the presence of 100 nM TRIP1 and increasing concentrations of BCL6 or SMARCA ligand. The resulting TR-FRET ratio was background subtracted and normalized to the DMSO vehicle control; data represent mean ± SD, n = 2 independent replicates. c , SplitHalo assay for BCL6-SMARCA2 interaction. Schematic of the in-cell splitHalo assay to probe induced protein-protein interactions (top). A HaloTag enzyme is split into two complementing parts, cpHalo and Hpep3 peptide, which only assemble into a functional self-labeling HaloTag enzyme when actively brought into proximity and supplied with TAMRA dye. HEK293T cells co-expressing BCL6-cpHalo and SMARCA2(BD)-Hpep3 were treated with a dilution series of TRIP1 or negative controls and incubated with compound and the covalent HaloTag dye TAMRA for 3 hours (bottom). Data is normalized to DMSO vehicle control, which corresponds to the baseline signal upon TAMRA addition; data represent mean ± SD, n = 3 independent replicates. d , SplitHalo competition assay for TRIP1-induced BCL6-SMARCA2 interaction. Schematic of the in-cell splitHalo competition assay to probe the inhibition of induced protein-protein interactions (top). Cells are pre-incubated with excess amounts of protein ligands to saturate binding pockets and prevent or reduce ternary complex formation. HEK293T cells co-expressing BCL6-cpHalo and SMARCA2(BD)-Hpep3 were pre-treated with a dilution series of SMARCA or BCL6 ligand for 30 minutes before adding 1 µM TRIP1 and TAMRA dye for 3 hours (bottom). Data is normalized to TRIP1 with DMSO vehicle control without ligand addition, corresponding to maximum ternary complex formation; data represent mean ± SD, n = 3 independent replicates. e , f , BCL6 transcriptional reporter competition. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO, the BCL6 ligand ( e ), or SMARCA ligand ( f ) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates. g , CaspaseGlo 3/7 apoptosis pre-degradation. KARPAS-422 cells were pre-treated with DMSO or SMARCA degrader (ACBI1) for 8 hours, followed by co-treatment with 1 µM of TRIP1 for 16 hours. Caspase 3/7 activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Figure Legend Snippet: a , In vitro TR-FRET ternary complex formation assay. The interaction between biotinylated SMARCA4 bromodomain (100 nM) and FITC-labeled BCL6 BTB domain (500 nM) was measured with increasing concentrations of TRIP1. The biotinylated SMARCA4 bromodomain was captured with streptavidin-terbium cryptate donor (SA-Tb, 2 nM), and TR-FRET signal was monitored upon complex formation. The resulting TR-FRET ratio was normalized to the DMSO vehicle control; data represent mean ± SD, n = 2 independent replicates. b , TR-FRET competition assay. The interaction between biotinylated SMARCA2/4 bromodomains (100 nM) and FITC-labeled BCL6 BTB domain (500 nM) was measured in the presence of 100 nM TRIP1 and increasing concentrations of BCL6 or SMARCA ligand. The resulting TR-FRET ratio was background subtracted and normalized to the DMSO vehicle control; data represent mean ± SD, n = 2 independent replicates. c , SplitHalo assay for BCL6-SMARCA2 interaction. Schematic of the in-cell splitHalo assay to probe induced protein-protein interactions (top). A HaloTag enzyme is split into two complementing parts, cpHalo and Hpep3 peptide, which only assemble into a functional self-labeling HaloTag enzyme when actively brought into proximity and supplied with TAMRA dye. HEK293T cells co-expressing BCL6-cpHalo and SMARCA2(BD)-Hpep3 were treated with a dilution series of TRIP1 or negative controls and incubated with compound and the covalent HaloTag dye TAMRA for 3 hours (bottom). Data is normalized to DMSO vehicle control, which corresponds to the baseline signal upon TAMRA addition; data represent mean ± SD, n = 3 independent replicates. d , SplitHalo competition assay for TRIP1-induced BCL6-SMARCA2 interaction. Schematic of the in-cell splitHalo competition assay to probe the inhibition of induced protein-protein interactions (top). Cells are pre-incubated with excess amounts of protein ligands to saturate binding pockets and prevent or reduce ternary complex formation. HEK293T cells co-expressing BCL6-cpHalo and SMARCA2(BD)-Hpep3 were pre-treated with a dilution series of SMARCA or BCL6 ligand for 30 minutes before adding 1 µM TRIP1 and TAMRA dye for 3 hours (bottom). Data is normalized to TRIP1 with DMSO vehicle control without ligand addition, corresponding to maximum ternary complex formation; data represent mean ± SD, n = 3 independent replicates. e , f , BCL6 transcriptional reporter competition. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO, the BCL6 ligand ( e ), or SMARCA ligand ( f ) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates. g , CaspaseGlo 3/7 apoptosis pre-degradation. KARPAS-422 cells were pre-treated with DMSO or SMARCA degrader (ACBI1) for 8 hours, followed by co-treatment with 1 µM of TRIP1 for 16 hours. Caspase 3/7 activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Techniques Used: In Vitro, Tube Formation Assay, Labeling, Control, Competitive Binding Assay, Protein-Protein interactions, Functional Assay, Expressing, Incubation, Inhibition, Binding Assay, Activity Assay

a , TRIP1 resistance CRISPR viability screen. KARPAS-422 iCas9 cells were mutagenized with a genome-wide sgRNA library and cultivated in the presence of DMSO or TRIP1 (400 nM) for 33 days. Gene-level enrichment score was calculated as -log10 (P value) × log2 (fold-change). BAF complex members are colored based on involvement in different sub-complexes or modules. n = 2 independent replicates. b , Competitive growth assay in KARPAS-422 cells. Control KARPAS-422 iCas9 cells expressing AAVS1-targeting sgRNA and BFP were mixed with KARPAS-422 iCas9 cells expressing BAF subunit-targeting sgRNAs and mCherry. Cell mixtures were treated with DMSO, TRIP1 (400 nM), BRD4 recruiting TCIP1 (4 nM), or p300/CBP recruiting TCIP3 (4 nM) and evaluated in regular intervals every 3 or 4 days via flow cytometry. Data were logit-transformed and normalized to 50% at day 0 to enable comparison across conditions. Data represent mean ± SD, n = 3 independent replicates. c , BCL6 transcriptional reporter pre-degradation. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO or BRD7/9 degrader (VZ-185) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Figure Legend Snippet: a , TRIP1 resistance CRISPR viability screen. KARPAS-422 iCas9 cells were mutagenized with a genome-wide sgRNA library and cultivated in the presence of DMSO or TRIP1 (400 nM) for 33 days. Gene-level enrichment score was calculated as -log10 (P value) × log2 (fold-change). BAF complex members are colored based on involvement in different sub-complexes or modules. n = 2 independent replicates. b , Competitive growth assay in KARPAS-422 cells. Control KARPAS-422 iCas9 cells expressing AAVS1-targeting sgRNA and BFP were mixed with KARPAS-422 iCas9 cells expressing BAF subunit-targeting sgRNAs and mCherry. Cell mixtures were treated with DMSO, TRIP1 (400 nM), BRD4 recruiting TCIP1 (4 nM), or p300/CBP recruiting TCIP3 (4 nM) and evaluated in regular intervals every 3 or 4 days via flow cytometry. Data were logit-transformed and normalized to 50% at day 0 to enable comparison across conditions. Data represent mean ± SD, n = 3 independent replicates. c , BCL6 transcriptional reporter pre-degradation. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO or BRD7/9 degrader (VZ-185) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Techniques Used: CRISPR, Genome Wide, Growth Assay, Control, Expressing, Flow Cytometry, Transformation Assay, Comparison, Activity Assay

a , Genes up- and down-regulated after TRIP1 treatment. 3’ RNA fingerprinting in KARPAS-422 cells after 8h (left) or 16h (right) treatment with 2 µM TRIP1. BCL6 target genes involved in apoptosis, B-cell differentiation, and cell-cycle are highlighted in brown/bold. Hits: Log 2 FC ≥ |1| and P adj < 0.05 (dotted lines). b , Gene expression changes (3’ RNA fingerprinting) in KARPAS-422 cells of selected BCL6 target genes after treatment with TRIP1 or negative control compounds at two different time points and concentrations. c , Lollipop plot summarizing gene set enrichment analysis (GSEA) of BCL6 target genes and genes shown to be regulated by BCL6 targeting TCIPs in Extended Data Figure 4d. Normalized enrichment scores are plotted for each condition compared to DMSO. d , LISA analysis of differentially expressed genes (DEGs) after TRIP1 treatment (16h, 0.5 µM). BCL6 (orange) is the top predicted transcriptional regulator of DEGs after TRIP1 treatment. Other transcriptional repressors with a high score are highlighted in brown. For a-d , differential gene expression analysis was performed using DESeq2. Log₂ fold changes were shrunk using the apeglm method. P-values were obtained from the Wald test and adjusted for multiple testing using the Benjamini-Hochberg procedure. Genes with an adjusted p-value (padj) < 0.05 and |log₂FC| > 1 are highlighted as differentially expressed; n= 3 independent replicates. e , Validation of TRIP1-induced expression changes at the protein level. Time-resolved (left) and dose-resolved (right) expression changes of BCL6, ARID3A, FOXO3 and p21 (CDKN1A) in KARPAS-422 cells. Loading controls were performed per gel. Western blots are representative of two independent replicates.

Figure Legend Snippet: a , Genes up- and down-regulated after TRIP1 treatment. 3’ RNA fingerprinting in KARPAS-422 cells after 8h (left) or 16h (right) treatment with 2 µM TRIP1. BCL6 target genes involved in apoptosis, B-cell differentiation, and cell-cycle are highlighted in brown/bold. Hits: Log 2 FC ≥ |1| and P adj < 0.05 (dotted lines). b , Gene expression changes (3’ RNA fingerprinting) in KARPAS-422 cells of selected BCL6 target genes after treatment with TRIP1 or negative control compounds at two different time points and concentrations. c , Lollipop plot summarizing gene set enrichment analysis (GSEA) of BCL6 target genes and genes shown to be regulated by BCL6 targeting TCIPs in Extended Data Figure 4d. Normalized enrichment scores are plotted for each condition compared to DMSO. d , LISA analysis of differentially expressed genes (DEGs) after TRIP1 treatment (16h, 0.5 µM). BCL6 (orange) is the top predicted transcriptional regulator of DEGs after TRIP1 treatment. Other transcriptional repressors with a high score are highlighted in brown. For a-d , differential gene expression analysis was performed using DESeq2. Log₂ fold changes were shrunk using the apeglm method. P-values were obtained from the Wald test and adjusted for multiple testing using the Benjamini-Hochberg procedure. Genes with an adjusted p-value (padj) < 0.05 and |log₂FC| > 1 are highlighted as differentially expressed; n= 3 independent replicates. e , Validation of TRIP1-induced expression changes at the protein level. Time-resolved (left) and dose-resolved (right) expression changes of BCL6, ARID3A, FOXO3 and p21 (CDKN1A) in KARPAS-422 cells. Loading controls were performed per gel. Western blots are representative of two independent replicates.

Techniques Used: Cell Differentiation, Gene Expression, Negative Control, Biomarker Discovery, Expressing, Western Blot

a, CUT&RUN binding profiles at BCL6 peaks. Binding profiles of BCL6 (left) and SMARCA4 (right) at BCL6 peaks for DMSO/TRIP1 (2 µM) treated cells after 4 and 8 hours. BCL6 peaks were called from DMSO-treated KARPAS-422 samples. Profiles were centered on peaks and extended 5 kb upstream and downstream of the peak location. b , Correlation between BCL6 and SMARCA4 (top), BCL6 and H3K27ac (middle), or SMARCA4 and H3K27ac (bottom) signal changes on gene bodies upon TRIP1 (8h, 2 µM) treatment compared to DMSO. Single dots represent hg38 genes. c, Relationship between TRIP1-induced BCL6/SMARCA4 binding change and H3K27 acetylation or SMARCA4 binding. Genes were ranked based on their differential binding of BCL6 (top) and SMARCA4 (bottom) after TRIP1 treatment (8h, 2 µM) and segmented into respective 20% quantiles. SMARCA4 (top) and H3K27ac (middle) changes at genes per BCL6 quintile. H3K27ac change (bottom) at genes segmented by SMARCA4 quintiles. After overall association was assessed by Kruskal-Wallis was successfully, pairwise comparisons were made by Dunn’s post-hoc test with Benjamini-Hochberg FDR correction. d , Relationship of TRIP1-induced gene expression changes with BCL6/SMARCA4 binding changes after 4 and 8 hours of TRIP1 treatment. Pearson correlation coefficient and R 2 were calculated. Dots represent single genes colored by their gene expression change upon TRIP1 treatment (2 µM after 16 hours vs. DMSO). Selected differentially expressed genes and BCL6 are highlighted. For b-d , Normalized CUT&RUN signal on gene bodies ±3 kb up and downstream to include regulatory regions was calculated for TRIP1 and DMSO. The scores were subtracted to calculate differential binding. e , CUT&RUN binding profiles at BCL6 peaks. Binding profiles of BCL6 at BCL6 peaks after short, low-dose TRIP1 (1 µM) treatment up to 2h. BCL6 peaks were called from DMSO-treated KARPAS-422 CUT&RUN samples. Profiles were centered on peaks and extended 5 kb upstream and downstream of the peak location. f-h , Genome tracks of the ARID3A gene locus. Time-resolved ( f ) BCL6 and SMARCA4 or ( g ) RNA Pol II serine 2/5 phosphorylation signal is computed along the gene locus to infer transcriptional dynamics. For g , Below, RNA-seq reads are mapped to the gene locus. h , BAF ATPase-dependent BCL6 eviction after 1 hour of DMSO or TRIP1 (1 µM) co-treatment with BRM-014 (1 µM). BCL6signal is computed along the ARID3A gene locus. Exon position and genome location are indicated below the genome tracks. All CUT&RUN data is from two merged independent replicates. i , CaspaseGlo 3/7 apoptosis ATPase pre-inhibition. KARPAS-422 cells were pre-treated with DMSO or SMARCA2/4 ATPase inhibitor (BRM-014) for 8 hours, followed by co-treatment with 1 µM of TRIP1 for 16 hours. Caspase 3/7 activity is normalized to DMSO control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates. j , BCL6 transcriptional reporter ATPase pre-inhibition. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO or SMARCA2/4 ATPase inhibitor (BRM-014) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Figure Legend Snippet: a, CUT&RUN binding profiles at BCL6 peaks. Binding profiles of BCL6 (left) and SMARCA4 (right) at BCL6 peaks for DMSO/TRIP1 (2 µM) treated cells after 4 and 8 hours. BCL6 peaks were called from DMSO-treated KARPAS-422 samples. Profiles were centered on peaks and extended 5 kb upstream and downstream of the peak location. b , Correlation between BCL6 and SMARCA4 (top), BCL6 and H3K27ac (middle), or SMARCA4 and H3K27ac (bottom) signal changes on gene bodies upon TRIP1 (8h, 2 µM) treatment compared to DMSO. Single dots represent hg38 genes. c, Relationship between TRIP1-induced BCL6/SMARCA4 binding change and H3K27 acetylation or SMARCA4 binding. Genes were ranked based on their differential binding of BCL6 (top) and SMARCA4 (bottom) after TRIP1 treatment (8h, 2 µM) and segmented into respective 20% quantiles. SMARCA4 (top) and H3K27ac (middle) changes at genes per BCL6 quintile. H3K27ac change (bottom) at genes segmented by SMARCA4 quintiles. After overall association was assessed by Kruskal-Wallis was successfully, pairwise comparisons were made by Dunn’s post-hoc test with Benjamini-Hochberg FDR correction. d , Relationship of TRIP1-induced gene expression changes with BCL6/SMARCA4 binding changes after 4 and 8 hours of TRIP1 treatment. Pearson correlation coefficient and R 2 were calculated. Dots represent single genes colored by their gene expression change upon TRIP1 treatment (2 µM after 16 hours vs. DMSO). Selected differentially expressed genes and BCL6 are highlighted. For b-d , Normalized CUT&RUN signal on gene bodies ±3 kb up and downstream to include regulatory regions was calculated for TRIP1 and DMSO. The scores were subtracted to calculate differential binding. e , CUT&RUN binding profiles at BCL6 peaks. Binding profiles of BCL6 at BCL6 peaks after short, low-dose TRIP1 (1 µM) treatment up to 2h. BCL6 peaks were called from DMSO-treated KARPAS-422 CUT&RUN samples. Profiles were centered on peaks and extended 5 kb upstream and downstream of the peak location. f-h , Genome tracks of the ARID3A gene locus. Time-resolved ( f ) BCL6 and SMARCA4 or ( g ) RNA Pol II serine 2/5 phosphorylation signal is computed along the gene locus to infer transcriptional dynamics. For g , Below, RNA-seq reads are mapped to the gene locus. h , BAF ATPase-dependent BCL6 eviction after 1 hour of DMSO or TRIP1 (1 µM) co-treatment with BRM-014 (1 µM). BCL6signal is computed along the ARID3A gene locus. Exon position and genome location are indicated below the genome tracks. All CUT&RUN data is from two merged independent replicates. i , CaspaseGlo 3/7 apoptosis ATPase pre-inhibition. KARPAS-422 cells were pre-treated with DMSO or SMARCA2/4 ATPase inhibitor (BRM-014) for 8 hours, followed by co-treatment with 1 µM of TRIP1 for 16 hours. Caspase 3/7 activity is normalized to DMSO control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates. j , BCL6 transcriptional reporter ATPase pre-inhibition. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO or SMARCA2/4 ATPase inhibitor (BRM-014) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Techniques Used: Binding Assay, Gene Expression, Phospho-proteomics, RNA Sequencing, Inhibition, Activity Assay, Control, Expressing

Image Search Results

BCL6 in T cells promotes spontaneous GC formation, anti-insulin B cell infiltration of islets, and spontaneous diabetes development in VH125 SD .NOD mice Cells from spleen, pLNs, mLNs, and pancreas were isolated from 8 to 12-week-old VH125 SD .NOD and VH125 SD .Bcl6 ΔCD4 .NOD mice (genotypes fully defined in ). (A) A representative flow cytometry plot from spleen shows the frequency of insulin-binding B cells (insulin+) identified using biotinylated human insulin/streptavidin fluorochrome as in among total B cells (live singlet CD45 + CD19 + lymphocytes). (B) Diabetes was monitored in cohorts of female VH125 SD .NOD mice ( n = 14, black line) and VH125 SD . Bcl6 ΔCD4 .NOD littermates ( n = 14, purple line) from 10 to 35 weeks of age. Mice were considered diabetic after two consecutive blood glucose readings >250 mg/dL, p < 0.0001, log-rank test. (C–G) Representative flow cytometry plots of pancreatic draining lymph nodes and pancreata gating on anti-insulin B cells as in (A) and as described in for pancreas are shown (C). The frequency of anti-insulin B cells (among total B cells) in (D) spleen, (E) mesenteric lymph nodes (mLNs), (F) pancreatic draining lymph nodes, and (G) pancreata are plotted for individual mice of the indicated genotypes. (H–L) Representative flow cytometry plots from the pancreatic lymph nodes (pLNs) of (H) Tfh cells (live singlet CD45 + CD4 + PD-1 hi CXCR5 hi Foxp3 − lymphocytes) are shown with (I and J) frequencies among total CD4 + Foxp3- CD45+ cells and (K and L) numbers of Tfh cells in pLNs and pancreata plotted for individual mice. (M–Q) Representative flow plots of (M) GC B cells from the pancreatic lymph nodes (live singlet CD45 + CD19 + Fas + GL7 + lymphocytes) with (N and O) frequencies of GC B cells among total B cells and (P and Q) numbers shown for pLNs and pancreata. (C–Q) n = 6–8 mice per group, 5 independent experiments, Mann-Whitney U test, bars representative of mean ± standard deviation. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ns, not significant.

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Figure Lengend Snippet: BCL6 in T cells promotes spontaneous GC formation, anti-insulin B cell infiltration of islets, and spontaneous diabetes development in VH125 SD .NOD mice Cells from spleen, pLNs, mLNs, and pancreas were isolated from 8 to 12-week-old VH125 SD .NOD and VH125 SD .Bcl6 ΔCD4 .NOD mice (genotypes fully defined in ). (A) A representative flow cytometry plot from spleen shows the frequency of insulin-binding B cells (insulin+) identified using biotinylated human insulin/streptavidin fluorochrome as in among total B cells (live singlet CD45 + CD19 + lymphocytes). (B) Diabetes was monitored in cohorts of female VH125 SD .NOD mice ( n = 14, black line) and VH125 SD . Bcl6 ΔCD4 .NOD littermates ( n = 14, purple line) from 10 to 35 weeks of age. Mice were considered diabetic after two consecutive blood glucose readings >250 mg/dL, p < 0.0001, log-rank test. (C–G) Representative flow cytometry plots of pancreatic draining lymph nodes and pancreata gating on anti-insulin B cells as in (A) and as described in for pancreas are shown (C). The frequency of anti-insulin B cells (among total B cells) in (D) spleen, (E) mesenteric lymph nodes (mLNs), (F) pancreatic draining lymph nodes, and (G) pancreata are plotted for individual mice of the indicated genotypes. (H–L) Representative flow cytometry plots from the pancreatic lymph nodes (pLNs) of (H) Tfh cells (live singlet CD45 + CD4 + PD-1 hi CXCR5 hi Foxp3 − lymphocytes) are shown with (I and J) frequencies among total CD4 + Foxp3- CD45+ cells and (K and L) numbers of Tfh cells in pLNs and pancreata plotted for individual mice. (M–Q) Representative flow plots of (M) GC B cells from the pancreatic lymph nodes (live singlet CD45 + CD19 + Fas + GL7 + lymphocytes) with (N and O) frequencies of GC B cells among total B cells and (P and Q) numbers shown for pLNs and pancreata. (C–Q) n = 6–8 mice per group, 5 independent experiments, Mann-Whitney U test, bars representative of mean ± standard deviation. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ns, not significant.

Article Snippet: NOD Bcl6 fl/fl , The Jackson Laboratory and Alexander Dent Laboratory , See also: The Jackson Laboratory, Strain 001976 and https://pubmed.ncbi.nlm.nih.gov/39556799/.

Techniques: Isolation, Flow Cytometry, Binding Assay, MANN-WHITNEY, Standard Deviation

T cell expression of BCL6 supports enhanced insulitis severity, but is dispensable for tertiary lymphoid structure organization in VH125 SD .NOD mice Pancreata were harvested from female, pre-diabetic VH125 SD .NOD and VH125 SD Bcl6 ΔCD4 .NOD mice at 8–12 and 13–16 weeks of age and were formalin fixed, and paraffin embedded. (A–H) About 10 μm pancreas sections were stained with hematoxylin and eosin (H&E) and blind scored, with individual mice plotted. (A and E) Representative H&E-stained sections are shown, with arrows pointing to islets. All islets were blind scored, with average insulitis scores shown for control VH125 SD .NOD mice (black) and VH125 SD Bcl6 ΔCD4 .NOD mice (purple) at (B) 8–12 weeks and (F) 13–16 weeks. The percentage of islets, which had no lymphocytic infiltrate present were calculated for (C) 8–12 weeks and (G) 13–16 weeks. (D and H) The percentage of islets with each score for all pancreata for 8–12 and 13–16 weeks is shown. (I–L) Pancreas sections from the 8–16-week-old cohort that had the highest insulitis infiltrate ( n = 6 mice per group) were obtained and stained with antibodies reactive against CD3 (T cells) or B220 (B cells). (I) Representative images show “disorganized” T cell-B cell infiltration (top), and “organized” TLS (bottom). Islets were scored separately for T cell (CD3) and B cell (B220) infiltrate as follows: 0 (no T/B infiltrate), 1 (>25% infiltrate), 2 (25%–50% infiltrate), 3 (50%–75% infiltrate), and 4 (>75% infiltrate). Average infiltration score for (J) CD3 + and (K) B220+ cells in islets is shown. (L) Infiltrated islets that scored 2 or above were blind scored as organized (blue) or disorganized (red) in both VH125 SD .NOD mice and VH125 SD Bcl6 ΔCD4 .NOD, n = number of islets scored. (A–L) n = 6–8 mice per group, Bars represent mean ± standard deviation, ns, not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, (B, C, F, G, J, and K) Mann-Whitney U test or (L) chi-square test.

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Figure Lengend Snippet: T cell expression of BCL6 supports enhanced insulitis severity, but is dispensable for tertiary lymphoid structure organization in VH125 SD .NOD mice Pancreata were harvested from female, pre-diabetic VH125 SD .NOD and VH125 SD Bcl6 ΔCD4 .NOD mice at 8–12 and 13–16 weeks of age and were formalin fixed, and paraffin embedded. (A–H) About 10 μm pancreas sections were stained with hematoxylin and eosin (H&E) and blind scored, with individual mice plotted. (A and E) Representative H&E-stained sections are shown, with arrows pointing to islets. All islets were blind scored, with average insulitis scores shown for control VH125 SD .NOD mice (black) and VH125 SD Bcl6 ΔCD4 .NOD mice (purple) at (B) 8–12 weeks and (F) 13–16 weeks. The percentage of islets, which had no lymphocytic infiltrate present were calculated for (C) 8–12 weeks and (G) 13–16 weeks. (D and H) The percentage of islets with each score for all pancreata for 8–12 and 13–16 weeks is shown. (I–L) Pancreas sections from the 8–16-week-old cohort that had the highest insulitis infiltrate ( n = 6 mice per group) were obtained and stained with antibodies reactive against CD3 (T cells) or B220 (B cells). (I) Representative images show “disorganized” T cell-B cell infiltration (top), and “organized” TLS (bottom). Islets were scored separately for T cell (CD3) and B cell (B220) infiltrate as follows: 0 (no T/B infiltrate), 1 (>25% infiltrate), 2 (25%–50% infiltrate), 3 (50%–75% infiltrate), and 4 (>75% infiltrate). Average infiltration score for (J) CD3 + and (K) B220+ cells in islets is shown. (L) Infiltrated islets that scored 2 or above were blind scored as organized (blue) or disorganized (red) in both VH125 SD .NOD mice and VH125 SD Bcl6 ΔCD4 .NOD, n = number of islets scored. (A–L) n = 6–8 mice per group, Bars represent mean ± standard deviation, ns, not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, (B, C, F, G, J, and K) Mann-Whitney U test or (L) chi-square test.

Article Snippet: NOD Bcl6 fl/fl , The Jackson Laboratory and Alexander Dent Laboratory , See also: The Jackson Laboratory, Strain 001976 and https://pubmed.ncbi.nlm.nih.gov/39556799/.

Techniques: Expressing, Staining, Control, Standard Deviation, MANN-WHITNEY

BCL6 in T cells increases activation and proliferation markers of insulin-binding B cells relative to non-insulin-binding B cells Cells were isolated from 8 to 12-week-old, female, pre-diabetic VH125 SD .NOD with and without Cd4 -Cre Bcl6 deletion from (A–G) spleen, pancreatic lymph nodes (pLNs) and pancreata. (A–C) Representative flow cytometry plots show Ki67 staining overlays of insulin-binding (left) or non-insulin-binding (right) B cells (identified as in ) from each genotype within live singlet GL7- CD95- CD45 + CD19 + lymphocytes in spleen (A), pancreatic lymph nodes (B), and pancreata (C). (D) Non-GC (Fas − GL7 − ) B cells were further gated on insulin-binding (ins+) and non-insulin-binding (ins−) and the frequency of cells that were Ki67+ (a marker of proliferation). (E–G) Insulin+ or insulin− B cell expression of (E) CD86 (T cell co-stimulatory molecule), (F) CD44 (activation marker), and (G) CD69 (activation marker) is shown for spleen (left), pancreatic lymph nodes (middle), and pancreata (right), with n = 6–8 individual mice plotted per group. (D–G) Bars represent mean ± standard deviation. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ns = not significant, Kruskal-Wallis test with post hoc test of multiple comparisons. All other comparisons not shown are not significant.

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Figure Lengend Snippet: BCL6 in T cells increases activation and proliferation markers of insulin-binding B cells relative to non-insulin-binding B cells Cells were isolated from 8 to 12-week-old, female, pre-diabetic VH125 SD .NOD with and without Cd4 -Cre Bcl6 deletion from (A–G) spleen, pancreatic lymph nodes (pLNs) and pancreata. (A–C) Representative flow cytometry plots show Ki67 staining overlays of insulin-binding (left) or non-insulin-binding (right) B cells (identified as in ) from each genotype within live singlet GL7- CD95- CD45 + CD19 + lymphocytes in spleen (A), pancreatic lymph nodes (B), and pancreata (C). (D) Non-GC (Fas − GL7 − ) B cells were further gated on insulin-binding (ins+) and non-insulin-binding (ins−) and the frequency of cells that were Ki67+ (a marker of proliferation). (E–G) Insulin+ or insulin− B cell expression of (E) CD86 (T cell co-stimulatory molecule), (F) CD44 (activation marker), and (G) CD69 (activation marker) is shown for spleen (left), pancreatic lymph nodes (middle), and pancreata (right), with n = 6–8 individual mice plotted per group. (D–G) Bars represent mean ± standard deviation. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ns = not significant, Kruskal-Wallis test with post hoc test of multiple comparisons. All other comparisons not shown are not significant.

Article Snippet: NOD Bcl6 fl/fl , The Jackson Laboratory and Alexander Dent Laboratory , See also: The Jackson Laboratory, Strain 001976 and https://pubmed.ncbi.nlm.nih.gov/39556799/.

Techniques: Activation Assay, Binding Assay, Isolation, Flow Cytometry, Staining, Marker, Expressing, Standard Deviation

BCL6 loss in CD4 + T cells impairs CD4 + T cell activation and promotes Treg formation in some sites but does not affect anti-insulin B cell proliferation in the pancreas CD4 + T cells were isolated from 12 to 14 week-old female donors and ∼5 × 10 6 BCL6+ or ΔBCL6 CD4 + T cells were CellTrace Violet (CTV)-labeled and adoptively transferred into 8–12 week-old VH125 SD .Bcl6 ΔCD4 .NOD recipients. Seven days after transfer, spleen, pancreatic lymph nodes, and pancreata were assayed as follows. (A) Experimental schematic. (B) (Left) Representative flow plots of CD4 + CTV+ labeled cells in FMO control, spleen, pancreatic lymph nodes, and pancreas. (Right) The proportions of CTV+ labeled cells in spleen, pancreatic lymph nodes, and pancreata are shown for individual recipients following BCL6+ or ΔBCL6 CD4 + transfers. (C–F) CTV+ (transferred) or CTV- (endogenous, non-transferred) CD4 + T cells were assessed regarding: (C) CD44 MFI levels, (D) % CD69 + , (E) % Tfh (CXCR5+ PD-1+), and (F) % Treg (FoxP3+). (G–H) % GC B cells (CD95 + GL7+) is shown among total B cells (CD19 + B220+ CD45 + live cells) and (H) % Ki67+ among non-GC (CD95 − GL7 − CD19 + B220+ CD45 + live) B cells in either insulin-binding (INS+) or non-insulin binding (INS−) B cells. (B–H) n = 5–6 mice per group, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ns, not significant, Kruskal-Wallis test with post hoc test of multiple comparisons (B–F and H) or Mann-Whitney U test (G) was used for analyses. All other comparisons not shown are not significant. Bars represent mean ± standard deviation.

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Figure Lengend Snippet: BCL6 loss in CD4 + T cells impairs CD4 + T cell activation and promotes Treg formation in some sites but does not affect anti-insulin B cell proliferation in the pancreas CD4 + T cells were isolated from 12 to 14 week-old female donors and ∼5 × 10 6 BCL6+ or ΔBCL6 CD4 + T cells were CellTrace Violet (CTV)-labeled and adoptively transferred into 8–12 week-old VH125 SD .Bcl6 ΔCD4 .NOD recipients. Seven days after transfer, spleen, pancreatic lymph nodes, and pancreata were assayed as follows. (A) Experimental schematic. (B) (Left) Representative flow plots of CD4 + CTV+ labeled cells in FMO control, spleen, pancreatic lymph nodes, and pancreas. (Right) The proportions of CTV+ labeled cells in spleen, pancreatic lymph nodes, and pancreata are shown for individual recipients following BCL6+ or ΔBCL6 CD4 + transfers. (C–F) CTV+ (transferred) or CTV- (endogenous, non-transferred) CD4 + T cells were assessed regarding: (C) CD44 MFI levels, (D) % CD69 + , (E) % Tfh (CXCR5+ PD-1+), and (F) % Treg (FoxP3+). (G–H) % GC B cells (CD95 + GL7+) is shown among total B cells (CD19 + B220+ CD45 + live cells) and (H) % Ki67+ among non-GC (CD95 − GL7 − CD19 + B220+ CD45 + live) B cells in either insulin-binding (INS+) or non-insulin binding (INS−) B cells. (B–H) n = 5–6 mice per group, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ns, not significant, Kruskal-Wallis test with post hoc test of multiple comparisons (B–F and H) or Mann-Whitney U test (G) was used for analyses. All other comparisons not shown are not significant. Bars represent mean ± standard deviation.

Article Snippet: NOD Bcl6 fl/fl , The Jackson Laboratory and Alexander Dent Laboratory , See also: The Jackson Laboratory, Strain 001976 and https://pubmed.ncbi.nlm.nih.gov/39556799/.

Techniques: Activation Assay, Isolation, Labeling, Control, Binding Assay, MANN-WHITNEY, Standard Deviation

Insulin-binding B cells exist in extrafollicular niches and skew toward CD11c + T-bet + and CD11b + CD11c + atypical B cell subsets, some of which are reduced by loss of Bcl6 Spleen, pancreatic draining lymph nodes, and pancreata were harvested from pre-diabetic, 8–12-week-old VH125 SD .NOD mice. (A) Around 10 μm sections of spleen form VH125 SD .NOD underwent immunofluorescence staining with IgD, Ki67, insulin, and CD3. Representative merge image shown on left with identified borders of B cell zone (BCZ), T cell zone (TCZ), germinal center (GC) and extrafollicular area (EF). Circled cells are representative insulin-autoreactive B cells, which express both IgD and insulin. Quantification of % insulin+ B cells in predefined niches on the left graph. Quantification of insulin+ IgD + colocalization, or insulin+ IgD-. n = 4–6 individual splenic sections per group. (B) Representative flow plots show CD11c+/CD11b+ cells among B220+ CD19 + live singlet lymphocytes for both non-insulin-reactive (insulin−, left) and insulin-autoreactive (insulin+, middle, right) B cell populations in the pancreas. (C) Quantification of the proportion of CD11c+ CD11b+ B cells in pancreata for both insulin− and insulin+ B cells for VH125 SD (black) and VH125 SD Bcl6 ΔCD4 (purple) mice. (D) Representative flow plots of CD11c+/Tbet+ cells among B220+ CD19 + live singlet lymphocytes for both insulin− and insulin+ B cell populations in the pancreas. (E) Quantification of the proportion of CD11c+/Tbet+ cells among both insulin− and insulin+ B cells in the pancreas of VH125 SD (black) and VH125 SD Bcl6 ΔCD4 (purple) mice. (F and G) Quantification of the proportion of CD11c+/Tbet+ B cells and CD11b+/CD11c+ B cells in pancreatic draining lymph nodes (F) and spleen (G). (A–G) One-way ANOVA (A, left), Mann-Whitney U test (A, right) (A) or Kruskal-Wallis test (C–G) with post hoc multiple comparison test were used, n = 4–8 mice per group. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001, ns, not significant. All other comparisons not shown are not significant. Bars represent mean ± standard deviation.

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Figure Lengend Snippet: Insulin-binding B cells exist in extrafollicular niches and skew toward CD11c + T-bet + and CD11b + CD11c + atypical B cell subsets, some of which are reduced by loss of Bcl6 Spleen, pancreatic draining lymph nodes, and pancreata were harvested from pre-diabetic, 8–12-week-old VH125 SD .NOD mice. (A) Around 10 μm sections of spleen form VH125 SD .NOD underwent immunofluorescence staining with IgD, Ki67, insulin, and CD3. Representative merge image shown on left with identified borders of B cell zone (BCZ), T cell zone (TCZ), germinal center (GC) and extrafollicular area (EF). Circled cells are representative insulin-autoreactive B cells, which express both IgD and insulin. Quantification of % insulin+ B cells in predefined niches on the left graph. Quantification of insulin+ IgD + colocalization, or insulin+ IgD-. n = 4–6 individual splenic sections per group. (B) Representative flow plots show CD11c+/CD11b+ cells among B220+ CD19 + live singlet lymphocytes for both non-insulin-reactive (insulin−, left) and insulin-autoreactive (insulin+, middle, right) B cell populations in the pancreas. (C) Quantification of the proportion of CD11c+ CD11b+ B cells in pancreata for both insulin− and insulin+ B cells for VH125 SD (black) and VH125 SD Bcl6 ΔCD4 (purple) mice. (D) Representative flow plots of CD11c+/Tbet+ cells among B220+ CD19 + live singlet lymphocytes for both insulin− and insulin+ B cell populations in the pancreas. (E) Quantification of the proportion of CD11c+/Tbet+ cells among both insulin− and insulin+ B cells in the pancreas of VH125 SD (black) and VH125 SD Bcl6 ΔCD4 (purple) mice. (F and G) Quantification of the proportion of CD11c+/Tbet+ B cells and CD11b+/CD11c+ B cells in pancreatic draining lymph nodes (F) and spleen (G). (A–G) One-way ANOVA (A, left), Mann-Whitney U test (A, right) (A) or Kruskal-Wallis test (C–G) with post hoc multiple comparison test were used, n = 4–8 mice per group. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001, ns, not significant. All other comparisons not shown are not significant. Bars represent mean ± standard deviation.

Article Snippet: NOD Bcl6 fl/fl , The Jackson Laboratory and Alexander Dent Laboratory , See also: The Jackson Laboratory, Strain 001976 and https://pubmed.ncbi.nlm.nih.gov/39556799/.

Techniques: Binding Assay, Immunofluorescence, Staining, MANN-WHITNEY, Comparison, Standard Deviation

BCL6 loss in T cells promotes formation of a phenotypically defined atypical memory B cell population in the pancreas of VH125 SD .NOD mice Pancreata were harvested from pre-diabetic, 8–12-week-old Bcl6 -sufficient (VH125 SD ) and Bcl6 -deficient (VH125 SD Bcl6 ΔCD4 .NOD) mice ( n = 12–14 mice) and flow cytometry staining was performed using the panel of markers in and (D). B220+ CD19 + CD45 + live singlets were concatenated and normalized via CyCombine and concatenated, with genotype metadata encoded. (A) t-SNE was used to perform dimensionality reduction based on phenotypic marker expression profiles. Clusters were manually defined as indicated. (B) The cluster frequency is shown for each genotype, with individual mice plotted and means indicated. ∗ p < 0.05, Kruskal-Wallis test with post hoc multiple comparison test. Bars represent mean ± standard deviation. (C) A rainbow intensity scale indicates expression levels of the phenotypic markers indicated at the top of each t-SNE plot. Insulin-binding, CD45, B220, CD19, and viability dye markers were omitted from the t-SNE analysis given they were used in parent population gating upstream of t-SNE analysis. (D) Heatmap shows relative expression of the indicated markers for each cluster defined as in (A) using a log-like arcsinh scale and appropriate arcsinh factors.

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Figure Lengend Snippet: BCL6 loss in T cells promotes formation of a phenotypically defined atypical memory B cell population in the pancreas of VH125 SD .NOD mice Pancreata were harvested from pre-diabetic, 8–12-week-old Bcl6 -sufficient (VH125 SD ) and Bcl6 -deficient (VH125 SD Bcl6 ΔCD4 .NOD) mice ( n = 12–14 mice) and flow cytometry staining was performed using the panel of markers in and (D). B220+ CD19 + CD45 + live singlets were concatenated and normalized via CyCombine and concatenated, with genotype metadata encoded. (A) t-SNE was used to perform dimensionality reduction based on phenotypic marker expression profiles. Clusters were manually defined as indicated. (B) The cluster frequency is shown for each genotype, with individual mice plotted and means indicated. ∗ p < 0.05, Kruskal-Wallis test with post hoc multiple comparison test. Bars represent mean ± standard deviation. (C) A rainbow intensity scale indicates expression levels of the phenotypic markers indicated at the top of each t-SNE plot. Insulin-binding, CD45, B220, CD19, and viability dye markers were omitted from the t-SNE analysis given they were used in parent population gating upstream of t-SNE analysis. (D) Heatmap shows relative expression of the indicated markers for each cluster defined as in (A) using a log-like arcsinh scale and appropriate arcsinh factors.

Article Snippet: NOD Bcl6 fl/fl , The Jackson Laboratory and Alexander Dent Laboratory , See also: The Jackson Laboratory, Strain 001976 and https://pubmed.ncbi.nlm.nih.gov/39556799/.

Techniques: Flow Cytometry, Staining, Marker, Expressing, Comparison, Standard Deviation, Binding Assay

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Article Snippet: Bcl6 fl/fl Cd4 -Cre.NOD mice were then crossed with VH125 SD .NOD mice, a gift from Dr. James W. Thomas (and available at The Jackson Laboratory, strain ID 029288), in which an anti-insulin IgH transgene is targeted to the IgH locus (site-directed, SD), to generate VH125 SD+/− .Bcl6 fl/fl .

Techniques: Isolation, Flow Cytometry, Binding Assay, MANN-WHITNEY, Standard Deviation

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Techniques: Expressing, Staining, Control, Standard Deviation, MANN-WHITNEY

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Techniques: Activation Assay, Binding Assay, Isolation, Flow Cytometry, Staining, Marker, Expressing, Standard Deviation

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Techniques: Activation Assay, Isolation, Labeling, Control, Binding Assay, MANN-WHITNEY, Standard Deviation

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Techniques: Binding Assay, Immunofluorescence, Staining, MANN-WHITNEY, Comparison, Standard Deviation

Journal: iScience

Article Title: BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

doi: 10.1016/j.isci.2026.115990

Techniques: Flow Cytometry, Staining, Marker, Expressing, Comparison, Standard Deviation, Binding Assay

BCL6 is highly expressed in human and mouse skeletal muscle and is essential for regulating muscle mass and function. (A) RNA-seq data for BCL6 tissue-specific expression pattern in various human tissues obtained from GTEx database. Expression values are shown in transcripts per million (TPM). (B) Unbiased clustering of snRNA-seq data of 5-month-old TA muscle represented on a UMAP (upper panel). UMAP (middle panel) and violin plots (lower panel) showing Bcl6 gene expression for different cell populations. Data obtained from MYOATLAS database. Expression values are shown as a probability distribution across clusters. (C) Expression pattern of Bcl6 from the gastrocnemius of old wild-type mice at various time points obtained from SarcoAtlas database. Expression values are shown in transcripts per million (TPM). (D) Quantification of Bcl6 mRNA expression in various muscles and non-muscle tissue from 10 to 12-week-old mice assessed by RT-qPCR. Data was normalized by the housekeeping gene 36B4 and expressed as fold change compared to M- Bcl6 WT. n = 4–9 per group. (E) Body mass of M- Bcl6 WT and KO male mice at the indicated ages. n = 4–9 per group. (F) Representative images of the various muscle groups from the M- Bcl6 WT (left) and M- Bcl6 KO (right) mice at 10–12 weeks of age. Scale bar = 0.5 cm. (G) Tissue weights of various muscles from male M- Bcl6 WT and M- Bcl6 KO mice at 10–12 weeks of age. Tissue weights are shown as percent of those from M- Bcl6 WT. n = 8 per group. (H) Tissue weights of various muscles from male M- Bcl6 WT and KO mice normalized to body mass at 10–12 weeks of age. Normalized data are shown as percent of M- Bcl6 WT. n = 8 per group. (I) Absolute forelimb grip strength measured in male M- Bcl6 WT and M- Bcl6 KO mice at 8 weeks of age. n = 13–14 per group. (J) Absolute tetanic force of gastrocnemius muscle of male M- Bcl6 WT and KO mice at 12–15 weeks of age. n = 7–8 per group. (K) Representative images of laminin staining of the TA muscle of male M- Bcl6 WT and KO mice, Scale bar = 50 μm; and (L) quantification of their myofiber cross sectional area (CSA) and (M) their average CSA. n = 4–6 per group. Data in D, E, G, H and L were analyzed with two-way ANOVA and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data in I, J, and M were analyzed with unpaired two tailed Student's t- test (∗p < 0.05 and ∗∗p < 0.01). Data are presented as mean ± SEM (with individual data points).

Journal: Molecular Metabolism

Article Title: BCL6 regulates skeletal muscle mass and mitochondrial bioenergetics

doi: 10.1016/j.molmet.2026.102367

Figure Lengend Snippet: BCL6 is highly expressed in human and mouse skeletal muscle and is essential for regulating muscle mass and function. (A) RNA-seq data for BCL6 tissue-specific expression pattern in various human tissues obtained from GTEx database. Expression values are shown in transcripts per million (TPM). (B) Unbiased clustering of snRNA-seq data of 5-month-old TA muscle represented on a UMAP (upper panel). UMAP (middle panel) and violin plots (lower panel) showing Bcl6 gene expression for different cell populations. Data obtained from MYOATLAS database. Expression values are shown as a probability distribution across clusters. (C) Expression pattern of Bcl6 from the gastrocnemius of old wild-type mice at various time points obtained from SarcoAtlas database. Expression values are shown in transcripts per million (TPM). (D) Quantification of Bcl6 mRNA expression in various muscles and non-muscle tissue from 10 to 12-week-old mice assessed by RT-qPCR. Data was normalized by the housekeeping gene 36B4 and expressed as fold change compared to M- Bcl6 WT. n = 4–9 per group. (E) Body mass of M- Bcl6 WT and KO male mice at the indicated ages. n = 4–9 per group. (F) Representative images of the various muscle groups from the M- Bcl6 WT (left) and M- Bcl6 KO (right) mice at 10–12 weeks of age. Scale bar = 0.5 cm. (G) Tissue weights of various muscles from male M- Bcl6 WT and M- Bcl6 KO mice at 10–12 weeks of age. Tissue weights are shown as percent of those from M- Bcl6 WT. n = 8 per group. (H) Tissue weights of various muscles from male M- Bcl6 WT and KO mice normalized to body mass at 10–12 weeks of age. Normalized data are shown as percent of M- Bcl6 WT. n = 8 per group. (I) Absolute forelimb grip strength measured in male M- Bcl6 WT and M- Bcl6 KO mice at 8 weeks of age. n = 13–14 per group. (J) Absolute tetanic force of gastrocnemius muscle of male M- Bcl6 WT and KO mice at 12–15 weeks of age. n = 7–8 per group. (K) Representative images of laminin staining of the TA muscle of male M- Bcl6 WT and KO mice, Scale bar = 50 μm; and (L) quantification of their myofiber cross sectional area (CSA) and (M) their average CSA. n = 4–6 per group. Data in D, E, G, H and L were analyzed with two-way ANOVA and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data in I, J, and M were analyzed with unpaired two tailed Student's t- test (∗p < 0.05 and ∗∗p < 0.01). Data are presented as mean ± SEM (with individual data points).

Article Snippet: Myf6-Cre ± mice (Jackson Laboratories, 010528) were bred with Bcl6 fl/fl mice (Jackson Laboratories, 023727) to generate Myf6-Cre + BCL6 fl/wt mice, which were then intercrossed to generate Myf6-Cre + Bcl6 fl/fl mice, herein referred to as M- Bcl6 KO.

Techniques: RNA Sequencing, Expressing, Gene Expression, Muscles, Quantitative RT-PCR, Staining, Two Tailed Test

AAV-KD of Bcl6 in adult mice causes muscle atrophy and weakness. (A) Schematic representation of experimental design created with BioRender.com . (B) Quantification of Bcl6 mRNA expression from male mice injected with either AAV-scrmb or AAV-sh Bcl6 knockdown (KD) in tibialis anterior (TA) muscle assessed by RT-qPCR. n = 7–8 per group. (C) Tissue mass of TA muscle from male mice injected with either AAV-scrmb and AAV-sh Bcl6 KD muscle. n = 8 per group. (D) Tissue mass of gastrocnemius (GAS) muscle from male mice injected with either AAV-scrmb and AAV-sh Bcl6 KD muscle. n = 8 per group. (E) Representative myosin heavy chain (MHC) immunolabelling of fiber type proportion in TA muscle of male mice injected with either AAV-scrmb or AAV-sh Bcl6 KD. Large Scale bars = 200 μm, inset scale bar = 50 μm. (F) Analysis of fiber type proportion in TA muscle of male mice injected with either AAV-scrmb or AAV-sh Bcl6 knockdown. n = 4–5 per group. (G) Average cross-sectional area (CSA) of TA muscle of male mice injected with either AAV-scrmb or AAV-sh Bcl6 knockdown. n = 11 per group. (H) In vivo isometric plantarflexor torque measured at 4 weeks post-AAV injection. n = 11 per group. Data in B, C, D, F and G were analyzed with paired two-tailed Student's t -test (∗p < 0.05). Data in H was analyzed with two-way ANOVA and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data are presented as mean ± SEM (with individual data points).

Journal: Molecular Metabolism

Article Title: BCL6 regulates skeletal muscle mass and mitochondrial bioenergetics

doi: 10.1016/j.molmet.2026.102367

Figure Lengend Snippet: AAV-KD of Bcl6 in adult mice causes muscle atrophy and weakness. (A) Schematic representation of experimental design created with BioRender.com . (B) Quantification of Bcl6 mRNA expression from male mice injected with either AAV-scrmb or AAV-sh Bcl6 knockdown (KD) in tibialis anterior (TA) muscle assessed by RT-qPCR. n = 7–8 per group. (C) Tissue mass of TA muscle from male mice injected with either AAV-scrmb and AAV-sh Bcl6 KD muscle. n = 8 per group. (D) Tissue mass of gastrocnemius (GAS) muscle from male mice injected with either AAV-scrmb and AAV-sh Bcl6 KD muscle. n = 8 per group. (E) Representative myosin heavy chain (MHC) immunolabelling of fiber type proportion in TA muscle of male mice injected with either AAV-scrmb or AAV-sh Bcl6 KD. Large Scale bars = 200 μm, inset scale bar = 50 μm. (F) Analysis of fiber type proportion in TA muscle of male mice injected with either AAV-scrmb or AAV-sh Bcl6 knockdown. n = 4–5 per group. (G) Average cross-sectional area (CSA) of TA muscle of male mice injected with either AAV-scrmb or AAV-sh Bcl6 knockdown. n = 11 per group. (H) In vivo isometric plantarflexor torque measured at 4 weeks post-AAV injection. n = 11 per group. Data in B, C, D, F and G were analyzed with paired two-tailed Student's t -test (∗p < 0.05). Data in H was analyzed with two-way ANOVA and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data are presented as mean ± SEM (with individual data points).

Techniques: Expressing, Injection, Knockdown, Quantitative RT-PCR, In Vivo, Two Tailed Test

Single-nucleus RNA sequencing shows no significant changes to fibre type composition of muscles with Bcl6 KO. (A) Schematic representation of experimental design created with BioRender.com . (B) Unbiased clustering showing nuclear transcriptomes of snRNA-seq data in 6-week-old M- Bcl6 WT (left) and M- Bcl6 KO (right) TA muscle represented on a UMAP, colored by cluster identity. (C) Dot plot from snRNA-seq showing gene markers used for cluster identification. Darker-colored dots indicate higher expression, and larger dots indicate a greater percentage of nuclei expression of the corresponding gene. (D) Bar graph showing proportions (left) and percentages (right) of the composition of nuclei showing myofibre and cell type in M- Bcl6 WT and KO TA muscle. (E) Unbiased clustering of snRNA-seq data for myonuclear population in M- Bcl6 WT (left) and KO (right) TA muscle represented on a UMAP, colored by cluster identity. (F) Bar graph showing proportions (left) and percentages (right) of the composition of myonuclei indicating fibre type in M- Bcl6 WT and KO TA muscle. The right and left TA muscles from 2 male mice were used per group.

Journal: Molecular Metabolism

Article Title: BCL6 regulates skeletal muscle mass and mitochondrial bioenergetics

doi: 10.1016/j.molmet.2026.102367

Figure Lengend Snippet: Single-nucleus RNA sequencing shows no significant changes to fibre type composition of muscles with Bcl6 KO. (A) Schematic representation of experimental design created with BioRender.com . (B) Unbiased clustering showing nuclear transcriptomes of snRNA-seq data in 6-week-old M- Bcl6 WT (left) and M- Bcl6 KO (right) TA muscle represented on a UMAP, colored by cluster identity. (C) Dot plot from snRNA-seq showing gene markers used for cluster identification. Darker-colored dots indicate higher expression, and larger dots indicate a greater percentage of nuclei expression of the corresponding gene. (D) Bar graph showing proportions (left) and percentages (right) of the composition of nuclei showing myofibre and cell type in M- Bcl6 WT and KO TA muscle. (E) Unbiased clustering of snRNA-seq data for myonuclear population in M- Bcl6 WT (left) and KO (right) TA muscle represented on a UMAP, colored by cluster identity. (F) Bar graph showing proportions (left) and percentages (right) of the composition of myonuclei indicating fibre type in M- Bcl6 WT and KO TA muscle. The right and left TA muscles from 2 male mice were used per group.

Techniques: RNA Sequencing, Muscles, Expressing

Bcl6 regulates Socs2 expression, the IGF1/AKT signalling pathway and proteostasis: (A) Volcano plot showing upregulated (red) and downregulated (blue) genes in M- Bcl6 KO vs M- Bcl6 WT male mice from snRNA-seq data. ( B ) UMAP plot of the myonuclear gene expression of Socs2 in 6-week-old male M- Bcl6 WT (top) and M- Bcl6 KO (bottom) mice derived from snRNA-seq data. ( C ) Quantification of Socs2 and Igf1 mRNA expression in quadriceps of 10–12-week-old male M- Bcl6 WT and M- Bcl6 KO mice obtained by RT-qPCR. mRNA expression levels are shown relative to housekeeping gene β-actin and are expressed as fold change compared to M- Bcl6 WT. n = 8–9 per group. ( D ) snATAC-seq data showing increased chromatin accessibility for Socs2 in 6-week-old male M- Bcl6 KO mice vs M- Bcl6 WT. ( E ) Immunoblot detection (left) of AKT and p-AKT from TA muscles and corresponding quantification (right) of the p-AKT/AKT ratio in 10–12-week-old male M- Bcl6 WT and M- Bcl6 KO mice. n = 4 per group. ( F ) UMAP plot of the myonuclear gene expression of Retreg1 (also known as Fam134b) in 6-week-old male M- Bcl6 WT (top) and M- Bcl6 KO (bottom) mice derived from snRNA-seq data. ( G ) Immunoblot detection (left) of Retreg1 from quadriceps of 10–12-week-old male M- Bcl6 WT and KO mice and corresponding quantification (right). n = 5 per group. Data in C, F and H were analyzed with paired two-tailed Student's t -test (∗p < 0.05). Data are presented as mean ± SEM (with individual data points).

Journal: Molecular Metabolism

Article Title: BCL6 regulates skeletal muscle mass and mitochondrial bioenergetics

doi: 10.1016/j.molmet.2026.102367

Figure Lengend Snippet: Bcl6 regulates Socs2 expression, the IGF1/AKT signalling pathway and proteostasis: (A) Volcano plot showing upregulated (red) and downregulated (blue) genes in M- Bcl6 KO vs M- Bcl6 WT male mice from snRNA-seq data. ( B ) UMAP plot of the myonuclear gene expression of Socs2 in 6-week-old male M- Bcl6 WT (top) and M- Bcl6 KO (bottom) mice derived from snRNA-seq data. ( C ) Quantification of Socs2 and Igf1 mRNA expression in quadriceps of 10–12-week-old male M- Bcl6 WT and M- Bcl6 KO mice obtained by RT-qPCR. mRNA expression levels are shown relative to housekeeping gene β-actin and are expressed as fold change compared to M- Bcl6 WT. n = 8–9 per group. ( D ) snATAC-seq data showing increased chromatin accessibility for Socs2 in 6-week-old male M- Bcl6 KO mice vs M- Bcl6 WT. ( E ) Immunoblot detection (left) of AKT and p-AKT from TA muscles and corresponding quantification (right) of the p-AKT/AKT ratio in 10–12-week-old male M- Bcl6 WT and M- Bcl6 KO mice. n = 4 per group. ( F ) UMAP plot of the myonuclear gene expression of Retreg1 (also known as Fam134b) in 6-week-old male M- Bcl6 WT (top) and M- Bcl6 KO (bottom) mice derived from snRNA-seq data. ( G ) Immunoblot detection (left) of Retreg1 from quadriceps of 10–12-week-old male M- Bcl6 WT and KO mice and corresponding quantification (right). n = 5 per group. Data in C, F and H were analyzed with paired two-tailed Student's t -test (∗p < 0.05). Data are presented as mean ± SEM (with individual data points).

Techniques: Expressing, Gene Expression, Derivative Assay, Quantitative RT-PCR, Western Blot, Muscles, Two Tailed Test

Ultrastructural abnormalities and mitochondrial dysfunction in Bcl6 KO muscle. (A) Manhattan plot of ontology terms and pathways associated with Bcl6 gene function in skeletal muscle tissue. All terms significantly associated with Bcl6 expression in muscle tissue (GMAS ≥0.268, <=-0.268) are highlighted. Data obtained from GeneBridge database. (B) Electron micrographs of M- Bcl6 WT (upper panel) and M- Bcl6 KO (lower panel) gastrocnemius (GAS) muscle at 10–12 weeks of age. Scale bar = 1 μm. Black arrows represent glycogen accumulation and white arrow represents altered mitochondria morphology. (C) Quantification of mitochondrial density analyzed from electron micrographs of M- Bcl6 WT and KO GAS muscle. (D) Mean quantification of morphometric and shape descriptors of intermyofibrillar mitochondria, including Form Factor (FF), Roundness, Aspect Ratio (AR), Feret's diameters (Feret), Circularity (Circ), Perimeter (Perim), and Area in male M- Bcl6 WT and KO mice. (E) Quantification of genes involved in mitochondrial quality control processes in quadriceps of 10-week-old male M- Bcl6 WT and KO mice. mRNA expression levels are shown relative to housekeeping gene β-actin and are expressed as fold change compared to M- Bcl6 WT as assessed by RT-qPCR. n = 6–9 per group. (F) Immunoblot detection (right) and corresponding protein quantification (left) of OPA1, VDAC1, Mfn2, and Drp1 in quadriceps of male M- Bcl6 WT and M–KO mice. n = 5 per group. (G) Mitochondrial DNA (mtDNA) copy number calculated as a ratio of mitochondrial encoded genes ( Cox2 or 16S ) to nuclear encoded genes ( CycA or HK2 ) as measured by RT-qPCR from quadriceps of 12-week-old M- Bcl6 WT and M- Bcl6 KO mice. n = 7 per group. (H) Quantification of mitochondrial biogenesis genes in quadriceps of 10-week-old male M- Bcl6 WT and KO mice. mRNA expression levels are shown relative to housekeeping gene β-actin and are expressed as fold change compared to M- Bcl6 WT assessed by RT-qPCR. n = 8–9 per group. (I) Immunoblot detection (left) and corresponding protein quantification (right) of representative subunits of the mitochondrial oxidative phosphorylation (mtOxPhos) from quadriceps of male M- Bcl6 WT and M- Bcl6 KO mice. n = 5 per group. ( J ) Mitochondrial respiration of male and female M- Bcl6 WT and M- Bcl6 KO mice performed by the addition of pyruvate and malate (PM) in permeabilized muscle fibers across a range of ADP concentrations, and succinate (Succ) in the presence of creatine. n = 7–11 per group. Data in C, D, E, F, H, and I were analyzed by unpaired two-tailed Student's t -test (∗p < 0.05). Data in J was analyzed with two-way ANOVA, and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data are presented as mean ± SEM (with individual data points).

Journal: Molecular Metabolism

Article Title: BCL6 regulates skeletal muscle mass and mitochondrial bioenergetics

doi: 10.1016/j.molmet.2026.102367

Figure Lengend Snippet: Ultrastructural abnormalities and mitochondrial dysfunction in Bcl6 KO muscle. (A) Manhattan plot of ontology terms and pathways associated with Bcl6 gene function in skeletal muscle tissue. All terms significantly associated with Bcl6 expression in muscle tissue (GMAS ≥0.268, <=-0.268) are highlighted. Data obtained from GeneBridge database. (B) Electron micrographs of M- Bcl6 WT (upper panel) and M- Bcl6 KO (lower panel) gastrocnemius (GAS) muscle at 10–12 weeks of age. Scale bar = 1 μm. Black arrows represent glycogen accumulation and white arrow represents altered mitochondria morphology. (C) Quantification of mitochondrial density analyzed from electron micrographs of M- Bcl6 WT and KO GAS muscle. (D) Mean quantification of morphometric and shape descriptors of intermyofibrillar mitochondria, including Form Factor (FF), Roundness, Aspect Ratio (AR), Feret's diameters (Feret), Circularity (Circ), Perimeter (Perim), and Area in male M- Bcl6 WT and KO mice. (E) Quantification of genes involved in mitochondrial quality control processes in quadriceps of 10-week-old male M- Bcl6 WT and KO mice. mRNA expression levels are shown relative to housekeeping gene β-actin and are expressed as fold change compared to M- Bcl6 WT as assessed by RT-qPCR. n = 6–9 per group. (F) Immunoblot detection (right) and corresponding protein quantification (left) of OPA1, VDAC1, Mfn2, and Drp1 in quadriceps of male M- Bcl6 WT and M–KO mice. n = 5 per group. (G) Mitochondrial DNA (mtDNA) copy number calculated as a ratio of mitochondrial encoded genes ( Cox2 or 16S ) to nuclear encoded genes ( CycA or HK2 ) as measured by RT-qPCR from quadriceps of 12-week-old M- Bcl6 WT and M- Bcl6 KO mice. n = 7 per group. (H) Quantification of mitochondrial biogenesis genes in quadriceps of 10-week-old male M- Bcl6 WT and KO mice. mRNA expression levels are shown relative to housekeeping gene β-actin and are expressed as fold change compared to M- Bcl6 WT assessed by RT-qPCR. n = 8–9 per group. (I) Immunoblot detection (left) and corresponding protein quantification (right) of representative subunits of the mitochondrial oxidative phosphorylation (mtOxPhos) from quadriceps of male M- Bcl6 WT and M- Bcl6 KO mice. n = 5 per group. ( J ) Mitochondrial respiration of male and female M- Bcl6 WT and M- Bcl6 KO mice performed by the addition of pyruvate and malate (PM) in permeabilized muscle fibers across a range of ADP concentrations, and succinate (Succ) in the presence of creatine. n = 7–11 per group. Data in C, D, E, F, H, and I were analyzed by unpaired two-tailed Student's t -test (∗p < 0.05). Data in J was analyzed with two-way ANOVA, and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data are presented as mean ± SEM (with individual data points).

Techniques: Expressing, Control, Quantitative RT-PCR, Western Blot, Phospho-proteomics, Two Tailed Test

AAV-KD of Bcl6 in adult mice reduces mitochondrial content and respiration . (A) Representative images of succinate dehydrogenase (SDH) histochemistry of TA muscle 4 weeks post AAV-injection. Scale bar = 50 μm. (B) Quantification of SDH staining from TA muscle injected with either AAV-scrmb or AAV-sh BCL6 knockdown. n = 10 per group. (C) Mitochondrial respiration of GAS muscle injected with either AAV-scrmb or AAV-sh BCL6 knockdown. GM: respiration rate driven by the addition of Glutamate and Malate. ADP: respiration rate driven by the subsequent addition of ADP. n = 10 per group. Data in B were analyzed with paired two-tailed Student's t -test (∗∗p < 0.01). Data in C were analyzed with two-way ANOVA, and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data are presented as mean ± SEM (with individual data points).

Journal: Molecular Metabolism

Article Title: BCL6 regulates skeletal muscle mass and mitochondrial bioenergetics

doi: 10.1016/j.molmet.2026.102367

Figure Lengend Snippet: AAV-KD of Bcl6 in adult mice reduces mitochondrial content and respiration . (A) Representative images of succinate dehydrogenase (SDH) histochemistry of TA muscle 4 weeks post AAV-injection. Scale bar = 50 μm. (B) Quantification of SDH staining from TA muscle injected with either AAV-scrmb or AAV-sh BCL6 knockdown. n = 10 per group. (C) Mitochondrial respiration of GAS muscle injected with either AAV-scrmb or AAV-sh BCL6 knockdown. GM: respiration rate driven by the addition of Glutamate and Malate. ADP: respiration rate driven by the subsequent addition of ADP. n = 10 per group. Data in B were analyzed with paired two-tailed Student's t -test (∗∗p < 0.01). Data in C were analyzed with two-way ANOVA, and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data are presented as mean ± SEM (with individual data points).

Techniques: Injection, Staining, Knockdown, Two Tailed Test

Bcl6 KO mice display blunted adaptation to endurance training . (A) Schematic representation of experimental design created with BioRender.com . (B–C) Distance (B) and time (C) achieved by untrained and trained M- Bcl6 WT and M- Bcl6 KO mice during a treadmill exhaustion test. (D–F) Quantification of mitochondrial biogenesis genes (D- Tfam , E- Tfb1m and F- Tfb1m ) in the quadriceps of untrained and trained male M- Bcl6 WT and KO mice. mRNA expression levels are shown relative to housekeeping gene 18s and are expressed as fold change compared to M- Bcl6 WT assessed by RT-qPCR. Data were analyzed by two-way ANOVAs followed by a Fisher's LSD post hoc test (∗p < 0.05). Data are presented as mean ± SEM (with individual data points).

Journal: Molecular Metabolism

Article Title: BCL6 regulates skeletal muscle mass and mitochondrial bioenergetics

doi: 10.1016/j.molmet.2026.102367

Figure Lengend Snippet: Bcl6 KO mice display blunted adaptation to endurance training . (A) Schematic representation of experimental design created with BioRender.com . (B–C) Distance (B) and time (C) achieved by untrained and trained M- Bcl6 WT and M- Bcl6 KO mice during a treadmill exhaustion test. (D–F) Quantification of mitochondrial biogenesis genes (D- Tfam , E- Tfb1m and F- Tfb1m ) in the quadriceps of untrained and trained male M- Bcl6 WT and KO mice. mRNA expression levels are shown relative to housekeeping gene 18s and are expressed as fold change compared to M- Bcl6 WT assessed by RT-qPCR. Data were analyzed by two-way ANOVAs followed by a Fisher's LSD post hoc test (∗p < 0.05). Data are presented as mean ± SEM (with individual data points).

Techniques: Expressing, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Leveraging the BAF chromatin remodeling complex for targeted transcriptional rewiring in cancer

doi: 10.64898/2026.03.30.715217

Figure Lengend Snippet: a , Schematic of chemically induced strategy for recruitment of the BAF complex (via SMARCA2/4) to BCL6. The BAF complex at BCL6 loci leads to eviction of nucleosomes and BCL6, increased chromatin accessibility, and de-repression of pro-apoptotic BCL6 target genes. b , Overview of chemoproteomic competition assay for SMARCA ligand characterization. SMARCA ligand with linker was immobilized on NHS-activated Sepharose beads and outcompeted with the SMARCA ligand (top). Each data point in the volcano plot (bottom) represents a dose-response score for a protein. The p-value indicates the significance of a dose-response according to the CurveCurator statistics pipeline, and the log2 fold change indicates the effect size of the dose-dependent depletion. Binding curves for highlighted BAF complex members in orange are shown in Extended Data Figure 1a. Protein with missing/imputed values in their dose-response curves were filtered out. Hits: Log 2 FC ≥ |1| and -log 10 P value ≥ 2 (dotted lines). c , Chemical structures of BCL6-SMARCA bifunctional compounds, TRIP1, TRIP1-neg1 (SMARCA non-binding), and TRIP1-neg2 (BCL6 non-binding). d , CellTiter-Glo cell viability assays in diffuse large B cell lymphoma (DLBCL) cell lines. KARPAS-422 (top) and SU-DHL4 (bottom) cells were treated with a dilution series of TRIP1 or negative controls for 72 hours. Viability with compound treatment is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. e , BCL6 transcriptional reporter de-repression. KARPAS-422 cells expressing a BCL6 transcriptional reporter (top) were treated with a dilution series of TRIP1 or negative controls for 24 hours. Reporter activity is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. f , CaspaseGlo 3/7 apoptosis detection. KARPAS-422 cells were treated with a dilution series of compounds for 16 hours. Caspase 3/7 activity is normalized to DMSO vehicle control; data represent mean ± SD, n = 3 independent replicates. g , SMARCA4 target engagement. Monoclonal HEK293T cells expressing endogenously tagged SMARCA4-HiBiT were pre-treated with a dilution series of TRIP1 or the SMARCA ligand for 2 hours, followed by co-treatment with 0.1 µM ACBI1 for 8 hours. SMARCA4-HiBiT levels are shown as a remaining fraction relative to DMSO vehicle control (no ACBI1 treatment). D max represents the fraction of SMARCA4 remaining when ACBI1 is co-treated with DMSO. Gray bar indicates early cytotoxic/pro-apoptotic concentrations of TRIP1. Data represent mean ± SD, n = 6 independent replicates.

Article Snippet: The following antibodies were used: IgG rabbit isotype control (Cell Signaling Technology, CST3900), BCL6 (Cell Signaling Technology, CST49360), SMARCA4 (Cell Signaling Technology, CST49360), H3K27ac (Cell Signaling Technology, CST8173), RNA Pol II Ser2 phospho (Cell Signaling Technology, CST13499), RNA Pol II Ser5 phospho (Cell Signaling Technology, CST13523).

Techniques: Competitive Binding Assay, Binding Assay, Control, Expressing, Activity Assay, Drug discovery

Journal: bioRxiv

Article Title: Leveraging the BAF chromatin remodeling complex for targeted transcriptional rewiring in cancer

doi: 10.64898/2026.03.30.715217

Figure Lengend Snippet: a , In vitro TR-FRET ternary complex formation assay. The interaction between biotinylated SMARCA4 bromodomain (100 nM) and FITC-labeled BCL6 BTB domain (500 nM) was measured with increasing concentrations of TRIP1. The biotinylated SMARCA4 bromodomain was captured with streptavidin-terbium cryptate donor (SA-Tb, 2 nM), and TR-FRET signal was monitored upon complex formation. The resulting TR-FRET ratio was normalized to the DMSO vehicle control; data represent mean ± SD, n = 2 independent replicates. b , TR-FRET competition assay. The interaction between biotinylated SMARCA2/4 bromodomains (100 nM) and FITC-labeled BCL6 BTB domain (500 nM) was measured in the presence of 100 nM TRIP1 and increasing concentrations of BCL6 or SMARCA ligand. The resulting TR-FRET ratio was background subtracted and normalized to the DMSO vehicle control; data represent mean ± SD, n = 2 independent replicates. c , SplitHalo assay for BCL6-SMARCA2 interaction. Schematic of the in-cell splitHalo assay to probe induced protein-protein interactions (top). A HaloTag enzyme is split into two complementing parts, cpHalo and Hpep3 peptide, which only assemble into a functional self-labeling HaloTag enzyme when actively brought into proximity and supplied with TAMRA dye. HEK293T cells co-expressing BCL6-cpHalo and SMARCA2(BD)-Hpep3 were treated with a dilution series of TRIP1 or negative controls and incubated with compound and the covalent HaloTag dye TAMRA for 3 hours (bottom). Data is normalized to DMSO vehicle control, which corresponds to the baseline signal upon TAMRA addition; data represent mean ± SD, n = 3 independent replicates. d , SplitHalo competition assay for TRIP1-induced BCL6-SMARCA2 interaction. Schematic of the in-cell splitHalo competition assay to probe the inhibition of induced protein-protein interactions (top). Cells are pre-incubated with excess amounts of protein ligands to saturate binding pockets and prevent or reduce ternary complex formation. HEK293T cells co-expressing BCL6-cpHalo and SMARCA2(BD)-Hpep3 were pre-treated with a dilution series of SMARCA or BCL6 ligand for 30 minutes before adding 1 µM TRIP1 and TAMRA dye for 3 hours (bottom). Data is normalized to TRIP1 with DMSO vehicle control without ligand addition, corresponding to maximum ternary complex formation; data represent mean ± SD, n = 3 independent replicates. e , f , BCL6 transcriptional reporter competition. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO, the BCL6 ligand ( e ), or SMARCA ligand ( f ) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates. g , CaspaseGlo 3/7 apoptosis pre-degradation. KARPAS-422 cells were pre-treated with DMSO or SMARCA degrader (ACBI1) for 8 hours, followed by co-treatment with 1 µM of TRIP1 for 16 hours. Caspase 3/7 activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Techniques: In Vitro, Tube Formation Assay, Labeling, Control, Competitive Binding Assay, Protein-Protein interactions, Functional Assay, Expressing, Incubation, Inhibition, Binding Assay, Activity Assay

Journal: bioRxiv

Article Title: Leveraging the BAF chromatin remodeling complex for targeted transcriptional rewiring in cancer

doi: 10.64898/2026.03.30.715217

Figure Lengend Snippet: a , TRIP1 resistance CRISPR viability screen. KARPAS-422 iCas9 cells were mutagenized with a genome-wide sgRNA library and cultivated in the presence of DMSO or TRIP1 (400 nM) for 33 days. Gene-level enrichment score was calculated as -log10 (P value) × log2 (fold-change). BAF complex members are colored based on involvement in different sub-complexes or modules. n = 2 independent replicates. b , Competitive growth assay in KARPAS-422 cells. Control KARPAS-422 iCas9 cells expressing AAVS1-targeting sgRNA and BFP were mixed with KARPAS-422 iCas9 cells expressing BAF subunit-targeting sgRNAs and mCherry. Cell mixtures were treated with DMSO, TRIP1 (400 nM), BRD4 recruiting TCIP1 (4 nM), or p300/CBP recruiting TCIP3 (4 nM) and evaluated in regular intervals every 3 or 4 days via flow cytometry. Data were logit-transformed and normalized to 50% at day 0 to enable comparison across conditions. Data represent mean ± SD, n = 3 independent replicates. c , BCL6 transcriptional reporter pre-degradation. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO or BRD7/9 degrader (VZ-185) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Techniques: CRISPR, Genome Wide, Growth Assay, Control, Expressing, Flow Cytometry, Transformation Assay, Comparison, Activity Assay

Journal: bioRxiv

Article Title: Leveraging the BAF chromatin remodeling complex for targeted transcriptional rewiring in cancer

doi: 10.64898/2026.03.30.715217

Figure Lengend Snippet: a , Genes up- and down-regulated after TRIP1 treatment. 3’ RNA fingerprinting in KARPAS-422 cells after 8h (left) or 16h (right) treatment with 2 µM TRIP1. BCL6 target genes involved in apoptosis, B-cell differentiation, and cell-cycle are highlighted in brown/bold. Hits: Log 2 FC ≥ |1| and P adj < 0.05 (dotted lines). b , Gene expression changes (3’ RNA fingerprinting) in KARPAS-422 cells of selected BCL6 target genes after treatment with TRIP1 or negative control compounds at two different time points and concentrations. c , Lollipop plot summarizing gene set enrichment analysis (GSEA) of BCL6 target genes and genes shown to be regulated by BCL6 targeting TCIPs in Extended Data Figure 4d. Normalized enrichment scores are plotted for each condition compared to DMSO. d , LISA analysis of differentially expressed genes (DEGs) after TRIP1 treatment (16h, 0.5 µM). BCL6 (orange) is the top predicted transcriptional regulator of DEGs after TRIP1 treatment. Other transcriptional repressors with a high score are highlighted in brown. For a-d , differential gene expression analysis was performed using DESeq2. Log₂ fold changes were shrunk using the apeglm method. P-values were obtained from the Wald test and adjusted for multiple testing using the Benjamini-Hochberg procedure. Genes with an adjusted p-value (padj) < 0.05 and |log₂FC| > 1 are highlighted as differentially expressed; n= 3 independent replicates. e , Validation of TRIP1-induced expression changes at the protein level. Time-resolved (left) and dose-resolved (right) expression changes of BCL6, ARID3A, FOXO3 and p21 (CDKN1A) in KARPAS-422 cells. Loading controls were performed per gel. Western blots are representative of two independent replicates.

Techniques: Cell Differentiation, Gene Expression, Negative Control, Biomarker Discovery, Expressing, Western Blot

Journal: bioRxiv

Article Title: Leveraging the BAF chromatin remodeling complex for targeted transcriptional rewiring in cancer

doi: 10.64898/2026.03.30.715217

Figure Lengend Snippet: a, CUT&RUN binding profiles at BCL6 peaks. Binding profiles of BCL6 (left) and SMARCA4 (right) at BCL6 peaks for DMSO/TRIP1 (2 µM) treated cells after 4 and 8 hours. BCL6 peaks were called from DMSO-treated KARPAS-422 samples. Profiles were centered on peaks and extended 5 kb upstream and downstream of the peak location. b , Correlation between BCL6 and SMARCA4 (top), BCL6 and H3K27ac (middle), or SMARCA4 and H3K27ac (bottom) signal changes on gene bodies upon TRIP1 (8h, 2 µM) treatment compared to DMSO. Single dots represent hg38 genes. c, Relationship between TRIP1-induced BCL6/SMARCA4 binding change and H3K27 acetylation or SMARCA4 binding. Genes were ranked based on their differential binding of BCL6 (top) and SMARCA4 (bottom) after TRIP1 treatment (8h, 2 µM) and segmented into respective 20% quantiles. SMARCA4 (top) and H3K27ac (middle) changes at genes per BCL6 quintile. H3K27ac change (bottom) at genes segmented by SMARCA4 quintiles. After overall association was assessed by Kruskal-Wallis was successfully, pairwise comparisons were made by Dunn’s post-hoc test with Benjamini-Hochberg FDR correction. d , Relationship of TRIP1-induced gene expression changes with BCL6/SMARCA4 binding changes after 4 and 8 hours of TRIP1 treatment. Pearson correlation coefficient and R 2 were calculated. Dots represent single genes colored by their gene expression change upon TRIP1 treatment (2 µM after 16 hours vs. DMSO). Selected differentially expressed genes and BCL6 are highlighted. For b-d , Normalized CUT&RUN signal on gene bodies ±3 kb up and downstream to include regulatory regions was calculated for TRIP1 and DMSO. The scores were subtracted to calculate differential binding. e , CUT&RUN binding profiles at BCL6 peaks. Binding profiles of BCL6 at BCL6 peaks after short, low-dose TRIP1 (1 µM) treatment up to 2h. BCL6 peaks were called from DMSO-treated KARPAS-422 CUT&RUN samples. Profiles were centered on peaks and extended 5 kb upstream and downstream of the peak location. f-h , Genome tracks of the ARID3A gene locus. Time-resolved ( f ) BCL6 and SMARCA4 or ( g ) RNA Pol II serine 2/5 phosphorylation signal is computed along the gene locus to infer transcriptional dynamics. For g , Below, RNA-seq reads are mapped to the gene locus. h , BAF ATPase-dependent BCL6 eviction after 1 hour of DMSO or TRIP1 (1 µM) co-treatment with BRM-014 (1 µM). BCL6signal is computed along the ARID3A gene locus. Exon position and genome location are indicated below the genome tracks. All CUT&RUN data is from two merged independent replicates. i , CaspaseGlo 3/7 apoptosis ATPase pre-inhibition. KARPAS-422 cells were pre-treated with DMSO or SMARCA2/4 ATPase inhibitor (BRM-014) for 8 hours, followed by co-treatment with 1 µM of TRIP1 for 16 hours. Caspase 3/7 activity is normalized to DMSO control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates. j , BCL6 transcriptional reporter ATPase pre-inhibition. KARPAS-422 cells expressing a BCL6 transcriptional reporter were pre-treated with DMSO or SMARCA2/4 ATPase inhibitor (BRM-014) for 8 hours, followed by co-treatment with 0.5 µM of TRIP1 for 24 hours. Reporter activity is normalized to DMSO vehicle control without TRIP1 co-treatment; data represent mean ± SD, n = 6 independent replicates.

Techniques: Binding Assay, Gene Expression, Phospho-proteomics, RNA Sequencing, Inhibition, Activity Assay, Control, Expressing

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