Journal: Nature Communications

Article Title: Tumor-specific lncRNA IGF1R-AS1 trans-regulates chromatin interactions associated with oncogenic MYC signaling

doi: 10.1038/s41467-026-70814-4

Figure Lengend Snippet: a Heatmap of differentially expressed annotated ( n = 595) and unannotated ( n = 191) lncRNAs across benign prostate tissue (Normal, n = 52), localized prostate cancer (PCa, n = 500), and metastatic prostate cancer (Mets, n = 101) samples. RNA-seq data for Normal and PCa samples were obtained from the TCGA cohort, while Mets samples were sourced from the SU2C study. Rows represent lncRNAs, with annotated lncRNAs in the upper panel and unannotated lncRNAs in the lower panel. Columns represent individual samples grouped by disease state. Expression values are Z-score normalized across samples. Representative lncRNAs with established roles in prostate cancer progression (e.g., CTBP1-AS , MALAT1 , and SChLAP1 ) are highlighted. b Super-enhancer-associated lncRNAs and protein-coding genes in VCaP cells based on ranked ordered H3K27ac ChIP-seq signal. c Identification of super-enhancer-associated coding genes and lncRNA transcripts elevated during prostate cancer progression. d Detected IGF1R-AS1 isoforms by short- and long-read sequencing approaches. e Integrated genome view of IGF1R-AS1 and its associated super-enhancer in VCaP cells. The data tracks represent RNA-Seq; ChIP-Seq for H3K4me3, H3K27ac, and RNA Pol II; ChIA-PET for RNA Pol II. f ChIP-qPCR measurement of BRD4 binding at the IGF1R-AS1 -associated super-enhancer region in JQ1-treated VCaP cells and DMSO control cells ( n = 3 biological replicates). Unpaired Student’s t -test, two-sided. p *=0.0161, p **=0.0011, p ****<0.0001. g RNA expression levels of IGF1R-AS1 , IGF1R , and the known super-enhancer target gene MYC measured in VCaP cells treated with JQ1 and DMSO ( n = 3 biological replicates). Unpaired Student’s t -test, two-sided. p ****<0.0001, ns not significant. h SMARCA4 ChIP-seq binding in the IGF1R-AS1 -associated super-enhancer across different prostate cancer cell lines and ATAC-seq signals under SMARCA4 depletion and control VCaP cells at this super-enhancer region. i H3K27ac HiChIP heatmap within the IGF1R-AS1 locus in VCaP cells treated with or without the SWI/SNF PROTAC degrader AU-15330 for 4 h (bin size = 5 kb). Overlaid on the heatmap are ATAC-seq read-density tracks from the same treatment conditions. Gray shading indicates typical- and super-enhancers, while blue shading marks the IGF1R-AS1 promoter. The loop represents a read-supported cis -interaction within the locus, with interaction reads (IR) denoting the strength of this interaction. Data in ( f , g ) shown are mean ± SEM. Source data of this Figure are provided as Source data file.

Article Snippet: Full length and fragments of SMARCA4 and SMARCA1 coding DNA sequences were cloned through PCR amplification from ORF clones of SMARCA4 (RG219258) and SMARCA1 (RC239926) purchased from OriGene and inserted into pCDH-EF1-IRES-eGFP vector. siRNAs targeting IGF1R-AS1 were purchased from Dharmacon. siRNAs targeting SMARCA1 (SR304469) or SMARCA4 (SR321835) were purchased from OriGene. shRNA oligos targeting IGF1R-AS1 were constructed in pLKO-vector (#10878) from Addgene.

Techniques: RNA Sequencing, Expressing, ChIP-sequencing, Sequencing, ChIA Pet Assay, ChIP-qPCR, Binding Assay, Control, RNA Expression, HiChIP

Journal: Nature Communications

Article Title: Tumor-specific lncRNA IGF1R-AS1 trans-regulates chromatin interactions associated with oncogenic MYC signaling

doi: 10.1038/s41467-026-70814-4

Figure Lengend Snippet: a FISH analysis of IGF1R-AS1 in VCaP cells. (Left) IGF1R-AS1 FISH signal (magenta) in wild-type (WT) cells (top) and after siRNA knockdown (bottom). DAPI stains nuclei (blue). Scale bar: 5 μm. (Right) Quantification of IGF1R-AS1 foci distribution in WT cells ( n = 4 biological replicates). Unpaired Student’s t -test, two-sided. p ***=0.0009. b Workflow for detecting the IGF1R-AS1 interactome. c High-confidence interactions between IGF1R-AS1 and chromatin remodeling complex subfamily members are highlighted, with spectral counts indicating interaction strength. Additional interacting proteins are listed in Supplementary Data . d Binding of IGF1R-AS1 to SMARCA1 and SMARCA4 was detected in IGF1R-AS1 RNA pulldown assays. IGF1R sense RNA ( IGF1R-S1 ) and streptavidin beads alone served as negative controls. e RIP-qPCR analysis of IGF1R-AS1 enriched by anti-SMARCA1 and anti-SMARCA4 antibodies in VCaP cells ( n = 3 biological replicates). SChLAP1 , a known SWI/SNF-interacting lncRNA, was included as a positive control. Unpaired Student’s t -test, two-sided. p ****<0.0001. f , g Western blotting analysis of recombinant SMARCA4 proteins and SMARCA1 proteins pulled down by IGF1R-AS1 RNA in vitro binding assay. IGF1R-S1 and streptavidin beads served as negative controls. h The secondary structure model of IGF1R-AS1 is predicted by RNAfold webserver based on minimum free energy algorithm. i Western blotting analysis of SMARCA1 and SMARCA4 pulled down by full-length and truncated IGF1R-AS1 . j , k Schematic representation of Flag-tagged full-length SMARCA4 ( j ) and SMARCA1 ( k ), and its truncations, and RIP-qPCR in VCaP cells showing IGF1R-AS1 enrichment by Flag-tagged full-length SMARCA4 ( j ) and SMARCA1 ( k ), and its truncations ( n = 3 biological replicates). l , m GSEA analysis of RNA-seq data from SMARCA4 knockdown ( l ) and SMARCA1 knockdown ( m ) in VCaP cells. n–q qPCR analysis ( n , o , n = 3 biological replicates) and western blotting analysis ( p , q ) of MYC mRNA and protein levels following SMARCA4 or SMARCA1 depletion in VCaP cells. Unpaired Student’s t -test in ( n and o ), two-sided. p ****<0.0001. r , s VCaP cell proliferation curves following SMARCA4 ( r , n = 3 biological replicates) or SMARCA1 ( s , n = 3 biological replicates) depletion. Unpaired Student’s t -test, two-sided. p **** ( r , siNC vs siSMARCA4-1#, day 5) < 0.0001, p *** ( r , siNC vs siSMARCA4-2#, day 5) = 0.0002, p *** ( s , siNC vs siSMARCA1-1#, day 5) = 0.0002, p *** ( s , siNC vs siSMARCA1-2#, day 5) = 0.0004. Data in ( a , e , j , k , n , o , r , and s) are shown as mean ± SEM. Experiments in ( d , f , g , i , p , and q ) were biologically repeated three times. Source data of this Figure are provided as Source data file.

Article Snippet: Full length and fragments of SMARCA4 and SMARCA1 coding DNA sequences were cloned through PCR amplification from ORF clones of SMARCA4 (RG219258) and SMARCA1 (RC239926) purchased from OriGene and inserted into pCDH-EF1-IRES-eGFP vector. siRNAs targeting IGF1R-AS1 were purchased from Dharmacon. siRNAs targeting SMARCA1 (SR304469) or SMARCA4 (SR321835) were purchased from OriGene. shRNA oligos targeting IGF1R-AS1 were constructed in pLKO-vector (#10878) from Addgene.

Techniques: Knockdown, Binding Assay, Positive Control, Western Blot, Recombinant, In Vitro, RNA Sequencing

Journal: Nature Communications

Article Title: Tumor-specific lncRNA IGF1R-AS1 trans-regulates chromatin interactions associated with oncogenic MYC signaling

doi: 10.1038/s41467-026-70814-4

Figure Lengend Snippet: a Genome-wide chromatin accessibility changes following IGF1R-AS1 depletion. Heatmap shows ATAC-seq signal intensity (± 1 kb from peak center) for 9244 regions with decreased accessibility (top) and 12808 regions with increased accessibility (bottom). b Overlap of IGF1R-AS1 depletion-induced accessibility changes with SMARCA4 binding sites. 3800 regions with reduced accessibility coincide with SMARCA4 ChIP-seq peaks (right). These peaks are completely lost upon treatment with the SWI/SNF PROTAC degrader AU-15330. c Schematic for constructing an enhancer-promoter (E-P) interaction network. d Top eight E-P interaction subnetworks by node size highlight key regulatory hubs like MYC , with node size and color intensity reflecting centrality within each network. e IGF1R-AS1 predominantly influences the E-P subnetwork of MYC , as indicated by the MYC gene’s highest normalized centrality score. f K-Means clustering of 40 prostate cancer models, including mCRPC organoids, xenografts, and cell lines, based on ATAC-seq signals from the top 10 MYC-connected enhancers. IGF1R-AS1 expression was mainly found in the high MYC enhancer activity group. g Kaplan–Meier (K–M) survival analysis of mCRPC patients ( n = 499) stratified by IGF1R-AS1 network activity score at optimal cutpoints. Cox proportional-hazards regression model (CoxPH) p -value indicates significance of IGF1R-AS1 network activity as a predictor in a penalized spline Cox proportional hazards model. h Kaplan–Meier survival analysis of patients ( n = 499) stratified by both IGF1R-AS1 network activity and MYC pathway activity levels demonstrates poorest survival in IGF1R-AS1 -high/ MYC -high group. p -value indicates significance of IGF1R-AS1 network activity as a predictor in a penalized spline Cox proportional hazards model after stratification by MYC expression.

Article Snippet: Full length and fragments of SMARCA4 and SMARCA1 coding DNA sequences were cloned through PCR amplification from ORF clones of SMARCA4 (RG219258) and SMARCA1 (RC239926) purchased from OriGene and inserted into pCDH-EF1-IRES-eGFP vector. siRNAs targeting IGF1R-AS1 were purchased from Dharmacon. siRNAs targeting SMARCA1 (SR304469) or SMARCA4 (SR321835) were purchased from OriGene. shRNA oligos targeting IGF1R-AS1 were constructed in pLKO-vector (#10878) from Addgene.

Techniques: Genome Wide, Binding Assay, ChIP-sequencing, Expressing, Activity Assay

Journal: Nature Communications

Article Title: Tumor-specific lncRNA IGF1R-AS1 trans-regulates chromatin interactions associated with oncogenic MYC signaling

doi: 10.1038/s41467-026-70814-4

Figure Lengend Snippet: a Multi-panel visualization of the enhancer regions upstream of MYC : (top) Log fold change of interaction frequencies (siNC/siAS1) at different regulatory elements, showing significant change at E3, n = 3 biological replicates; (middle) 3C-qPCR analysis demonstrating loss of chromatin looping between MYC promoter and the E3 enhancer locus upon IGF1R-AS1 depletion, n = 3 biological replicates; (bottom) genome browser tracks displaying ATAC-seq profiles in control and IGF1R-AS1 -depleted cells, SMARCA4 binding by ChIP-seq, and H3K27ac HiChIP interactions connecting the MYC promoter to upstream enhancers. Unpaired Student’s t -test, two-sided. p **=0.0049. ns not significant. b CTCF HiChIP-seq heat maps within the MYC gene locus in VCaP cells treated with DMSO or the SWI/SNF degrader AU-15330 (1 μM) for 4 h (bin size = 50 kb). ChIP-seq read-density tracks for CTCF are overlaid for both conditions. HiChIP loops indicate read-supported cis interactions within the locus. Grey highlights mark key regulatory regions, including enhancers and the MYC promoter. IR interaction reads. Gene annotations are shown below the heat maps, highlighting the MYC locus and adjacent lncRNA genes. c ChIP-qPCR shows decreased binding of CTCF, cohesin (SMC1A), and chromatin remodelers (SMARCA1/A4) at the E3 enhancer after IGF1R-AS1 knockdown. n = 2 biological replicates. d CTCF HiChIP-seq heat maps within the MYC gene locus in VCaP cells in control and after IGF1R-AS1 knockdown. (bin size = 25 kb). Overlaid is ChIP-seq read-density track for CTCF in DMSO from ( b ). Grey highlights mark key regulatory regions. IR interaction reads. e Genome-wide aggregate peak analysis (APA) plots in CTCF HiChIP using square root VC normalization. APA values indicate ratio between signal at CTCF ChIP-seq peaks (middle pixel) and mean signal at bottom left 3 × 3 pixels. f APA values for each chromosome (chr1-22, chrX) in control and IGF1R-AS1 knockdown cells, showing that CTCF signal enrichment goes down in IGF1R-AS1 knockdown cells across all chromosomes ( n = 23, chr1-22, chrX). Paired Wilcoxon signed-rank test, p ****<0.0001. Boxplots are shown as the median (center line), interquartile range (box bounds, 25th to 75th percentile), and whiskers extending to 1.5× the interquartile range, and points beyond the whiskers represent outliers. g RIP-qPCR analysis of IGF1R-AS1 enrichment by anti-CTCF antibody in VCaP cells ( n = 3 biological replicates). Unpaired Student’s t -test, two-sided. p ****<0.0001. h Western blotting analysis of CTCF pulled down by full-length and truncated IGF1R-AS1 . i Co-immunoprecipitation (co-IP) of SMARCA4 with CTCF in VCaP cells upon IGF1R-AS1 depletion. j A model demonstrating IGF1R-AS1 promotes MYC transcription by mediating SWI/SNF-CTCF interaction and facilitating chromatin loops between MYC enhancer and promoter. Data in ( a and g ) are shown as mean ± SEM. Experiments in ( h and i ) were biologically repeated three times. Source data of this Figure are provided as Source data file.

Article Snippet: Full length and fragments of SMARCA4 and SMARCA1 coding DNA sequences were cloned through PCR amplification from ORF clones of SMARCA4 (RG219258) and SMARCA1 (RC239926) purchased from OriGene and inserted into pCDH-EF1-IRES-eGFP vector. siRNAs targeting IGF1R-AS1 were purchased from Dharmacon. siRNAs targeting SMARCA1 (SR304469) or SMARCA4 (SR321835) were purchased from OriGene. shRNA oligos targeting IGF1R-AS1 were constructed in pLKO-vector (#10878) from Addgene.

Techniques: Control, Binding Assay, ChIP-sequencing, HiChIP, ChIP-qPCR, Knockdown, Genome Wide, Western Blot, Immunoprecipitation, Co-Immunoprecipitation Assay