biostudies-arrayexpressTamina WeissHomo sapiensdeepToolsBBDuk (BBTools v39.01), Bowtie2 (v.2.4.5https://www.ebi.ac.uk/biostudies/studies/E-MTAB-17186Wnt/β-catenin signaling is commonly understood to activate transcription through pre-accessible regulatory elements. To investigate whether this pathway also directly modulates chromatin states, we performed CUT&RUN in human embryonic kidney 293T (HEK293T) and matched ΔCTNNB1 cells to profile β-catenin-associated chromatin complexes following pathway activation. Cells were pre-treated with the PORCN inhibitor LGK974 (10 nM) for 24 hours and subsequently stimulated with the GSK3 inhibitor CHIR99021 (10 µM) for 30 minutes, 90 minutes, or 4 hours to activate Wnt/β-catenin signaling. Chromatin binding of β-catenin, pan-TCF/LEF (LEF1, TCF7, TCF7L1, and TCF7L2), CBP/p300, HDAC1, H3K27ac, and H3K4me1 was assessed across conditions. An IgG control was included for all experiments. For perturbation experiments, cells were additionally treated with either the CBP/p300 inhibitor A-485 (5 µM) or the HDAC inhibitor sodium butyrate (1 mM). For the rescue experiment, LGK treated ΔCTNNB1 cells were transfected with pcDNA3-S33Yβ-catenin (Addgene#19286) or the empty vector control for 24 hours. The dataset captures signal-dependent and signal-independent changes in factor occupancy at regulatory regions, including conventional Wnt-responsive elements; and latent enhancers where β-catenin functions as a pioneer-like factor together with HDAC1 and CBP, mediating chromatin remodeling and TCF/LEF occupancy.biostudies-arrayexpressSample Collection - Cells were harvested by incubation in Trypsin EDTA 0.25%, and washed with PBS, and subjected to three washes with nuclear extraction buffer (HEPES-KOH pH-8.2 [20 mM], KCl [10 mM], Spermidine [0.5 mM], IGEPAL [0.05%], Glycerol [20%], 1xPMSF).Growth Protocol - HEK293T cells WT and ΔCTNNB1 were cultured in DMEM (41966-029, Gibco – Thermo Fisher Scientific) supplemented with 10% Fetal Bovine Serum (12133C, Sigma-Aldrich) and 10 U/ml Penicillin-Streptomycin (15276355, Gibco - Thermo Fisher Scientific). Cells were passaged by incubation in Trypsin EDTA 0.25% (25-200-056, Thermo Fisher Scientific).Sequencing - Sequenced either as 35 bp pair-end reads on the NextSeq500 using llumina NextSeq 500/550 High Output Kit v2.5 (75 cycles), or as 50 bp paired-end reads on a NextSeq 2000 (Illumina) using the NextSeq 1000/2000 P2 XLEAP-SBS Reagent Kit (100 cycles; Illumina).Nucleic Acid Extraction - For each sample, 250,000 nuclei were bound to 10 µL magnetic ConA beads (Cell Signaling Technology #93569) for 15 minutes at 4 °C with rotation, using the manufacturer’s activation buffer and instructions. Nuclei bound beads were washed with wash buffer (Hepes pH 7.5 [20 mM], NaCl [150 mM], Spermidine [0.5 mM], 1xPMSF) and incubated 5 minutes with wash buffer containing EDTA [0.2 mM], before they were incubated over-night in wash body containing EDTA [0.2 mM], BSA [0.01%], Digitonin [0.025%] and 1:100 antibody. For the panTCF condition, antibodies against LEF1, TCF1, TCF7L1 and TCF7L2 were mixed at equal ratios of 0.8 µL per antibody. For a list of all used antibodies, please refer to table 1. The next day, nuclei were washed five times and incubated 45 minutes at 4 °C in wash buffer containing digitonin [0.025%] and pAG-MNase [0.6 μg/ml]. After additional five washes, nuclei were incubated in 50 µL wash buffer containing CaCl2 [2 mM] for exactly 30 minutes at 4 °C for digestion. Digestion was terminated by adding 3 µL of a 250 mM EDTA/EGTA mix, and the initial fragment release was initiated by the addition of 2 µL NaCl [5 M] per 50 µL sample, followed by incubation at 37 °C for 30 min. This initial release was set aside, and nuclei were incubated in 50 µL urea stop buffer (NaCl [100 mM], EDTA [2 mM], EGTA [2 mM], IGEPAL [0.5%], Urea [8.8 M]) and incubated 30 minutes at RT for a secondary release. Both releases were combined and fragments were purified using phenol chloroform.Sample Treatment - Cells were pre-treated with the PORCN inhibitor LGK974 (10 nM) for 24 hours and subsequently stimulated with the GSK3 inhibitor CHIR99021 (10 µM) for 30 minutes, 90 minutes, or 4 hours to activate Wnt/β-catenin signaling. Chromatin binding of β-catenin, pan-TCF/LEF (LEF1, TCF7, TCF7L1, and TCF7L2), CBP/p300, HDAC1, H3K27ac, and H3K4me1 was assessed across conditions. An IgG control was included for all experiments. For perturbation experiments, cells were additionally treated with either the CBP/p300 inhibitor A-485 (5 µM) or the HDAC inhibitor sodium butyrate (1 mM). For the rescue experiment, LGK treated ΔCTNNB1 cells were transfected with pcDNA3-S33Yβ-catenin (Addgene#19286) or the empty vector control for 24 hours.Library Construction - Library preparation was performed using the KAPA EvoPrep Kit (Roche, #10096039001) according to the manufacturer’s instructions. For end repair and A-tailing, 14 µL of DNA was used in 0.4x reaction volumes. Adapter ligation was carried out in 0.4x reaction volumes using 0.15 µM KAPA Dual-Indexed adapters, followed by a post-ligation cleanup with Mag-Bind TotalPure NGS beads at a 1x ratio. Libraries were resuspended in 10 mM Tris-HCl (pH 8.0) and amplified using 0.5x reaction volumes. A post-amplification cleanup was performed with 1x beads. Libraries were size-selected by electrophoresis on a 2% E-Gel EX agarose gel (Invitrogen, #G402022) for 10 minutes using the E-Gel Power Snap Electrophoresis System. DNA fragments between 150 and 500 bp were excised and purified using the GeneJET Gel Extraction Kit (Thermo Scientific, #K0691). Libraries were quantified using the Qubit High Sensitivity DNA kit (Thermo Scientific, #Q32854), and pooled.OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - For visualization, n = 2 replicates were merged using SAMTools merge and genome-wide coverage tracks as BigWig format were generated using deepTools (bamCoverage) using CPM normalization and -bin10 --extendReads --centerReads --ignoreDublicates.Sequence Alignment - Paired-end sequencing reads were processed using a standard pipeline. Raw FASTQ files were first subjected to adapter trimming using BBDuk (BBTools suite v39.01). Both read pairs were filtered against built in adapters and artifact reference sequences, as well as custom low complexity sequences (poly-AT, poly-G and ployC-stretches). Trimmed reads were then aligned to the human genome (GRCh38/hg38) using Bowtie2 (v.2.4.5) in local alignment mode with the very-sensitive-local preset. Only properly paired reads were retained by disabling mixed and discordant alignments, and dovetail alignments were allowed. The insert size range was restricted to 0-500bp. Unaligned reads were excluded from downstream processing.UnknownTranscriptomicsGenomicsProteomicsKAPA EvoPrep KitIllumina HiSeq 2000CUT&RUNHomo sapiensClaudio CantùTamina WeissfalseCUT&RUN LoV-U time-course profiling of Wnt transcriptional complex in ΔCTNNB1 HEK293T cells compared to wild-typeWnt/β-catenin signaling is commonly understood to activate transcription through pre-accessible regulatory elements. To investigate whether this pathway also directly modulates chromatin states, we performed CUT&RUN in human embryonic kidney 293T (HEK293T) and matched ΔCTNNB1 cells to profile β-catenin-associated chromatin complexes following pathway activation. Cells were pre-treated with the PORCN inhibitor LGK974 (10 nM) for 24 hours and subsequently stimulated with the GSK3 inhibitor CHIR99021 (10 µM) for 30 minutes, 90 minutes, or 4 hours to activate Wnt/β-catenin signaling. Chromatin binding of β-catenin, pan-TCF/LEF (LEF1, TCF7, TCF7L1, and TCF7L2), CBP/p300, HDAC1, H3K27ac, and H3K4me1 was assessed across conditions. An IgG control was included for all experiments. For perturbation experiments, cells were additionally treated with either the CBP/p300 inhibitor A-485 (5 µM) or the HDAC inhibitor sodium butyrate (1 mM). For the rescue experiment, LGK treated ΔCTNNB1 cells were transfected with pcDNA3-S33Yβ-catenin (Addgene#19286) or the empty vector control for 24 hours. The dataset captures signal-dependent and signal-independent changes in factor occupancy at regulatory regions, including conventional Wnt-responsive elements; and latent enhancers where β-catenin functions as a pioneer-like factor together with HDAC1 and CBP, mediating chromatin remodeling and TCF/LEF occupancy.2026-06-27T00:00:00Z2026-06-27T01:00:44.157Z2026-06-18T14:42:07.211ZE-MTAB-17186ERP195332E-MTAB-16975EFO_0002944EFO_0009973EFO_0004170EFO_0003789EFO_0004917EFO_0005518EFO_0003816EFO_0004184EFO_0003969