Identification of methylation-sensitive human transcription factors using meSMiLE-seq

. 2024 Nov 12 ; () : . [epub] 20241112

Status PubMed-not-MEDLINE Jazyk angličtina Země Spojené státy americké Médium electronic

Typ dokumentu časopisecké články, preprinty

Perzistentní odkaz   https://www.medvik.cz/link/pmid39605503

Grantová podpora
R01 HG013328 NHGRI NIH HHS - United States
P30 CA008748 NCI NIH HHS - United States
P30 AR070549 NIAMS NIH HHS - United States
U24 HG013078 NHGRI NIH HHS - United States
R01 AI173314 NIAID NIH HHS - United States
R01 AR073228 NIAMS NIH HHS - United States
R21 HG012258 NHGRI NIH HHS - United States

Transcription factors (TFs) are key players in eukaryotic gene regulation, but the DNA binding specificity of many TFs remains unknown. Here, we assayed 284 mostly poorly characterized, putative human TFs using selective microfluidics-based ligand enrichment followed by sequencing (SMiLE-seq), revealing 72 new DNA binding motifs. To investigate whether some of the 158 TFs for which we did not find motifs preferably bind epigenetically modified DNA (i.e. methylated CG dinucleotides), we developed methylation-sensitive SMiLE-seq (meSMiLE-seq). This microfluidic assay simultaneously probes the affinity of a protein to methylated and unmethylated DNA, augmenting the capabilities of the original method to infer methylation-aware binding sites. We assayed 114 TFs with meSMiLE-seq and identified DNA-binding models for 48 proteins, including the known methylation-sensitive binding modes for POU5F1 and RFX5. For 11 TFs, binding to methylated DNA was preferred or resulted in the discovery of alternative, methylation-dependent motifs (e.g. PRDM13), while aversion towards methylated sequences was found for 13 TFs (e.g. USF3). Finally, we uncovered a potential role for ZHX2 as a putative binder of Z-DNA, a left-handed helical DNA structure which is adopted more frequently upon CpG methylation. Altogether, our study significantly expands the human TF codebook by identifying DNA binding motifs for 98 TFs, while providing a versatile platform to quantitatively assay the impact of DNA modifications on TF binding.

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