DNA shape analysis has demonstrated the potential to reveal structure-based mechanisms of protein-DNA binding. However, information about influence chemical modification is limited. Cytosine methylation, most frequent modification, represents addition a methyl group at major groove edge cytosine base. In mammalian genomes, methylation frequently occurs CpG dinucleotides. changing signature C/G base pairs, can affect structure. Since original discovery efforts have been made understand its...

Protein-DNA binding is a fundamental component of gene regulatory processes, but it still not completely understood how proteins recognize their target sites in the genome. Besides hydrogen bonding major groove (base readout), minor-groove geometry using positively charged amino acids (shape readout). The underlying mechanism DNA shape readout involves correlation between width and electrostatic potential (EP). To probe this biophysical effect directly, rather than as an indirect measure for...

DNA-binding proteins play important roles in various cellular processes, but the mechanisms by which recognize genomic target sites remain incompletely understood. Functional groups at edges of base pairs (bp) exposed DNA grooves represent physicochemical signatures. As these signatures enable to form specific contacts between protein residues and bp, their study can provide mechanistic insights into protein-DNA binding. Existing experimental methods, such as X-ray crystallography, reveal...

Genome-wide binding profiles of estrogen receptor (ER) and FOXA1 reflect cancer state in ER + breast cancer. However, routine profiling tumor transcription factor (TF) is impractical the clinic. Here, we show that plasma cell-free DNA (cfDNA) contains high-resolution for Enrichment TF footprints reflects strength originating tissue. We defined pure vivo signatures using xenografts, which can distinguish xenografts with distinct states. Furthermore, state-specific ER-binding partition human...

We demonstrate here that the α subunit C-terminal domain of Escherichia coli RNA polymerase (αCTD) recognizes upstream promoter (UP) DNA element via its characteristic minor groove shape and electrostatic potential. In two compositionally distinct crystallized assemblies, a pair αCTD subunits bind in tandem to UP consensus A-tract is 6 bp length (A6-tract), each with their arginine 265 guanidinium group inserted into groove. The A6-tract significantly narrowed these crystal structures, as...

Cell-free DNA (cfDNA) has the potential to enable non-invasive detection of disease states and progression. Beyond its sequence, cfDNA also represents nucleosomal landscape cell(s)-of-origin captures dynamics epigenome. In this review, we highlight emergence epigenomic methods that assess beyond scope mutant tumour genotyping. Detection mutations is gold standard for sequencing in clinical oncology. However, limitations inherent mutation targeting cfDNA, possibilities uncovering molecular...

Many anecdotal observations exist of a regulatory effect DNA methylation on gene expression (Dantas Machado et al., 2015). However, in general, the underlying mechanisms this are poorl...

The MspI-HpaII digestion patterns of Gryllotalpa fossor (Scudder) DNA indicated the methylation internal cytosine in sequence 5′ -CCGG-3′. amount 5-methylcytosine (5mC) was estimated by HPLC analysis to be about 0.6–0.8%, which constitutes approximately 3% total genome. This is first example a non-mammalian animal with appreciable levels 5mC.Key words: insect genome, 5-methylcytosine, methylation.

Abstract Enhancers harbor binding motifs that recruit transcription factors (TFs) for gene activation. While cooperative of TFs at enhancers is known to be critical transcriptional activation a handful developmental enhancers, the extent TF cooperativity genome-wide unknown. Here, we couple high-resolution nuclease footprinting with single-molecule methylation profiling characterize active in Drosophila genome. Enrichment short MNase-protected DNA segments indicates majority two or more...

Abstract How transcription factors (TF) selectively occupy a minute subset of their binding sites from sizeable pool putative in large mammalian genomes remains an important unanswered question. In part, nucleosomes help by creating formidable barriers to TF binding. concentration itself plays crucial role the competition between TFs and nucleosomes. case nuclear receptors, ligand adds another layer complexity. Estrogen receptor alpha (ER) is classic example where its main estradiol (E2) can...

e21198 Background: Despite revolutionizing cancer therapy, immune checkpoint inhibitors (ICI) still do not benefit a significant proportion of patients. The risks associated with ICI-related adverse events, mixed performance PD-L1 staining in predicting treatment response and its high cost, present clinical need for more precise methods to define disease states the context ICI treatment. is thought depend on phenotype tumor response, especially functional state CD8+ T cells microenvironment...

Enhancers harbor binding motifs that recruit transcription factors (TFs) for gene activation. While cooperative of TFs at enhancers is known to be critical transcriptional activation a handful developmental enhancers, the extent TF cooperativity genome-wide unknown. Here, we couple high-resolution nuclease footprinting with single-molecule methylation profiling characterize active in Drosophila genome. Enrichment short MNase-protected DNA segments indicates majority two or more sites, and...

Here, we present a pipeline to map states of protein-binding DNA in vivo. Our infers as well quantifies cooperative binding. Using dual-enzyme single-molecule footprinting (dSMF) data, show how our workflow identifies binding at an enhancer Drosophila S2 cells. Data from cells lacking endogenous methylation are prerequisite for this pipeline. For complete details on the use and execution protocol, please refer Rao et al. (2021) Krebs (2017).

Abstract Cell-free DNA (cfDNA) contains a composite map of the epigenomes its cells-of-origin. Tissue-specific transcription factor (TF) binding inferred from cfDNA could enable us to track disease states in humans minimally invasive manner. Here, by enriching for short fragments, we directly TF footprints at single sites plasma. We show that enrichment plasma reflects strength tissue-of-origin. Based on this principle, were able identify subset genome-wide selected TFs leave TF-specific...