Biology subjects

Barzaghi, G.

Publications and source records attributed to Barzaghi, G..

4 recordsLinked to original sources

bioRxiv2025

FootprintCharter: unsupervised detection and quantification of footprints in single molecule footprinting data

SummarySingle molecule footprinting profiles the heterogeneity of TF occupancy at cis-regulatory elements across cell populations at unprecedented resolution. The single molecule nature of the data in principle allows for observing the footprint of individual transcription factors and nucleosomes. However, we currently lack algorithms to quantify these occupancy patterns of chromatin binding factors in an automated way and without prior assumptions on their genomic location. Here we present FootprintCharter, an unsupervised tool to detect and quantify footprints for transcription factors (TFs) and nucleosomes from single molecule footprinting data. FootprintCharter allows for the quantification of complex molecular states such as positioning of unphased nucleosomes and combinatorial co-binding of multiple TFs. Availability and implementationFootprintCharter is freely available on Bioconductor with version 2.1.6 of https://bioconductor.org/packages/SingleMoleculeFootprinting with the functions FootprintCharter, PlotFootprints and Plot_FootprintCharter_SM.

bioinformatics↗

bioRxiv2025

Cumulative TF binding and H3K27 Acetylation drive enhancer activation frequency

In eukaryotes, transcription factors (TFs) must continuously compete with nucleosomes to access their binding sites, leading to cell-to-cell variability in chromatin accessibility at regulatory regions. Although critical to understand enhancer function in transcription, the mechanisms that define how frequently an enhancer is active in a cell population remain unclear. Here, we used single molecule footprinting to quantify the frequency at which chromatin is accessible at enhancers in response to TF perturbations, and changes of their chromatin environment. We find that individually most TFs open chromatin in a small fraction of the cells, and that cumulative TF binding controls enhancers activation frequency. Moreover, testing the functionality of hundreds of enhancers when inserted at an ectopic genomic location identifies H3K27Ac as a chromatin mark required for their full activation. Our data support a model where enhancer activation frequency depends on the cumulative function of multiple TFs, enhanced by activating chromatin marks.

molecular biology↗

bioRxiv2023

Identification of transcription factor co-binding patterns with non-negative matrix factorization

Transcription factor (TF) binding to DNA is critical to transcription regulation. Although the binding properties of numerous individual TFs are well-documented, a more detailed comprehension of how TFs interact cooperatively with DNA, forming either complex or co-binding to the same region, is required. Indeed, the combinatorial binding of TFs is essential to cell differentiation, development, and response to external stimuli. We present COBIND, a novel method based on non-negative matrix factorization (NMF) to identify TF co-binding patterns automatically. COBIND applies NMF to one-hot encoded regions flanking known TF binding sites (TFBSs) to pinpoint enriched DNA patterns at fixed distances. We applied COBIND to 8,293 TFBS datasets from UniBind for 404 TFs in seven species. The method uncovered already established co-binding patterns (e.g., between POU5F1 and SOX2 or SOX17) and new co-binding configurations not yet reported in the literature and inferred through motif similarity and protein-protein interaction knowledge. Our extensive analyses across species revealed that 84% of the studied TFs share a co-binding motif with other TFs from the same structural family. The co-binding patterns captured by COBIND are likely functionally relevant as they harbor higher evolutionarily conservation than isolated TFBSs. Open chromatin data from matching human cell lines further supported the co-binding predictions. Finally, we used single-molecule footprinting data from mouse embryonic stem cells to confirm that the co-binding events captured by COBIND were likely occurring on the same DNA molecules.

bioinformatics↗

bioRxiv2022

Single molecule multi-omics reveals context-dependent regulation of enhancers by DNA methylation

Enhancers are cis-regulatory elements that control the establishment of cell identities during development. In mammals, enhancer activation is tightly coupled with local DNA demethylation. Yet, whether this epigenetic remodelling is necessary for enhancer activation is unknown. Here, we developed a single molecule multi-omics approach to measure chromatin accessibility and transcription factor binding as a function of the presence of methylation on the same DNA molecules. We leveraged natural epigenetic heterogeneity at active enhancers to test the impact of DNA methylation on their activity in multiple cell lineages. While reduction of DNA methylation appears dispensable for the activity of most enhancers, we identify a class of cell-type-specific enhancers where presence of DNA methylation antagonises the binding of transcription factors. Genetic perturbations reveal that chromatin accessibility and transcription factor binding are a direct function of active demethylation at these loci. Thus, in addition to safeguarding the genome from spurious activation, DNA methylation directly controls transcription factor occupancy at cell-type-specific active enhancers.

genomics↗