Figure 1: Integrative genomics approach to study cooperativity and antagonism between the multiple layers that regulate gene expression.

Figure 1: Integrative genomics approach to study cooperativity and antagonism between the multiple layers that regulate gene expression.

Figure 2: Single-molecule footprinting enables us to measure co-occupancy of transcription factors in vivo to understand cooperativity mechanisms.

Figure 2: Single-molecule footprinting enables us to measure co-occupancy of transcription factors in vivo to understand cooperativity mechanisms.

The Krebs group combines single-cell and single-molecule genomics with large-scale genome engineering to understand fundamental mechanisms for controlling gene expression.

Previous and current research

About 7% of the three billion bases contained in our genomes encode cis-regulatory elements, which control the activity of genes. Transcription factors interpret this genetic information to create the >250 different cellular identities required to form a multicellular organism. Expression of a gene is controlled by the collective action of multiple transcription factors, which recruit cofactors that alter chromatin, reshape genome organisation, and initiate transcription. The aim of our research is to understand how these multiple layers of regulatory information, and the factors that interpret them, integrate to precisely define gene expression levels (Fig. 1). How do Transcription factors cooperate? What is the influence of epigenetics on their action? How do they communicate signals across large genomic distances?

We recently developed single-molecule footprinting (SMF), a methodology to quantify protein–DNA contacts at the level of single DNA molecules in vivo (Fig. 2). SMF overcomes some of the limitations of bulk and single-cell genomics assays to quantify the patterns of occupancy of transcription factors at regulatory regions (Fig. 2) (Krebs, Trends in Genetics 2021). Analysis of the binding patterns at promoters with molecular resolution has revealed the mechanisms regulating the dynamics of the early regulatory steps of transcription activation (Krebs et al., Mol Cell 2017). Genome-wide analysis of the molecular co-occupancy of transcription factors has revealed principles ruling their combinatorial action at regulatory regions (Fig. 2) (Sönmezer et al., Mol Cell 2021).

Future projects and goals

Our group will develop experimental and computational strategies to understand the regulatory genome. We attempt to measure how multiple regulators interact at the molecular level at regulatory elements to identify cooperative and antagonistic relationships between them (Fig. 1). Our goal is to perform perturbation experiments at a scale and a resolution that allows us to understand the general principles governing the action of transcriptional regulators.

Transcription factor cooperativity mechanisms

Cooperativity between transcription factors is widespread at regulatory elements (Sönmezer et al., Mol Cell 2021), but the underlying mechanisms are mostly unknown. We will leverage natural genetic variation and large collections of synthetic variants of cis-regulatory regions to generate controlled genotype-to-phenotype associations. Quantitative modelling of the data will enable us to decipher how transcription factor cooperativity is encoded at regulatory regions.

Epigenetic control of transcription

Epigenetic modifications and transcription factors mutually influence each other to control genes during development. We will develop methods to measure the effect of epigenetic and chromatin modifications on the activity of transcription factors on individual DNA molecules. We will use chemical and genetic perturbation to establish the directionality of the interactions between the genetic and epigenetic regulation of transcription.