Research

DNA Organization, Dynamics & Repair

Folding chromosomes by DNA loop extrusion

Cells organize and compact the long and flexible DNA molecules within chromosomes to precisely control gene expression patterns, to support DNA repair processes and DNA recombination events, and to faithfully segregate chromosomes during cell division. What determines chromosome superstructure and how it impacts upon genome expression and genome maintenance are major open questions in biology. We aim to elucidate fundamental principles of chromosome folding by studying microbial model organisms including the Gram-positive bacterium Bacillus subtilis and budding yeast Saccharomyces cerevisiae. 

SMC proteins & DNA loop extrusion

Dynamic SMC architecture

Our work focuses on two widely distributed factors in chromosome organization: SMC protein complexes and ParABS systems. SMC complexes form multi-subunit ATPases with a characteristic ring topology. They organize DNA as genome linkers by clamping together pairs of chromosomal DNA segments. To uncover how they bring together the “right” pairs of DNA double helices, we investigate how SMC complexes interact with DNA and chromosomes. The emerging view is that DNA loop extrusion by SMC DNA motors plays the key role in the remarkable process of chromosome organization. In bacteria, SMC complexes load onto chromosomes at defined entry sites — parS sites — and then translocate onto flanking DNA helping to shape entire bacterial chromosomes. We want to understand how local SMC loading and subsequent DNA translocation brings about global chromosome folding.

ParABS & cellular regulation

ParB CTPase self-loading at a parS site.

ParABS systems promote chromosome segregation and plasmid maintenance in many bacteria and some archaea. It also supports additional regulatory functions in the cell. ParABS systems work by ParB protein binding to centromeric parS sequences to form large nucleoprotein complexes that act in concert with specific ATPases, ParA and Smc, to partition plasmid and chromosome copies. We have recently discovered that ParB proteins are enzymes — the first known CTP hydrolases — which form DNA sliding clamps that self-load onto parS DNA. We are particularly interested how ParB CTP binding and hydrolysis act together with ParA and Smc ATPases to promote chromosome organization and segregation in bacteria.

Our approach

Knowledge of the architecture and structure of macromolecular assemblies is often a prerequisite for a basic mechanistic understanding. We integrate genetics, molecular & cell biology and biochemistry with structural biology approaches to reveal the molecular basis for protein function and cellular activity.

We investigate protein-DNA complexes and DNA organization in vitro using an array of biophysical techniques and structural biology and in vivo for example by ChIP-Seq, Hi-C, directed or random mutagenesis and site-specific cross-linking. Bacillus subtilis develops natural competence under starvation and can readily be genetically manipulated, supporting easy and high-throughput allelic replacements.

New focus

In relatively new lines of investigation, we are focusing on the roles of SMC and SMC-like proteins (Rad50, RecN and the Smc5/6 complex) in the maintenance of DNA integrity in bacteria and eukaryotes.