Exploring SMC motor proteins and how to stop them

Genetic information encoded in the DNA sequences is maintained, replicated, and transcribed across the tree of life. Organisms evolved multiple hierarchical layers of chromosome organization which ensure that DNA can be contained but also processed within individual cells. This thesis focuses on SMC (Structural Maintenance of Chromosomes)...

Single-molecule visualization of twin-supercoiled domains generated during transcription

Transcription-coupled supercoiling of DNA is a key factor in chromosome compaction and the regulation of genetic processes in all domains of life. It has become common knowledge that, during transcription, the DNA-dependent RNA polymerase (RNAP) induces positive supercoiling ahead of it (downstream) and negative supercoils in its wake ...

Author Correction: Testing pseudotopological and nontopological models for SMC-driven DNA loop extrusion against roadblock-traversal experiments (Scientific Reports, (2023), 13, 1, (8100), 10.1038/s41598-023-35359-2)

Correction to: Scientific Reports, published online 19 May 2023 The original version of this Article contained an error in Figure 1b-1, where the fore- and background order of the strands “DNA” (in black) and “Brn1 Kleisin” (in green), were switched. The original Figure 1 and accompanying legend appear below. (Figure presented.) Description...

Looping the Genome with SMC Complexes

SMC (structural maintenance of chromosomes) protein complexes are an evolutionarily conserved family of motor proteins that hold sister chromatids together and fold genomes throughout the cell cycle by DNA loop extrusion. These complexes play a key role in a variety of functions in the packaging and regulation of chromosomes, and they have...

Dynamin A as a one-component division machinery for synthetic cells

Membrane abscission, the final cut of the last connection between emerging daughter cells, is an indispensable event in the last stage of cell division and in other cellular processes such as endocytosis, virus release or bacterial sporulation. However, its mechanism remains poorly understood, impeding its application as a cell-division...

Testing pseudotopological and nontopological models for SMC-driven DNA loop extrusion against roadblock-traversal experiments

DNA loop extrusion by structural-maintenance-of-chromosome (SMC) complexes has emerged as a primary organizing principle for chromosomes. The mechanism by which SMC motor proteins extrude DNA loops is still unresolved and much debated. The ring-like structure of SMC complexes prompted multiple models where the extruded DNA is topologically or...

CTCF is a DNA-tension-dependent barrier to cohesin-mediated loop extrusion

In eukaryotes, genomic DNA is extruded into loops by cohesin¹. By restraining this process, the DNA-binding protein CCCTC-binding factor (CTCF) generates topologically associating domains (TADs)^2,3 that have important roles in gene regulation and recombination during development and disease^1,4–7. How CTCF...

Dynamic ParB–DNA interactions initiate and maintain a partition condensate for bacterial chromosome segregation

In most bacteria, chromosome segregation is driven by the ParABS system where the CTPase protein ParB loads at the parS site to trigger the formation of a large partition complex. Here, we present in vitro studies of the partition complex for Bacillus subtilis ParB, using single-molecule fluorescence microscopy and AFM imaging to show that...

SMC complexes can traverse physical roadblocks bigger than their ring size

Ring-shaped structural maintenance of chromosomes (SMC) complexes like condensin and cohesin extrude loops of DNA. It remains, however, unclear how they can extrude DNA loops in chromatin that is bound with proteins. Here, we use in vitro single-molecule visualization to show that nucleosomes, RNA polymerase, and dCas9 pose virtually no...

Spatially coherent diffusion of human RNA Pol II depends on transcriptional state rather than chromatin motion

Gene transcription by RNA polymerase II (RNAPol II) is a tightly regulated process in the genomic, temporal, and spatial context. Recently, we have shown that chromatin exhibits spatially coherently moving regions over the entire nucleus, which is enhanced by transcription. Yet, it remains unclear how the mobility of RNA Pol II molecules is...

ParB proteins can bypass DNA-bound roadblocks via dimer-dimer recruitment

The ParABS system is essential for prokaryotic chromosome segregation. After loading at parS on the genome, ParB (partition protein B) proteins rapidly redistribute to distances of ~15 kilobases from the loading site. It has remained puzzling how this large-distance spreading can occur along DNA loaded with hundreds of proteins. Using in...

Navigating the crowd: visualizing coordination between genome dynamics, structure, and transcription

The eukaryotic genome is hierarchically structured yet highly dynamic. Regulating transcription in this environment demands a high level of coordination to permit many proteins to interact with chromatin fiber at appropriate sites in a timely manner. We describe how recent advances in quantitative imaging techniques overcome caveats of...

Coupling chromatin structure and dynamics by live super-resolution imaging

Chromatin conformation regulates gene expression and thus, constant remodeling of chromatin structure is essential to guarantee proper cell function. To gain insight into the spatiotemporal organization of the genome, we use high-density photoactivated localization microscopy and deep learning to obtain temporally resolved super-resolution...

Dynamics as a cause for the nanoscale organization of the genome

Chromatin ‘blobs’ were recently identified by live super-resolution imaging of labeled nucleosomes as pervasive but fleeting structural entities. However, the mechanisms leading to the formation of these blobs and their functional implications are unknown. We explore here whether causal relationships exist between parameters that characterize...