Biology subjects

Girvan, P.

Publications and source records attributed to Girvan, P..

3 recordsLinked to original sources

bioRxiv2026

INO80 rapidly shuttles nucleosomes between chromatin barriers

ATP-dependent chromatin remodelers establish nucleosome organization, but how they move nucleosomes over extended distances and respond to neighboring chromatin remains unclear. Here, we combine correlative optical tweezers and fluorescence microscopy, and single-molecule FRET to visualize human INO80 activity on naked and chromatinized DNA. hINO80 undergoes free one-dimensional diffusion along DNA with multiple, nucleotide-regulated, diffusive states. Upon engaging a nucleosome, hINO80 drives rapid, processive nucleosome sliding over thousands of base pairs in bidirectional bursts. Neighboring nucleosomes prevent passage and redirect translocation, confining mobile nucleosomes to repeated movement between chromatin boundaries. Single-molecule FRET reveals transitions consistent with INO80 switching between opposing nucleosome-binding orientations, while fluorescence stoichiometry and mass photometry show that two hINO80 complexes can simultaneously occupy one nucleosome. These findings identify non-exclusive mechanisms for directional reversal and reveal how long-range hINO80 translocation is converted into boundary-constrained nucleosome repositioning, providing a dynamic framework for understanding nucleosome organization within chromatin.

biophysics↗

bioRxiv2024

Herpes simplex virus type 1 origin binding protein UL9 tethers and loops origin- and non-origin-DNA intra- and intermolecularly

Herpesviruses are ubiquitous human pathogens, which are the causative agent of mild to severe symptoms ranging from cold sore to nasopharyngeal carcinoma. Even though replication of the linear dsDNA genome has been studied for decades, we still lack a complete molecular understanding of its mechanism. It has been proposed, but never shown directly, that the HSV-1 origin binding protein UL9 binds two closely located binding sites within the oriS origin sequence, thereby mediating origin looping, which in turn facilitates replication initiation. Here, we used an array of single-molecule approaches to test this long-standing hypothesis directly. Surprisingly, the data show that UL9 does not loop oriS efficiently. However, we demonstrate that UL9 can form large DNA loops at non-origin sequences very efficiently, as well as tether two oriS DNA molecules intermolecularly. Contrary to the origin bending hypothesis, our findings indicate that UL9 does not loop oriS DNA, but rather may play an alternative role in replication initiation, such as tethering two separate molecules to facilitate recombination.

microbiology↗

bioRxiv2024

Tetramethylrhodamine self-quenching is a probe of conformational change on the scale of 15 - 25 Angstrom

Tetramethylrhodamine (TMR) is a fluorescent dye whose self-quenching has been used as a probe of multiple biological phenomena. We determine the distance-dependence of self-quenching and place bounds on the timescale of TMR dissociation. Our results validate fluorescence self-quenching as an alternative to FRET and enable future assays to be designed with confidence.

biophysics↗