Biology subjects

Garvis, S.

Publications and source records attributed to Garvis, S..

3 recordsLinked to original sources

bioRxiv2025

Structural and Binding Properties of Dps: A Nucleoid Associated Protein

Prokaryotic chromosomal DNA is compacted and protected by nucleoid-associated proteins (NAPs). Among these, the DNA-binding proteins from starved cells (Dps) combine ferroxidase activity with nonspecific DNA binding to form dodecameric assemblies that condense and safeguard the genome under stress. Here, we present an integrated structural and biophysical analysis of Escherichia coli (E. coli) Dps across a pH range from 3 to 11, combining (i) single-particle cryo-EM reconstructions at [~]1.75 [A] resolution, (ii) adaptive Poisson-Boltzmann electrostatic modelling, (iii) electrophoretic mobility shift assays (EMSAs) on a 448 bp DNA fragment, and (iv) cryo-electron tomography (cryo-ET) of Dps-DNA assemblies. Through this analysis, we show that (1) the canonical ferritin-like fold is maintained at all pH values; (2) surface electrostatics shift from highly positive to net negative as the pH increases, modulating DNA affinity (EC50 values of 73.4 nM at the lowest measurement of pH 5, to 815.5 nM at the highest measurement of pH 11); and (3) short-fragment Dps-DNA complexes form amorphous [~]50 nm globular or tubular complexes rather than lattice co-crystals observed on kilobase-length DNA scaffolds. The findings reveal a possible two-stage assembly model, in which protonation-driven nucleation occurs via the Dps N-terminal lysine residues, followed by lattice ordering on long DNA. Our results further define Dps as a pH-responsive nucleoid compaction factor in bacteria.

biophysics↗

bioRxiv2025

FLUID-CELL: Flow-enabled Light and Ultrastructural Imaging Device for Correlative Electron and Light Localization

We present a novel microfluidic flow cell, or Flow-enabled Light and Ultrastructural Imaging Device for Correlative Electron and Light Localization (FLUID-CELL), that connects fluorescence light microscopy (FLM) and cryo-electron microscopy (cryo-EM) for advanced biological imaging and ultrastructural and structural analyses. The design of the FLUID-CELL features a precisely engineered microchannel that maintains native cell culturing conditions, supporting correlation and enabling real-time observation by FLM, as well as subsequent cryo-EM analysis. In this study, this device enabled practical FLM imaging over extended experimental periods, consistent sample handling, and the ability to perform correlative imaging. This capability connects dynamic cellular events imaged by fluorescence light microscopy with high-resolution ultrastructural data collected with cryo-EM. Our dual-modality approach streamlines the workflow and opens new possibilities for investigating the relationship between cellular function and molecular architecture at the nanoscale.

biophysics↗

bioRxiv2021

Loss of SET1/COMPASS methyltransferase activity reduces lifespan and fertility in Caenorhabditis elegans

Post-translational modification of histones, particularly lysine methylation, are thought to play a crucial role in the aging process. Histone 3 lysine 4 (H3K4) methylation, a modification associated with active chromatin, is mediated by a family of SET1 methyltransferases acting within conserved multiprotein complexes known as COMPASS. Previous work in model organisms with partial or complete deletion of COMPASS subunits has yielded conflicting results about the requirement for H3K4 methylation during aging. Here, we reassessed the role of SET1/COMPASS-dependent H3K4 methylation in Caenorhabditis elegans lifespan regulation and fertility by generating set-2(syb2085) mutant animals that express a catalytically inactive form of SET-2, the C. elegans homolog of SET1. We show that animals bearing catalytically inactive SET-2 retain the ability to form COMPASS complexes but have a marked global loss of H3K4 dimethylation and trimethylation. Consistent with previous work, reduced H3K4 methylation was accompanied by loss of fertility; however, in striking contrast to earlier studies, set-2(syb2085) mutants displayed a significantly shortened, not extended, lifespan and had normal intestinal fat stores. Furthermore, other commonly used set-2 mutants were also short-lived, as was a cfp-1 mutant that lacks a non-catalytic SET1/COMPASS component and displays reduced H3K4 methylation. These results challenge previously held views and establish that wild-type H3K4 methylation levels are necessary to achieve a normal lifespan in C. elegans.

genetics↗