Biology subjects

Smith, M. M.

Publications and source records attributed to Smith, M. M..

3 recordsLinked to original sources

bioRxiv2025

The human transcription factor occupancy landscape viewed using high-resolution in situ base-conversion strand-specific single-molecule chromatin accessibility mapping

Chromatin accessibility profiling is a key tool for mapping the location of cis-regulatory elements (cREs) in the genome and tracking chromatin state dynamics during development, in response to various external and internal stimuli, and in disease contexts. Single-molecule footprinting (SMF) methods that rely on the labeling of individual accessible DNA bases have emerged in recent years as a powerful chromatin accessibility mapping approach, as they provide not just an average readout of accessibility over a given genomic position but also the distribution of accessibility states within a population at the level of individual original DNA molecules. However, methylation-based SMF approaches have often been limited either in their resolution or in labeling readout accuracy. We developed a deaminase-based high-resolution strand-specific single molecule footprinting approach that uses highly active sequence context-independent endogenous methylation-insensitive double-strand DNA (dsDNA) deaminases (CseDa01 and LbDa02 C[->]U/T-ssSMF), which convert accessible cytosines into uracils (which are converted to thymine after PCR amplification). We demonstrate the application of the method to mapping single-molecule accessibility states in human cell lines in both a short and a long-read format, and quantifying the occupancy states of individual transcription factors (TFs) as well as TF co-accessibility and strand-specific accessibility patterns.

genomics↗

bioRxiv2021

Exogenous pigments shield microorganisms from spaceflight-induced changes

Research has indicated that pigments commonly produced by microorganisms may be protective against the environmental stresses inherent to spaceflight. However, few studies have directly tested the protective capabilities of microbial pigments applied externally as shielding materials. In this study, liquid cultures of Bacillus subtilis were shielded by various pigment solutions, and solid media cultures of Bacillus subtilis were co-inoculated with the highly pigmented microorganisms Aspergillus niger and Neurospora crassa. These experiments were conducted in a compact, automated payload aboard the International Space Station (ISS) interior for 30 days. Post-flight phenotypic analyses of liquid cultures showed that solutions of carotenoid pigments were effective at minimizing detrimental effects of spaceflight. Elevated growth rate was observed for solid cultures, and distinct morphology changes were identified in both liquid and solid samples and quantified as markers of spaceflight-induced stress. These findings collectively progress our understanding of microbial pigments for the development of space-related applications.

microbiology↗

bioRxiv2017

Super-Resolution Microscopy Reveals Structural Mechanisms Driving the Nanoarchitecture of a Viral Chromatin Tether

By tethering their circular genomes (episomes) to host chromatin, DNA tumor viruses ensure retention and segregation of their genetic material during cell divisions. Despite functional genetic and crystallographic studies, there is little information addressing the three-dimensional structure of these tethers in cells, issues critical for understanding persistent infection by these viruses. Here, we have applied direct stochastic optical reconstruction microscopy (dSTORM) to establish the nanoarchitecture of tethers within cells latently infected with the oncogenic human pathogen, Kaposi's sarcoma-associated herpesvirus (KSHV). Each KSHV tether comprises a series of homodimers of the latency-associated nuclear antigen (LANA) that bind with their C-termini to the tandem array of episomal terminal repeats (TRs) and with their N-termini to host chromatin. Super-resolution imaging revealed that individual KSHV tethers possess similar overall dimensions and, in aggregate, fold to occupy the volume of a prolate ellipsoid. Using plasmids with increasing numbers of TRs, we found that tethers display polymer power-law scaling behavior with a scaling exponent characteristic of active chromatin. For plasmids containing a two-TR tether, we determined the size, separation, and relative orientation of two distinct clusters of bound LANA, each corresponding to a single TR. From these data, we have generated a three-dimensional model of the episomal half of the tether that integrates and extends previously established findings from epi-fluorescent, crystallographic, and epigenetic approaches. Our findings also validate the use of dSTORM in establishing novel structural insights into the physical basis of molecular connections linking host and pathogen genomes.

microbiology↗