Biology subjects

Pinckert, M. L.

Publications and source records attributed to Pinckert, M. L..

3 recordsLinked to original sources

bioRxiv2023

comrades-OO: An Object-Oriented R Package for Comprehensive Analysis of RNA Structure Generated using RNA crosslinking experiments

SummaryRNA (Ribonucleic Acid) molecules have secondary and tertiary structures in vivo which play a crucial role in cellular processes such as the regulation of gene expression, RNA processing, and localization. The ability to investigate these structures will enhance our understanding of their function and contribute to the diagnosis and treatment of diseases caused by RNA dysregulation. However, there are no mature pipelines or packages for processing and analysing complex in vivo RNA structural data. Here, we present COMRADES Object-Oriented (comradesOO), a novel software package for the comprehensive analysis of data derived from the COMRADES (Crosslinking of Matched RNA and Deep Sequencing) method. comradesOO offers a comprehensive pipeline from raw sequencing reads to the identification of RNA structural features. It includes read processing and alignment, clustering of duplexes, data exploration, folding and comparisons of RNA structures. comradesOO also enables comparisons between conditions, the identification of inter-RNA interactions, and the incorporation of reactivity data to improve structure prediction. Availability and ImplementationcomradesOO is freely available to non-commercial users and implemented in R, with the source code and documentation accessible at [https://CRAN.R-project.org/package=comradesOO]. The software is supported on Linux, macOS, and Windows platforms. Contactjlp76@cam.ac.uk Supplementary informationSupplementary data are available at Bioinformatics online.

bioinformatics↗

bioRxiv2020

Patterns of within-host genetic diversity in SARS-CoV-2

Monitoring the spread of SARS-CoV-2 and reconstructing transmission chains has become a major public health focus for many governments around the world. The modest mutation rate and rapid transmission of SARS-CoV-2 prevents the reconstruction of transmission chains from consensus genome sequences, but within-host genetic diversity could theoretically help identify close contacts. Here we describe the patterns of within-host diversity in 1,181 SARS-CoV-2 samples sequenced to high depth in duplicate. 95% of samples show within-host mutations at detectable allele frequencies. Analyses of the mutational spectra revealed strong strand asymmetries suggestive of damage or RNA editing of the plus strand, rather than replication errors, dominating the accumulation of mutations during the SARS-CoV-2 pandemic. Within and between host diversity show strong purifying selection, particularly against nonsense mutations. Recurrent within-host mutations, many of which coincide with known phylogenetic homoplasies, display a spectrum and patterns of purifying selection more suggestive of mutational hotspots than recombination or convergent evolution. While allele frequencies suggest that most samples result from infection by a single lineage, we identify multiple putative examples of co-infection. Integrating these results into an epidemiological inference framework, we find that while sharing of within-host variants between samples could help the reconstruction of transmission chains, mutational hotspots and rare cases of superinfection can confound these analyses.

genomics↗

bioRxiv2019

Ebselen inhibits mycobacterial ESX-1 secretion through modification of EccA1

The type VII secretion system ESX-1 mediates virulence in Mycobacterium tuberculosis and Mycobacterium marinum. We find that in M. marinum, the synthetic organoselenium compound ebselen inhibits secretion of ESAT-6, a major ESX-1 substrate. We find that ebselen inhibits the in vitro activity of the ESX-1 AAA+ ATPase EccA1, which potentiates ESX-1 substrate secretion and function. Ebselen modifies a cysteine in its N-terminal tetratricopeptide repeat domain that is required for EccA1s in vitro ATPase activity. Surprisingly, mutational analyses show this this cysteine is not required for ESX-1 secretion or ebselens activity, showing that ebselen inhibits ESX-1 secretion independently of inhibiting EccA1 activity in vitro. While the mechanism by which ebselen inhibits ESX-1 secretion remains elusive, we show that it attenuates ESX-1-mediated damage of M. marinum-containing macrophage phagosomes and inhibits intramacrophage growth. Extending our studies to M. tuberculosis, we find that ebselen inhibits ESX-1 secretion and phagosomal membrane damage in this organism. This work provides insight into EccA1 biology. Ebselen is an orally active drug in clinical trials for other conditions and this work suggests its potential in tuberculosis therapy.

microbiology↗