Biology subjects

Curtis, B.

Publications and source records attributed to Curtis, B..

4 recordsLinked to original sources

bioRxiv2023

Influenza A virus NS1 effector domain is required for PA-X mediated host shutoff

Many viruses inhibit general host gene expression to limit innate immune responses and gain preferential access to the cellular translational apparatus for their own protein synthesis. This process is known as host shutoff. Influenza A viruses (IAVs) encode two host shutoff proteins: nonstructural protein 1 (NS1) and polymerase acidic X (PA-X). NS1 inhibits host nuclear pre-messenger RNA maturation and export, and PA-X is an endoribonuclease that preferentially cleaves host spliced nuclear and cytoplasmic messenger RNAs. Emerging evidence suggests that in circulating human IAVs NS1 and PA-X co-evolve to ensure optimal magnitude of general host shutoff without compromising viral replication that relies on host cell metabolism. However, the functional interplay between PA-X and NS1 remains unexplored. In this study, we sought to determine if NS1 function has a direct effect on PA-X activity by analyzing host shutoff in A549 cells infected with wild type or mutant IAVs with NS1 effector domain deletion. This was done using conventional quantitative reverse transcription polymerase chain reaction techniques and direct RNA sequencing using nanopore technology. Our previous research on the molecular mechanisms of PA-X function identified two prominent features of IAV infected cells: nuclear accumulation of cytoplasmic poly(A) binding protein (PABPC1) and increase in nuclear poly(A) RNA abundance relative to the cytoplasm. Here we demonstrate that NS1 effector domain function augments PA-X host shutoff and is necessary for nuclear PABPC1 accumulation. By contrast, nuclear poly(A) RNA accumulation is not dependent on either NS1 or PA-X mediated host shutoff and is accompanied by nuclear retention of viral transcripts. Our study demonstrates for the first time that NS1 and PA-X effects on host gene expression are not simply additive and that these factors functionally interact in mediating host shutoff. IMPORTANCERespiratory viruses including influenza A virus continue to cause annual epidemics with high morbidity and mortality due to limited effectiveness of vaccines and antiviral drugs. Among the strategies evolved by viruses to evade immune responses is host shutoff - a general blockade of host messenger RNA and protein synthesis. Disabling influenza A virus host shutoff is being explored in live attenuated vaccine development as an attractive strategy for increasing their effectiveness by boosting antiviral responses. Influenza A virus encodes two proteins that function in host shutoff: the non-structural protein 1 (NS1) and the polymerase acidic X (PA-X). We and others have characterised some of the NS1 and PA-X mechanisms of action and the additive effects that these viral proteins may have in ensuring the blockade of host gene expression. In this work we examined whether NS1 and PA-X functionally interact and discovered that NS1 is required for PA-X to function effectively. This work significantly advances our understanding of influenza A virus host shutoff and identifies new potential targets for therapeutic interventions against influenza and further informs development of improved live attenuated vaccines.

microbiology↗

bioRxiv2023

Evolutionarily related host and microbial pathways regulate fat desaturation

Fatty acid desaturation is central to metazoan lipid metabolism and provides building blocks of membrane lipids and precursors of diverse signaling molecules. Nutritional conditions and associated microbiota regulate desaturase expression1-4, but the underlying mechanisms have remained unclear. Here, we show that endogenous and microbiota-dependent small molecule signals promote lipid desaturation via the nuclear receptor NHR-49/PPAR in C. elegans. Untargeted metabolomics of a {beta}-oxidation mutant, acdh-11, in which expression of the stearoyl-CoA desaturase FAT-7/SCD1 is constitutively increased, revealed accumulation of a {beta}- cyclopropyl fatty acid, becyp#1, that potently activates fat-7 expression via NHR-49. Biosynthesis of becyp#1 is strictly dependent on expression of cyclopropane synthase by associated bacteria, e.g., E. coli. Screening for structurally related endogenous metabolites revealed a {beta}-methyl fatty acid, bemeth#1, whose activity mimics that of microbiota-dependent becyp#1, but is derived from a methyltransferase, fcmt-1, that is conserved across Nematoda and likely originates from bacterial cyclopropane synthase via ancient horizontal gene transfer. Activation of fat-7 expression by these structurally similar metabolites is controlled by distinct mechanisms, as microbiota-dependent becyp#1 is metabolized by a dedicated {beta}-oxidation pathway, while the endogenous bemeth#1 is metabolized via -oxidation. Collectively, we demonstrate that evolutionarily related biosynthetic pathways in metazoan host and associated microbiota converge on NHR-49/PPAR to regulate fat desaturation.

biochemistry↗

bioRxiv2023

Extreme mitochondrial reduction in a novel group of free-living metamonads.

Metamonads are a diverse group of heterotrophic microbial eukaryotes adapted to living in hypoxic environments. All metamonads but one harbour metabolically altered mitochondrion-related organelles (MROs) with reduced functions relative to aerobic mitochondria, however the degree of reduction varies markedly over the metamonad tree. To further investigate metamonad MRO diversity, we generated high quality draft genomes, transcriptomes, and predicted proteomes for five recently discovered free-living metamonads. Phylogenomic analyses placed these organisms in a group we informally named the BaSk (Barthelonids+Skoliomonads) clade, which emerges as a deeply branching sister group to the Fornicata, a metamonad phylum that includes parasitic and free-living flagellates. Extensive bioinformatic analyses of the manually curated gene models showed that these organisms are predicted to have extremely reduced MRO proteomes in comparison to other free-living metamonads. Loss of the mitochondrial iron-sulfur cluster (ISC) assembly system in some organisms in this group appears to be linked to the acquisition in their common ancestral lineage of a SUF-like minimal system (SMS) Fe/S cluster pathway through lateral gene transfer (LGT). One of the isolates, Skoliomonas litria, appears to have undergone further mitochondrial reduction having lost all other known MRO pathways. No proteins were confidently assigned to the predicted MRO proteome of this organism suggesting that the organelle has been lost. The extreme mitochondrial reduction observed within this free-living anaerobic protistan clade is unprecedented and demonstrates that mitochondrial functions, under some conditions, may be completely lost even in free-living organisms.

genomics↗

bioRxiv2021

Chromosome-scale assemblies of Acanthamoeba castellanii genomes provide insights into Legionella pneumophila infection-related chromatin re-organization

The unicellular amoeba Acanthamoeba castellanii is ubiquitous in aquatic environments, where it preys on bacteria. The organism also hosts bacterial endosymbionts, some of which are parasitic, including human pathogens such as Chlamydia and Legionella spp. Here we report complete, high quality genome sequences for two extensively studied A. castellanii strains, Neff and C3. Combining long- and short-read data with Hi-C, we generated near chromosome-level assemblies for both strains with 90% of the genome contained in 29 scaffolds for the Neff strain and 31 for the C3 strain. Comparative genomics revealed strain-specific functional enrichment, most notably genes related to signal transduction in the C3 strain, and to viral replication in Neff. Furthermore, we characterized the spatial organization of the A. castellanii genome and showed that it is reorganized during infection by Legionella pneumophila. Infection-dependent chromatin loops were found to be enriched in genes for signal transduction and phosphorylation processes. In genomic regions where chromatin organization changed during Legionella infection, we found functional enrichment for genes associated with metabolism, organelle assembly, and cytoskeleton organization, suggesting that changes in chromosomal folding are associated with host cell biology during infection.

genomics↗