Biology subjects

Pajon, C.

Publications and source records attributed to Pajon, C..

3 recordsLinked to original sources

bioRxiv2026

Loss of nsp14-exonuclease activity impairs the replication, proofreading, fitness and pathogenesis of SARS-CoV-2

Coronaviruses (CoVs) replicate their RNA genomes with increased fidelity than other RNA viruses, a mechanism mediated by the proofreading and recombination activities of the exoribonuclease domain of replicase nonstructural protein 14 (nsp14-ExoN). Both murine hepatitis virus (MHV) and SARS-CoV tolerate nsp14-ExoN loss-of-function mutations (ExoN-) (D90A and E92A) but have impaired replication fidelity and pathogenesis yet identical substitutions in MERS-CoV and SARS-CoV-2 have been reported to be lethal. Here, we report a saturation mutagenesis approach facilitating the recovery and analysis of several constellations of SARS-CoV-2 nsp14 ExoN-inactivating, loss-of-function substitutions, including the canonical D90A and E92A. Biochemical assays with purified WT or ExoN- nsp10-14 fusion proteins confirmed that active site substitutions abolished ExoN activity (ExoN-). SARS-CoV-2 ExoN- viruses had impaired replication, RNA synthesis, and recombination, as well as decreased replication fidelity and loss of fitness in vitro. ExoN- viruses were significantly attenuated for replication in human primary airway epithelial cells and were attenuated for replication and pathogenesis in WT mice as well as the highly susceptible K18 transgenic mice. In the absence of interferon signaling in vivo, SARS-CoV and SARS-CoV-2 ExoN- viral replication could be nearly fully restored. These results demonstrate that SARS-CoV-2 ExoN- are viable but highly impaired for replication, fitness, and fidelity in vitro as well as innate immune antagonism and pathogenesis in vivo. Collectively, our results solidify the multiple critical roles for nsp14-ExoN across CoV genera and establish new approaches for rescue and analysis of loss-of-function substitutions for studies of CoV replication, pathogenesis, and evolution.

microbiology↗

bioRxiv2025

A neutralizing human antibody induces movement of the HCoV-229E receptor binding domain

HCoV-229E is an endemic Alphacoronavirus that typically causes common cold-like disease in most healthy adults, but can also cause severe respiratory disease in the very young and the elderly. Although the virus was discovered over sixty years ago and undergoes continuous antigenic drift, remarkably little is known about the humoral immune response to HCoV-229E infection. Here we report the isolation of two receptor binding domain-targeting neutralizing human antibodies raised in response to natural HCoV-229E infection. One of these, DH1533, potently neutralizes HCoV-229E, binds to spike with sub-nanomolar affinity and prevents the association between the RBD and the host cell receptor aminopeptidase N. Structural characterization of this antibody bound to HCoV-229E spike delineated a neutralization-sensitive epitope on the RBD and revealed that DH1533 induces conformational flexibility in neighboring RBDs, reminiscent of the "up-and-down" kinetics observed in the related Betacoronavirus spikes. These findings provide insight into the humoral immune response to HCoV-229E infection and will serve as a guide for the design of future therapeutic interventions.

immunology↗

bioRxiv2022

Interactions between metabolism and growth can determine the co-existence of Staphylococcus aureus and Pseudomonas aeruginosa

Most bacteria exist and interact within polymicrobial communities. These interactions produce unique compounds, increased virulence and augmented antibiotic resistance. One community associated with negative healthcare outcomes consists of Pseudomonas aeruginosa and Staphylococcus aureus. When co-cultured, virulence factors secreted by P. aeruginosa reduce metabolism and growth in S. aureus. When grown in vitro this allows P. aeruginosa to drive S. aureus towards extinction. However, when found in vivo, both species can co-exist. Previous work has noted that this may due to altered gene expression or mutations. However, little is known about how the growth environment could influence co-existence of both species. Using a combination of mathematical modeling and experimentation, we show that changes to bacterial growth and metabolism caused by differences in the growth environment can determine final population composition. We found that changing the carbon source in growth medium affects the ratio of ATP to growth rate for both species, a metric we call absolute growth. We found that as a growth environment increases absolute growth for one species, that species will dominate the co-culture. This is due to interactions between growth, metabolism and metabolism altering virulence factors produced by P. aeruginosa. Finally, we show that the relationship between absolute growth and final population composition can be perturbed by altering the spatial structure in the community. Our results demonstrate that differences in growth environment can account for conflicting observations regarding the co-existence of these bacterial species in the literature, and may offer a novel mechanism to manipulate polymicrobial populations.

microbiology↗