Biology subjects

Elfenbein, J. R.

Publications and source records attributed to Elfenbein, J. R..

3 recordsLinked to original sources

bioRxiv2020

Sulfate import in Salmonella Typhimurium impacts bacterial aggregation and the neutrophil respiratory burst

During enteric salmonellosis, neutrophil generated reactive oxygen species alter the gut microenvironment favoring survival of Salmonella Typhimurium. While the type-3 secretion system-1 (T3SS-1) and flagellar motility are potent Salmonella Typhimurium agonists of the neutrophil respiratory burst in vitro, neither of these pathways alone are responsible for stimulation of a maximal respiratory burst. In order to identify Salmonella Typhimurium genes that impact the magnitude of the neutrophil respiratory burst, we performed a two-step screen of defined mutant libraries in co-culture with neutrophils. We first screened Salmonella Typhimurium mutants lacking defined genomic regions, followed by the individual mutants mapping to genomic regions under selection. Mutants in four genes, STM1696 (sapF), STM2201 (yeiE), STM2112 (wcaD), and STM2441 (cysA), induced an attenuated respiratory burst. We linked the altered respiratory burst to reduced T3SS-1 expression and/or altered flagellar motility for two mutants ({Delta}STM1696 and {Delta}STM2201). The {Delta}STM2441 mutant, defective for sulfate transport, formed aggregates in minimal media and adhered to surfaces in rich media, suggesting a role for sulfur homeostasis in regulation of aggregation/adherence. We linked the aggregation/adherence phenotype of the {Delta}STM2441 mutant to biofilm-associated protein A and flagellins and hypothesize that aggregation caused the observed reduction in the magnitude of the neutrophil respiratory burst. Our data demonstrate that Salmonella Typhimurium has numerous mechanisms to limit the magnitude of the neutrophil respiratory burst. These data further inform our understanding of how Salmonella may alter neutrophil antimicrobial defenses.

microbiology↗

bioRxiv2020

Bacterial retrons encode tripartite toxin/antitoxin systems

Retrons are genetic retroelements, commonly found in bacterial genomes and recently repurposed as genome editing tools. Their encoded reverse transcriptase (RT) produces a multi-copy single-stranded DNA (msDNA). Despite our understanding of their complex biosynthesis, the function of msDNAs and therefore, the physiological role of retrons has remained elusive. We establish that the retron-Sen2 in Salmonella Typhimurium encodes a toxin, which we have renamed as RcaT (Retron cold-anaerobic Toxin). RcaT is activated when msDNA biosynthesis is perturbed and its toxicity is higher at ambient temperatures or during anaerobiosis. The RT and msDNA form together the antitoxin unit, with the RT binding RcaT, and the msDNA enabling the antitoxin activity. Using another E. coli retron, we establish that this toxin/antitoxin function is conserved, and that RT-toxin interactions are cognate. Altogether, retrons constitute a novel family of tripartite toxin/antitoxin systems.

microbiology↗

bioRxiv2020

Phage proteins block and trigger retron toxin/antitoxin systems

Bacteria carry dozens of Toxin/Antitoxin systems (TAs) in their chromosomes. Upon growth, the antitoxin is co-expressed and neutralizes the toxin. TAs can be activated and inhibit growth, but when and how this occurs has largely remained enigmatic, hindering our understanding of their physiological roles. We developed TIC/TAC (Toxin Inhibition/Activation Conjugation), a high-throughput reverse genetics approach, to systematically identify molecular blockers and triggers of TAs. By applying TIC/TAC to a tripartite TA, the retron-Sen2 of Salmonella Typhimurium, we have identified multiple blockers and triggers of phage origin. We demonstrate that diverse phage functionalities are sensed by the DNA-part of the antitoxin and ultimately activate the retron toxin. Phage-origin proteins can circumvent activation by directly blocking the toxin. Some identified triggers and blockers also act on an E. coli retron-TA, Eco9. We propose that retron-TAs act as abortive-infection anti-phage defense systems, and delineate mechanistic principles by which phages trigger or block them.

microbiology↗