Biology subjects

Fedorenko, T.

Publications and source records attributed to Fedorenko, T..

5 recordsLinked to original sources

bioRxiv2025

Extracellular activity of a bacterial protease associated with reduced phage infectivity

To defend against bacteriophage (phage) infection, bacteria have developed various defense systems, dozens of which were discovered and mechanistically studied recently. To date, almost all defense systems whose mechanisms were deciphered were shown to operate within the bacterial cell. Here we describe a secreted protease from the Actinobacterium Salinispora mooreana which, when expressed heterologously in Streptomyces coelicolor, reduces titers of two taxonomically related Siphoviridae phages. Antiphage effects were maintained when concentrated supernatant from S. coelicolor expressing the Salinispora protease was added externally to phage-containing medium, even in the absence of bacterial cells, supporting an extracellular mechanism. We further show that phages can escape the antiphage effect of the Salinispora protease by mutating a tail-associated protein. The antiphage effect is associated with an increased proportion of phage particles devoid of DNA. Our data suggest antiphage activity of a secreted bacterial protease.

microbiology↗

bioRxiv2022

Discovery of phage determinants that confer sensitivity to bacterial immune systems

Over the past few years, numerous anti-phage defense systems have been discovered in bacteria. While the mechanism of defense for some of these systems is understood, a major unanswered question is how these systems sense phage infection. To systematically address this question, we isolated 192 phage mutants that escape 19 different defense systems. In many cases, these escaper phages were mutated in the gene sensed by the defense system, enabling us to map the phage determinants that confer sensitivity to bacterial immunity. Our data identify specificity determinants of diverse retron systems and reveal phage-encoded triggers for multiple abortive infection systems. We find general themes in phage sensing and demonstrate that mechanistically diverse systems have converged to sense either the core replication machinery of the phage, phage structural components, or host takeover mechanisms. Combining our data with previous findings, we formulate key principles on how bacterial immune systems sense phage invaders.

microbiology↗

bioRxiv2022

Cryo-EM structure of the RADAR supramolecular anti-phage defense complex

RADAR is a two-protein bacterial defense system which was reported to defend against phage by editing messenger RNA. Here we determine cryo-EM structures of the RADAR defense complex, revealing RdrA as a heptameric, two-layered AAA+ ATPase and RdrB as a dodecameric, hollow complex with twelve surface-exposed deaminase active sites. RdrA and RdrB join to form a giant assembly up to 10 MDa, with RdrA docked as a funnel over the RdrB active site. Surprisingly, our structures reveal a RdrB active site that targets mononucleotides, not RNA. We show that RdrB catalyzes ATP-to-ITP conversion in vitro and induces the accumulation of inosine mononucleotides during phage infection in vivo, limiting phage replication. Our results define ATP mononucleotide deamination as a determinant of RADAR immunity and reveal supramolecular assembly of a nucleotide-modifying machine as a novel mechanism of anti-phage defense.

biochemistry↗

bioRxiv2021

Antiviral defense via nucleotide depletion in bacteria

DNA viruses and retroviruses need to consume large quantities of deoxynucleotides (dNTPs) when replicating within infected cells. The human antiviral factor SAMHD1 takes advantage of this vulnerability in the viral life cycle, and inhibits viral replication by degrading dNTPs into their constituent deoxynucleosides and inorganic phosphate. In this study we report that bacteria employ a similar strategy to defend against phage infection. We found a family of defensive dCTP deaminase proteins that, in response to phage infection, convert dCTP into deoxy-uracil nucleotides. A second family of phage resistance genes encode dGTPase enzymes, which degrade dGTP into phosphate-free deoxy-guanosine (dG) and are distant homologs of the human SAMHD1. Our results show that the defensive proteins completely eliminate the specific deoxynucleotide (either dCTP or dGTP) from the nucleotide pool during phage infection, thus starving the phage of an essential DNA building block and halting its replication. Both defensive genes are found in a diverse set of bacterial species and are specifically enriched in Vibrio genomes. Our study demonstrates that manipulation of the deoxynucleotide pool is a potent antiviral strategy shared by both prokaryotes and eukaryotes.

microbiology↗

bioRxiv2020

Bacterial retrons function in anti-phage defense

Retrons are bacterial genetic elements comprised of a reverse transcriptase (RT) and a non-coding RNA. The RT uses the non-coding RNA as a template, generating a chimeric RNA/DNA molecule in which the RNA and DNA components are covalently linked. Although retrons were discovered three decades ago, their function remained unknown. In this study we report that retrons function as anti-phage defense systems. The defensive unit is composed of three components: the RT, the non-coding RNA, and an effector protein. Retron-containing systems are abundant in genomic "defense islands", suggesting a role for most retrons in phage resistance. By cloning multiple retron systems into a retron-less Escherichia coli strain, we show that these systems confer defense against a broad range of phages, with different retrons defending against different phages. Focusing on a single retron, Ec48, we show evidence that it is a "guardian" of RecBCD, a complex with central anti-phage functions in the bacterial cell. Inhibition of RecBCD by dedicated phage proteins activates the retron, leading to abortive infection and cell death. Thus, the Ec48 retron forms a second line of defense that is triggered if the first lines of defense have collapsed. Our results expose a new family of anti-phage defense systems abundant in bacteria.

microbiology↗