Biology subjects

Shamovsky, I.

Publications and source records attributed to Shamovsky, I..

6 recordsLinked to original sources

bioRxiv2026

H-NS silences antiviral immunity through 3D chromatin compaction

Bacterial antiviral immunity systems can be toxic to their hosts and must be tightly regulated. Immunity genes are often encoded by AT-rich mobile genetic elements subjected to silencing by nucleoid binding protein H-NS. Using RNA-seq, ChIP-seq, and chromosome 3D reconstruction with Micro-C, we find that H-NS binds and compacts bacterial immunity loci. H-NS deletion reveals anti-phage activity in model Escherichia coli strains generally considered phage-sensitive. Extending this approach to environmental isolates, we show that removal of H-NS silencing enhances defense, allowing bacteria to restrict phages with anti-defense proteins. We also discover Madara, a novel immunity system that complements the co-regulated BREX defense. Our results establish H-NS as a master regulator of bacterial immunity and highlight the importance of expression levels for anti-phage activity in native hosts.

molecular biology↗

bioRxiv2026

OLD sentinel: an abortive tRNase surveys phage replication and DNA defects in RecBCD-compromised cells

OLD, an abortive immunity protein from prophage P2, consists of an ABC ATPase sensor and a TOPRIM nuclease effector -- a core architecture shared by a large protein family, including components of antiphage systems Gabija, PARIS, Septu, and Lamassu. OLD was originally identified for its lethality in recB-deficient cells and inhibition of bacteriophage {lambda} infection, but the mechanisms governing its activation have remained elusive. Here, we present the cryo-EM structure of an inactive OLD tetramer and show that destabilization into dimeric form opens the TOPRIM catalytic site, stimulating tRNA cleavage. This activity arrests translation, a phenotype rescued by phage-encoded tRNAs. We demonstrate that OLD activation is not strictly RecBCD-dependent: OLD binds aberrant DNA structures in recB-deficient cells, but activation during infection requires recognition of single-stranded DNA hairpins at the phage replication origin. Collectively, our findings reveal how host and phage DNA processing factors create a complex landscape controlling OLD-mediated immunity.

molecular biology↗

bioRxiv2024

Unraveling cysteine deficiency-associated rapid weight loss

Forty percent of the US population and 1 in 6 individuals worldwide are obese, and the incidence of this disease is surging globally1,2. Various dietary interventions, including carbohydrate and fat restriction, and more recently amino acid restriction, have been explored to combat this epidemic3-6. We sought to investigate the impact of removing individual amino acids on the weight profiles of mice. Compared to essential amino acid restriction, induction of conditional cysteine restriction resulted in the most dramatic weight loss, amounting to 20% within 3 days and 30% within one week, which was readily reversed. This weight loss occurred despite the presence of substantial cysteine reserves stored in glutathione (GSH) across various tissues7. Further analysis demonstrated that the weight reduction primarily stemmed from an increase in the utilization of fat mass, while locomotion, circadian rhythm and histological appearance of multiple other tissues remained largely unaffected. Cysteine deficiency activated the integrated stress response (ISR) and NRF2-mediated oxidative stress response (OSR), which amplify each other, leading to the induction of GDF15 and FGF21, hormones associated with increased lipolysis, energy homeostasis and food aversion8-10. We additionally observed rapid tissue coenzyme A (CoA) depletion, resulting in energetically inefficient anaerobic glycolysis and TCA cycle, with sustained urinary excretion of pyruvate, orotate, citrate, -ketoglutarate, nitrogen rich compounds and amino acids. In summary, our investigation highlights that cysteine restriction, by depleting GSH and CoA, exerts a maximal impact on weight loss, metabolism, and stress signaling compared to other amino acid restrictions. These findings may pave the way for innovative strategies for addressing a range of metabolic diseases and the growing obesity crisis.

physiology↗

bioRxiv2023

Persistence of backtracking by human RNA polymerase II

RNA polymerase II (pol II) can backtrack during transcription elongation, exposing the 3 end of nascent RNA. Nascent RNA sequencing can approximate the location of backtracking events that are quickly resolved; however, the extent and genome wide distribution of more persistent backtracking is unknown. Consequently, we developed a novel method to directly sequence the extruded, "backtracked" 3 RNA. Our data shows that pol II slides backwards more than 20 nucleotides in human cells and can persist in this backtracked state. Persistent backtracking mainly occurs where pol II pauses near promoters and intron-exon junctions, and is enriched in genes involved in translation, replication, and development, where gene expression is decreased if these events are unresolved. Histone genes are highly prone to persistent backtracking, and the resolution of such events is likely required for timely expression during cell division. These results demonstrate that persistent backtracking has the potential to affect diverse gene expression programs.

molecular biology↗

bioRxiv2023

General Transcription Factor from E. coli with a Distinct Mechanism of Action

Gene expression in E. coli is controlled by well-established mechanisms that activate or repress transcription. Here, we identify CedA as an unconventional transcription factor specifically associated with the RNA polymerase (RNAP) {sigma}70 holoenzyme. Structural and biochemical analysis of CedA bound to RNAP reveal that it bridges distant domains of {beta} and {sigma}70 subunits to stabilize an open-promoter complex. Remarkably, CedA does so without contacting DNA. We further show that cedA is strongly induced in response to amino acid starvation, oxidative stress, and aminoglycosides. CedA provides a basal level of tolerance to these clinically relevant antibiotics, as well as to rifampicin and peroxide. Finally, we show that CedA modulates transcription of hundreds of bacterial genes, which explains its pleotropic effect on cell physiology and pathogenesis. One sentence summaryAn integrated structure-function approach uncovers CedA as a general transcription initiation factor in E. coli and elucidates its multifaceted role and unique mechanism.

molecular biology↗

bioRxiv2018

YcjW: a transcription factor that controls the emergency generator of H2S in E. coli.

Hydrogen sulfide (H2S) is a ubiquitous gaseous molecule that is endogenously produced in both eukaryotes and prokaryotes. Its role as a pleiotropic signaling molecule has been well characterized in mammals1,2. In contrast, the physiological role of H2S in bacteria only recently became apparent; H2S acts as a cytoprotectant against antibiotics-induced stress and affect the cells ability to maintain redox homeostasis 3-5. In E. coli, endogenous H2S production is primarily dependent on 3-mercaptopyruvate sulfurtransferase (3MST), encoded by mstA, previously known as sseA3,4. Here, we show that cells lacking 3MST acquired a unique phenotypic suppressor mutation resulting in compensatory H2S production and tolerance to antibiotics and oxidative stress. Using whole genome sequencing, we mapped a non-synonymous single nucleotide polymorphism (SNP) to uncharacterized Laci-type transcription factor, YcjW. We identified transcriptional regulatory targets of YcjW and discovered a major target, thiosulfate sulfurtransferase PspE, as an alternative mechanism for H2S biosynthesis. Deletion of pspE was sufficient to antagonize phenotypic suppression. Our results reveal a complex interaction between cell metabolism and H2S production and the role, a hithero uncharacterized transcription factor, YcjW, plays in linking the two.

microbiology↗