Biology subjects

Doron, S.

Publications and source records attributed to Doron, S..

7 recordsLinked to original sources

bioRxiv2025

Two opposing redox signals mediated by 2-Cys peroxiredoxin shape the redox proteome during photosynthetic induction

Photosynthetic induction, characterized by the lag in CO2 assimilation rates typically observed upon plant transition from darkness to light, has traditionally been attributed to Rubisco activase activity and stomatal opening. Yet, the faster induction of photosynthesis in the 2-Cys peroxiredoxins (Prxs) mutant (2cpab) highlighted the critical role of chloroplast redox state in regulating photosynthetic rates during this phase. Since 2-Cys Prxs are involved in transmission of oxidative signals to target enzymes, it was hypothesized that it slows down photosynthesis during the induction phase. SPEAR, a redox proteomics approach for simultaneous protein expression and redox analysis, was used to systematically map redox changes occurring at the proteome level during photosynthesis induction and to unravel the role of 2-Cys Prxs in shaping these redox alterations. No significant difference was observed in protein expression levels between WT and 2cpab plants, suggesting that protein abundance does not account for the 2cpab phenotype. During the transition from dark to low light, 82 and 54 cysteine-containing peptides were reduced or oxidized, respectively, in WT plants. Most redox-regulated cysteines in photosynthetic proteins were found oxidized in the dark and became reduced in response to light, including ATP synthase gamma chain 1 (ATPC1) and glyceraldehyde-3-phosphate dehydrogenase (GAPB). A reverse pattern was observed among redox-regulated cysteines in proteins involved in starch degradation and chloroplast glycolysis, which shifted from a reduced to an oxidized state in response to light. These findings demonstrate the initiation of two opposing redox responses, affecting distinct sets of metabolic proteins during the induction phase. Remarkably, a significantly lower number of cysteines were reduced or oxidized in 2cpab plants, highlighting the crucial role 2-Cys Prxs play in shaping both signals. Taken together, rotational shifts between metabolic pathways during the photosynthesis induction phase are regulated by two opposing redox signals mediated by 2-Cys Prx activity.

plant biology↗

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↗

bioRxiv2022

An expanding arsenal of immune systems that protect bacteria from phages

Bacterial anti-phage defense systems are frequently clustered in microbial genomes, forming defense islands. This genomic property enabled the recent discovery of multiple defense systems based on their genomic co-localization with known systems, but the full arsenal of anti-phage mechanisms in bacteria is still unknown. In this study we report the discovery of 21 new defense systems that protect bacteria from phages, based on computational genomic analyses and phage infection experiments. We find multiple systems with protein domains known to be involved in eukaryotic anti-viral immunity, including ISG15-like proteins, dynamin-like proteins, and SEFIR domains, and show that these domains participate in bacterial defense against phages. Additional systems include protein domains predicted to manipulate DNA and RNA molecules, as well as multiple toxin-antitoxin systems shown here to function in anti-phage defense. The systems we discovered are widely distributed in bacterial and archaeal genomes, and in some bacteria form a considerable fraction of the immune arsenal. Our data substantially expand the known inventory of defense systems utilized by bacteria to counteract phage infection.

microbiology↗

bioRxiv2021

The DarTG toxin-antitoxin system provides phage defense by ADP-ribosylating viral DNA

Toxin-antitoxin (TA) systems are broadly distributed, yet poorly conserved, genetic elements whose biological functions are unclear and controversial. Some TA systems may provide bacteria with immunity to infection by their ubiquitous viral predators, the bacteriophage. To identify TA systems that protect E. coli MG1655 against phage, we searched for those frequently encoded near known phage defense genes in bacterial genomes. Two of the systems tested provide strong protection against phage infection and are homologs of DarTG, a recently discovered family of TA systems whose biological functions and natural activating conditions were unclear. We demonstrate that phage infection triggers the release of DarT toxin, a DNA ADP-ribosyltransferase, to modify viral DNA and prevent replication, thereby blocking the production of mature virions. Phages can evolve to overcome DarTG defense either through mutations to their DNA polymerase or to an anti-DarT factor, gp61.2, encoded by many T-even phages. Collectively, our results indicate that phage defense may be a common function for TA systems and reveal the mechanism by which DarTG systems inhibit phage infection.

microbiology↗

bioRxiv2021

SPEAR: a proteomics approach for simultaneous protein expression and redox analysis

Oxidation and reduction of protein cysteinyl thiols serve as molecular switches, which is considered the most central mechanism for redox regulation of biological processes, altering protein structure, biochemical activity, subcellular localization, and binding affinity. Redox proteomics allows for the global identification of redox-modified cysteine (Cys) sites and quantification of their oxidation/reduction responses, serving as a hypothesis-generating platform to stimulate redox biology mechanistic research. Here, we developed Simultaneous Protein Expression and Redox (SPEAR) analysis, a new redox-proteomics approach based on differential labeling of oxidized and reduced cysteines with light and heavy isotopic forms of commercially available isotopically-labeled N-ethylmaleimide (NEM). The presented method does not require enrichment for labeled peptides, thus enabling simultaneous quantification of Cys oxidation state and protein abundance. Using SPEAR, we were able to quantify the in-vivo oxidation state of thousands of cysteines across the Arabidopsis proteome under steady-state and oxidative stress conditions. Functional assignment of the identified redox-sensitive proteins demonstrated the widespread effect of oxidative conditions on various cellular functions and highlighted the enrichment of chloroplast-targeted proteins. SPEAR provides a simple, straightforward, and cost-effective means of studying redox proteome dynamics. The presented data provide a global quantitative view of cysteine oxidation of well-known redox-regulated active sites and many novel redox-sensitive sites whose role in plant acclimation to stress conditions remains to be further explored.

biochemistry↗

bioRxiv2021

Antiviral activity of bacterial TIR domains via signaling molecules that trigger cell death

The Toll/interleukin-1 receptor (TIR) domain is a canonical component of animal and plant immune systems. In plants, intracellular pathogen sensing by immune receptors triggers their TIR domains to generate a molecule which is a variant of cyclic ADP-ribose (v-cADPR). This molecule is hypothesized to activate plant cell death via a yet unresolved pathway. TIR domains were recently also shown to be involved in a bacterial anti-phage defense system called Thoeris, but the mechanism of Thoeris defense remained unknown. In this study we report that phage infection triggers Thoeris TIR-domain proteins to produce an isomer of cyclic ADP-ribose. This molecular signal activates a second protein, ThsA, which then depletes the cell of the essential molecule nicotinamide adenine dinucleotide (NAD) and leads to abortive infection and cell death. We further show that similar to eukaryotic innate immune systems, bacterial TIR-domain proteins determine the immunological specificity to the invading pathogen. Our results describe a new antiviral signaling pathway in bacteria, and suggest that generation of intracellular signaling molecules is an ancient immunological function of TIR domains conserved in both plant and bacterial immunity.

microbiology↗

bioRxiv2018

A matrix protein acts as a cue to preserve collective motility of B. subtilis biofilm cells

Bacteria in nature are usually found in complex multicellular communities, termed biofilms. Biofilms are generally seen as sessile structures, resulting from downregulation of motility. However, during interspecies competition and predation, biofilm cells were shown to migrate towards competitor colonies. Here, we show that a specific extracellular matrix (ECM) protein, TasA, is essential for collective migration toward potential competitors and serves as a developmental cue that increases the formation of motile offspring from sessile chains. We reveal an effective strategy to maintain migration capacities in bacterial biofilms: besides providing a three-dimensional adhesive scaffold for the cells, TasA acts as a signal within the bacterial community.

microbiology↗