Biology subjects

Bernheim, A.

Publications and source records attributed to Bernheim, A..

27 records · Page 2Linked to original sources

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

Multiple phage resistance systems inhibit infection via SIR2-dependent NAD+ depletion

Defense-associated sirtuins (DSR) comprise a family of proteins that defend bacteria from phage infection via an unknown mechanism. These proteins are common in bacteria and harbor an N-terminal sirtuin (SIR2) domain. In this study we report that DSR proteins degrade nicotinamide adenine dinucleotide (NAD+) during infection, depleting the cell of this essential molecule and aborting phage propagation. Our data show that one of these proteins, DSR2, directly identifies phage tail tube proteins and then becomes an active NADase in Bacillus subtilis. Using a phage mating methodology that promotes genetic exchange between pairs of DSR2-sensitive and DSR2-resistant phages, we further show that some phages express anti-DSR2 proteins that bind and repress DSR2. Finally, we demonstrate that the SIR2 domain serves as an effector NADase in a diverse set of phage defense systems outside the DSR family. Our results establish the general role of SIR2 domains in bacterial immunity against phages.

microbiology↗

bioRxiv2021

Reversing anterior insular cortex neuronal hypoexcitability attenuates compulsive behavior in juvenile rats

Development of self-regulatory competencies during adolescence is partially dependent on normative brain maturation. Here we report that juvenile rats as compared to adults exhibit impulsive and compulsive-like behavioral traits, the latter being associated with lower expression of mRNA levels of the immediate early gene zif268 in the anterior insula (AI). This observation suggests that deficits in AI function in juvenile rats could explain their immature pattern of interoceptive cue integration in rational decision-making and compulsive phenotype. In support of this, here we report hypoexcitability of juvenile layer-V pyramidal neurons in the AI, concomitant with reduced glutamatergic synaptic input to these cells. Chemogenetic activation of the AI attenuated the compulsive trait suggesting that delayed maturation of the AI results in suboptimal integration of sensory and cognitive information in adolescents and this contributes to inflexible behaviors in specific conditions of reward availability.

neuroscience↗

bioRxiv2021

Systematic and quantitative view of the antiviral arsenal of prokaryotes

Facing the abundance and diversity of phages, bacteria have developed multiple anti-phage mechanisms. In the past three years, the number of known anti-phage mechanisms has been expanded by at least 5-fold rendering our view of prokaryotic immunity obsolete. Most anti-phage systems have been studied as standalone mechanisms, however many examples demonstrate strains encode not one but several anti-viral mechanisms. How these different systems integrate into an anti-viral arsenal at the strain level remains to be elucidated. Much could be learned from establishing fundamental description of features such as the number and diversity of anti-phage systems encoded in a given genome. To address this question, we developed DefenseFinder, a tool that automatically detects known anti-phage systems in prokaryotic genomes. We applied DefenseFinder to >20 000 fully sequenced genomes, generating a systematic and quantitative view of the anti-viral arsenal of prokaryotes. We show prokaryotic genomes encode on average five anti-phage systems from three different families of systems. This number varies drastically from one strain to another and is influenced by the genome size and the number of prophages encoded. Distributions of different systems are also very heterogenous with some systems being enriched in prophages and in specific clades. Finally, we provide a detailed comparison of the anti-viral arsenal of 15 common bacterial species, revealing drastic differences in anti-viral strategies. Overall, our work provides a free and open-source software, available as a command line tool or, on a webserver. It allows the rapid detection of anti-phage systems, enables a comprehensive description of the anti-viral arsenal of prokaryotes and paves the way for large scale genomics study in the field of anti-phage defense.

microbiology↗

bioRxiv2021

Prophage-encoded hotspots of bacterial immune systems

The arms race between bacteria and phages led to the emergence of a variety of genetic systems used by bacteria to defend against viral infection, some of which were repurposed as powerful biotechnological tools. While numerous defense systems have been identified in genomic regions termed defense islands, it is believed that many more remain to be discovered. Here, we show that P2- like prophages and their P4-like satellites have genomic hotspots that represent a significant source of novel anti-phage systems. We validate the defense activity of 14 systems spanning various protein domains and describe PARIS, an abortive infection system triggered by a phage-encoded anti-restriction protein. Immunity hotspots are present across prophages of distant bacterial species, highlighting their biological importance in the competition between bacteria and phages.

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↗

bioRxiv2019

Engineering gene overlaps to sustain genetic constructs in vivo

Evolution is often an obstacle to the engineering of stable biological systems due to the selection of mutations inactivating costly gene circuits. Gene overlaps induce important constraints on sequences and their evolution. We show that these constraints can be harnessed to increase the stability of synthetic circuits by purging loss-of-function mutations. We combine computational and synthetic biology approaches to rationally design an overlapping reading frame expressing an essential gene within an existing gene to protect. Our algorithm succeeded in creating overlapping reading frames in 80% of E. coli genes. Experimentally, scoring mutations in both genes of such overlapping construct, we found that a significant fraction of mutations impacting the gene to protect have a deleterious effect on the essential gene. Such an overlap thus protects a costly gene from removal by natural selection by associating the benefit of this removal with a larger or even lethal cost. In our synthetic constructs, the overlap converts many of the possible mutants into evolutionary dead-ends, effectively changing the fitness landscape and reducing the evolutionary potential of the system.

synthetic biology↗

bioRxiv2019

Co-occurrence of multiple CRISPRs and cas clusters suggests epistatic interactions

Prokaryotes use CRISPR-Cas for adaptive immunity, but the reasons for the existence of multiple CRISPR and cas clusters remain poorly understood. We found that more than 40% of the genomes encoding a system show atypical genetic organizations. Their analysis revealed negative and positive epistatic interactions between Cas subtypes. The latter often result in one single complex locus with a shared adaptation module and diverse interference mechanisms, presumably to produce more effective immune systems. We typed CRISPRs that could not be unambiguously associated with a cas cluster and found that such complex loci tend to have unique type I repeats in multiple CRISPRs. In contrast, under-represented co-occurrences caused by functional interference or redundancy may lead to CRISPRs distant from cas genes. To investigate the origin of atypical CRISPR-Cas organizations, we analyzed plasmids and phages. Sets of nearly 2000 phages and 10000 prophages were almost devoid of CRISPR-Cas systems, but a sizeable fraction of plasmids had them. Isolated CRISPRs in plasmids were often compatible with the chromosomal cas clusters, suggesting that plasmids use CRISPRs to subvert host immunity. These results point to an important role for the interactions between multiple CRISPR and Cas in the function and evolution of bacterial immunity.

genomics↗

bioRxiv2017

Inhibition of NHEJ repair by type II-A CRISPR-Cas systems

CRISPR-Cas systems introduce double strand breaks into DNA of invading genetic material and use DNA fragments to acquire novel spacers during adaptation. Double strand breaks are the substrate of several bacterial DNA repair pathways, paving the way for interactions between them and CRISPR-Cas systems. Here, we hypothesized that non-homologous end joining (NHEJ) interferes with type II CRISPR-Cas systems. We tested this idea by studying the patterns of co-occurrence of the two systems in bacterial genomes. We found that NHEJ and type II-A CRISPR-Cas systems only co-occur once among 5563 fully sequenced prokaryotic genomes. We investigated experimentally the possible molecular interactions causing this negative association using the NHEJ pathway from Bacillus subtilis and the type II-A CRISPR-Cas systems from Streptococcus thermophilus and Streptococcus pyogenes. Our results suggest that the NHEJ system has no effect on type II-A CRISPR-Cas interference and adaptation. On the other hand, we provide evidence for the inhibition of NHEJ repair by the Csn2 protein from type II-A CRISPR-Cas system. Our findings give insights on the complex interactions between CRISPR- Cas systems and repair mechanisms in bacteria and contribute to explain the scattered distribution of CRISPR-Cas systems in bacterial genomes.

microbiology↗