Research

How bacteria defend themselves against their viruses ?

Facing the abundance and diversity of their viruses, bacteria and archaea have developed multiple lines of defense that can be referred to as « prokaryotic immune systems« . My research focuses on these anti-phage immune systems.

I’m trying to understand evolutionary patterns and molecular mechanisms of these systems but also how to use them for medical applications. I work at several scales: from computational genomic analysis on thousands of prokaryotic genomes to experimental molecular genetics and diverse microbiology tools.

My PhD focused on the adaptive immune systems of bacteria, CRISPR-Cas systems. I focused on why they are encoded by some bacteria but not others. During my post-doc, I discovered and characterized novel immune systems of bacteria, namely retrons and viperins that showed incredible conservation with eukaryotic immune systems.

I’m currently working on the ecology and evolution of anti-phage systems, their conservation with eukaryotic immune systems and how we can harness them for the fight against pathogens.
You can discover my work through a seminar I gave in July 2020

Prokaryotic viperins, a novel family of defense systems that produce antiviral molecules


Viperin is an important anti-viral protein of humans that is conserved in animals. It has been shown to inhibit the replication of multiple human viruses by producing a molecule called ddhCTP, which acts as a chain terminator for viral RNA polymerase (Gizzi et al. 2018).

We showed that eukaryotic viperin originated from a clade of bacterial and archaeal proteins that protect against phage infection. Prokaryotic viperins produce a set of modified ribonucleotides that include ddhCTP, ddhGTP and ddhUTP. We further showed that prokaryotic viperins protect against T7 phage infection by inhibiting viral polymerase-dependent transcription, suggesting that it has an antiviral mechanism of action similar to that of animal viperin.

This study showed for the first time that natural antiviral compounds are produced by bacterial immune systems, opening avenues to look for more anti-viral molecules generated by bacteria. It’s also the first time such a strong conservation between a eukaryotic and prokaryotic immune system was demonstrated.

Publication: Prokaryotic viperins produce diverse antiviral molecules.
Bernheim A. Millman A., Ofir G., Meitav G., Abraham C., Shomar H., Rosenberg M., Tal N., Melamed S., Amitai G., Sorek R. Nature, in press (2020)
My Twitter thread explaining the discovery
Scientific Commentary: Cell Host and Microbes
General public summary: English or French:
Press: Times of Israel, Jerusalem Post, News1, israel21, Sciences et avenir


Retrons, mysterious bacterial elements function in anti-phage defense


With two collaborators (Adi Millman and Avigail Stokar-Avihail), we elucidated the physiological functions of genetic elements that had been a mystery for 30 years: retrons. Retrons are bacterial genetic elements comprised of a reverse transcriptase (RT) and a non-coding RNA (ncRNA). We stumbled upon a defense system that encodes a retron. This led us to hypothesize that all retrons function as defense systems. We found out that the defensive unit is composed of three components: the RT, the ncRNA, and an effector protein. We examined multiple retron systems and show that they confer defense against a broad range of phages via abortive infection. Focusing on retron Ec48, we showed evidence that it ‘‘guards’’ RecBCD, a complex with central anti-phage functions in bacteria. Inhibition of RecBCD by 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.
This study solved a three-decades old question, and demonstrated a novel concept in bacterial immunology (the guard hypothesis) inspired by the plant immunology field and conserved in prokaryotes.

Publication: Bacterial retrons function in anti- phage defense
Millman A*, Bernheim A*, Stokar-Avihail A.*, Fedorenko T., Voichek M., Leavitt A., Sorek R.
Cell, in press, (2020)

Our Twitter thread explaining the discovery
Scientific Commentaries: Science, The CRISPR journal, Nature Reviews Microbiology
General public summary:English
Press: Jerusalem Post, Phys.org, Drug and Target Review, Science Daily


Diversity and ecology of defense systems


This perspective provides a conceptual framework for the evolutionary forces that lead to the diverse immune systems and their original distribution in bacterial genomes.

The recent discovery of the unexpected diversity of prokaryotic immune arsenal have led to seemingly contradictory observations: on one hand, individual microorganisms often encode multiple distinct defense systems, some of which are acquired by horizontal gene transfer, suggesting they yield a fitness benefit. On the other hand, defense systems are frequently lost from prokaryotic genomes on short evolutionary time scales, suggesting that they impose a fitness cost. We introduced the ‘pan-immune system’ model in which we suggest that, although a single strain cannot carry all possible defense systems owing to their burden on fitness, it can employ horizontal gene transfer to access immune defense mechanisms encoded by closely related strains. Thus, we propose that the ‘effective’ immune system is not the one encoded by the genome of a single microorganism but rather by its pan-genome, comprising the sum of all immune systems available for a microorganism to horizontally acquire and use.

Publication:
The bacterial pan-immune system: anti-phage defense as a community resource

Bernheim A, Sorek R
Nature Reviews Microbiology 18, 113–119 (2020)


The downsides of CRISPR-Cas systems


A small video that I made in collaboration with Adrien Bernheim, a graphic designer to summarize my PhD findings (in french)
THe main interaction I elucidate during my Phd: the antagonism of NHEJ and type II-A CRISPR-Cas

For the past ten years, CRISPR-Cas systems have passionated the scientific community both because of their role as an adaptive immune system in bacteria and of their use in many biotechnological applications especially in genome editing. However, much remains to be studied on their evolution. My PhD research stemmed from the observation that only 50% of bacterial genomes harbor a CRISPR-Cas system despite their apparent fitness advantage and their high rate of horizontal transfer.

Hypothesis such as the cost of autoimmunity or the trade off between a constitutive or an inducible defense system have been put forward to explain this paradox. I proposed that the genetic background plays an important role in the process of maintaining a CRISPR-Cas system after its transfer. More precisely I hypothesized that CRISPR-Cas systems interact with DNA repair pathways. To test this idea, we detected DNA repair pathways and CRISPR-Cas systems in bacterial genomes and studied their co-occurences. We report both positive and negative associations that we interpret as potential antagonistic or synergistic interactions. We then focused on one interaction to validate our result experimentally and explored molecular mechanisms behind those interactions.

My findings gave insights on the complex interactions between CRISPR-Cas systems and DNA repair mechanisms in bacteria and provide a first example on the necessity of accommodation of CRISPR-Cas systems to a specific genetic context to be selected and maintained in bacterial genomes.

Publications:
Atypical organizations and epistatic interactions of CRISPRs and cas clusters in genomes and their mobile gemetic elements.
Bernheim A, Bikard D, Touchon M, Rocha EPC
Nucleic Acids Research 48 (2), 748-760 (2019)
A matter of background: DNA repair, pathways as a possible cause for the sparse distribution of CRISPR–Cas, systems in bacteria.
Bernheim A, Bikard D, Touchon M, Rocha EPC
Philosophical Transactions of the Royal Society B 374 (1772), 2018008 (2019)
Inhibition of NHEJ repair by type II-A CRISPR-Cas systems
Bernheim A, Calvo-Villaman A, Basier C, Cui L, Rocha EPC, Touchon M, Bikard D,
Nature Communications 8 (1), 1-9 (2017)