Quorum Sensing Controls Adaptive Immunity through the Regulation of Multiple CRISPR-Cas Systems

Bacteria commonly exist in high cell density populations, making them prone to viral predation and horizontal gene transfer (HGT) through transformation and conjugation. To combat these invaders, bacteria possess an arsenal of defenses, such as CRISPR-Cas adaptive immunity. Many bacterial populations coordinate their behavior as cell density increases, using quorum sensing (QS) signaling. In this study, we demonstrate that QS regulation results in increased expression of the type I-E, I-F, and III-A CRISPR-Cas systems in $\textit{Serratia}$ cells in high-density populations. Strains unable to communicate via QS were less effective at defending against invaders targeted by any of the three CRISPR-Cas systems. Additionally, the acquisition of immunity by the type I-E and I-F systems was impaired in the absence of QS signaling. We propose that bacteria can use chemical communication to modulate the balance between community-level defense requirements in high cell density populations and host fitness costs of basal CRISPR-Cas activity.This work was supported by a Rutherford Discovery Fellowship (P.C.F.) from the Royal Society of New Zealand (RSNZ) and the Marsden Fund, RSNZ. A.G.P. was supported by a University of Otago Doctoral Scholarship. G.P.C.S. is funded by the Biotechnology and Biological Sciences Research Council, UK

Different genetic and morphological outcomes for phages targeted by single or multiple CRISPR-Cas spacers.

CRISPR-Cas systems provide bacteria and archaea with adaptive immunity against genetic invaders, such as bacteriophages. The systems integrate short sequences from the phage genome into the bacterial CRISPR array. These 'spacers' provide sequence-specific immunity but drive natural selection of evolved phage mutants that escape the CRISPR-Cas defence. Spacer acquisition occurs by either naive or primed adaptation. Naive adaptation typically results in the incorporation of a single spacer. By contrast, priming is a positive feedback loop that often results in acquisition of multiple spacers, which occurs when a pre-existing spacer matches the invading phage. We predicted that single and multiple spacers, representative of naive and primed adaptation, respectively, would cause differing outcomes after phage infection. We investigated the response of two phages, ϕTE and ϕM1, to the Pectobacterium atrosepticum type I-F CRISPR-Cas system and observed that escape from single spacers typically occurred via point mutations. Alternatively, phages escaped multiple spacers through deletions, which can occur in genes encoding structural proteins. Cryo-EM analysis of the ϕTE structure revealed shortened tails in escape mutants with tape measure protein deletions. We conclude that CRISPR-Cas systems can drive phage genetic diversity, altering morphology and fitness, through selective pressures arising from naive and primed acquisition events. This article is part of a discussion meeting issue 'The ecology and evolution of prokaryotic CRISPR-Cas adaptive immune systems'.This work was supported by a Rutherford Discovery Fellow- ship from the Royal Society of New Zealand (RSNZ) (to P.C.F.), the Marsden Fund, RSNZ, the Bio-protection Research Centre (Tertiary Education Commission), a University of Otago Doctoral Scholarship (to B.N.J.W.), University of Otago Division of Health Sciences Career Development Post-doctoral Fellowship and a Veni grant (grant no. 016.Veni.171.047) from the The Netherlands Organization for Scienti- fic Research (to R.H.J.S.). G.P.C.S. was supported by the BBSRC, UK

Coevolution between bacterial CRISPR-Cas systems and their bacteriophages

This is the author accepted manuscript. The final version is available from Cell Press via the DOI in this record CRISPR-Cas systems provide bacteria and archaea with adaptive, heritable immunity against their viruses (bacteriophages and phages) and other parasitic genetic elements. CRISPR-Cas systems are highly diverse, and we are only beginning to understand their relative importance in phage defense. In this review, we will discuss when and why CRISPR-Cas immunity against phages evolves, and how this, in turn, selects for the evolution of immune evasion by phages. Finally, we will discuss our current understanding of if, and when, we observe coevolution between CRISPR-Cas systems and phages, and how this may be influenced by the mechanism of CRISPR-Cas immunity.VENIBiotechnology and Biological Sciences Research CouncilBiotechnology and Biological Sciences Research Counci

Keeping crispr in check: diverse mechanisms of phage-encoded anti-crisprs

CRISPR-Cas represents the only adaptive immune system of prokaryotes known to date. These immune systems are widespread among bacteria and archaea, and provide protection against invasion of mobile genetic elements, such as bacteriophages and plasmids. As a result of the arms-race between phages and their prokaryotic hosts, phages have evolved inhibitors known as anti-CRISPR (Acr) proteins to evade CRISPR immunity. In the recent years, several Acr proteins have been described in both temperate and virulent phages targeting diverse CRISPR-Cas systems. Here, we describe the strategies of Acr discovery and the multiple molecular mechanisms by which these proteins operate to inhibit CRISPR immunity. We discuss the biological relevance of Acr proteins and speculate on the implications of their activity for the development of improved CRISPR-based research and biotechnological tools

The Role of CRISPR-Cas Systems in Virulence of Pathogenic Bacteria

Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) genes are present in many bacterial and archaeal genomes. Since the discovery of the typical CRISPR loci in the 1980s, well before their physiological role was revealed, their variable sequences have been used as a complementary typing tool in diagnostic, epidemiologic, and evolutionary analyses of prokaryotic strains. The discovery that CRISPR spacers are often identical to sequence fragments of mobile genetic elements was a major breakthrough that eventually led to the elucidation of CRISPR-Cas as an adaptive immunity system. Key elements of this unique prokaryotic defense system are small CRISPR RNAs that guide nucleases to complementary target nucleic acids of invading viruses and plasmids, generally followed by the degradation of the invader. In addition, several recent studies have pointed at direct links of CRISPR-Cas to regulation of a range of stress-related phenomena. An interesting example concerns a pathogenic bacterium that possesses a CRISPR-associated ribonucleoprotein complex that may play a dual role in defense and/or virulence. In this review, we describe recently reported cases of potential involvement of CRISPR-Cas systems in bacterial stress responses in general and bacterial virulence in particular