Bacteria and their viral predators (bacteriophages) are locked in a constant battle. In order to proliferate in phage-rich environments, bacteria have an impressive arsenal of defence mechanisms, and in response, phages have evolved counter-strategies to evade these antiviral systems. In this Review, we describe the various tactics that are used by phages to overcome bacterial resistance mechanisms, including adsorption inhibition, restriction-modification, CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR-associated proteins) systems and abortive infection. Furthermore, we consider how these observations have enhanced our knowledge of phage biology, evolution and phage-host interactions.

Revenge of the phages: defeating bacterial defences

Our knowledge of prokaryotic defense systems has vastly expanded as the result of comparative genomic analysis, followed by experimental validation. This expansion is both quantitative, including the discovery of diverse new examples of known types of defense systems, such as restriction-modification or toxin-antitoxin systems, and qualitative, including the discovery of fundamentally new defense mechanisms, such as the CRISPR-Cas immunity system. Large-scale statistical analysis reveals that the distribution of different defense systems in bacterial and archaeal taxa is non-uniform, with four groups of organisms distinguishable with respect to the overall abundance and the balance between specific types of defense systems. The genes encoding defense system components in bacterial and archaea typically cluster in defense islands. In addition to genes encoding known defense systems, these islands contain numerous uncharacterized genes, which are candidates for new types of defense systems. The tight association of the genes encoding immunity systems and dormancy- or cell death-inducing defense systems in prokaryotic genomes suggests that these two major types of defense are functionally coupled, providing for effective protection at the population level.

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Comparative genomics of defense systems in archaea and bacteria

The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas (CRISPR-associated proteins) is a prokaryotic adaptive immune system that is represented in most archaea and many bacteria. Among the currently known prokaryotic defense systems, the CRISPR-Cas genomic loci show unprecedented complexity and diversity. Classification of CRISPR-Cas variants that would capture their evolutionary relationships to the maximum possible extent is essential for comparative genomic and functional characterization of this theoretically and practically important system of adaptive immunity. To this end, a multipronged approach has been developed that combines phylogenetic analysis of the conserved Cas proteins with comparison of gene repertoires and arrangements in CRISPR-Cas loci. This approach led to the current classification of CRISPR-Cas systems into three distinct types and ten subtypes for each of which signature genes have been identified. Comparative genomic analysis of the CRISPR-Cas systems in new archaeal and bacterial genomes performed over the 3 years elapsed since the development of this classification makes it clear that new types and subtypes of CRISPR-Cas need to be introduced. Moreover, this classification system captures only part of the complexity of CRISPR-Cas organization and evolution, due to the intrinsic modularity and evolutionary mobility of these immunity systems, resulting in numerous recombinant variants. Moreover, most of the cas genes evolve rapidly, complicating the family assignment for many Cas proteins and the use of family profiles for the recognition of CRISPR-Cas subtype signatures. Further progress in the comparative analysis of CRISPR-Cas systems requires integration of the most sensitive sequence comparison tools, protein structure comparison, and refined approaches for comparison of gene neighborhoods.

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Annotation and Classification of CRISPR-Cas Systems.

Prokaryotes, bacteria and archaea, are the most abundant cellular organisms among those sharing the planet Earth with human beings (among others). However, numerous ecological studies have revealed that it is actually prokaryotic viruses that predominate on our planet and outnumber their hosts by at least an order of magnitude. An understanding of how this viral domain is organized and what are the mechanisms governing its evolution is therefore of great interest and importance. The vast majority of characterized prokaryotic viruses belong to the order Caudovirales, double-stranded DNA (dsDNA) bacteriophages with tails. Consequently, these viruses have been studied (and reviewed) extensively from both genomic and functional perspectives. However, albeit numerous, tailed phages represent only a minor fraction of the prokaryotic virus diversity. Therefore, the knowledge which has been generated for this viral system does not offer a comprehensive view of the prokaryotic virosphere. In this review, we discuss all families of bacterial and archaeal viruses that contain more than one characterized member and for which evolutionary conclusions can be attempted by use of comparative genomic analysis. We focus on the molecular mechanisms of their genome evolution as well as on the relationships between different viral groups and plasmids. It becomes clear that evolutionary mechanisms shaping the genomes of prokaryotic viruses vary between different families and depend on the type of the nucleic acid, characteristics of the virion structure, as well as the mode of the life cycle. We also point out that horizontal gene transfer is not equally prevalent in different virus families and is not uniformly unrestricted for diverse viral functions.

Genomics of Bacterial and Archaeal Viruses: Dynamics within the Prokaryotic Virosphere

Evolution of bacteria and archaea involves an incessant arms race against an enormous diversity of genetic parasites. Accordingly, a substantial fraction of the genes in most bacteria and archaea are dedicated to antiparasite defense. The functions of these defense systems follow several distinct strategies, including innate immunity; adaptive immunity; and dormancy induction, or programmed cell death. Recent comparative genomic studies taking advantage of the expanding database of microbial genomes and metagenomes, combined with direct experiments, resulted in the discovery of several previously unknown defense systems, including innate immunity centered on Argonaute proteins, bacteriophage exclusion, and new types of CRISPR-Cas systems of adaptive immunity. Some general principles of function and evolution of defense systems are starting to crystallize, in particular, extensive gain and loss of defense genes during the evolution of prokaryotes; formation of genomic defense islands; evolutionary connections between mobile genetic elements and defense, whereby genes of mobile elements are repeatedly recruited for defense functions; the partially selfish and addictive behavior of the defense systems; and coupling between immunity and dormancy induction/programmed cell death.

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Evolutionary Genomics of Defense Systems in Archaea and Bacteria.

Present in the genomes of bacteria and eukaryotic organelles, group II introns are an ancient class of ribozymes and retroelements that are believed to have been the ancestors of nuclear pre-mRNA introns. Despite long-standing speculation, there is limited understanding about the actual pathway by which group II introns evolved into eukaryotic introns. In this review, we focus on the evolution of group II introns themselves. We describe the different forms of group II introns known to exist in nature and then address how these forms may have evolved to give rise to spliceosomal introns and other genetic elements. Finally, we summarize the structural and biochemical parallels between group II introns and the spliceosome, including recent data that strongly support their hypothesized evolutionary relationship.

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Evolution of group II introns

Structural characterization of Nonstructural protein 1 from SARS-CoV-2.

Reverse transcriptases (RTs) are RNA-dependent DNA polymerases that usually function in the replication of selfish DNAs such as retrotransposons and retroviruses. Here, we have biochemically characterized a RT-related protein, AbiK, which is required for abortive phage infection in the Gram-positive bacterium Lactococcus lactis. In vitro, AbiK does not exhibit the properties expected for an RT, but polymerizes long DNAs of 'random' sequence, analogous to a terminal transferase. Moreover, the polymerized DNAs appear to be covalently attached to the AbiK protein, presumably because an amino acid serves as a primer. Mutagenesis experiments indicate that the polymerase activity resides in the RT motifs and is essential for phage resistance in vivo. These results establish a novel biochemical property and a non-replicative biological role for a polymerase.

A reverse transcriptase-related protein mediates phage resistance and polymerizes untemplated DNA in vitro

Group II introns are catalytic RNAs (ribozymes) and retroelements found in the genomes of bacteria, archaebacteria, and organelles of some eukaryotes. The prototypical retroelement form consists of a structurally conserved RNA and a multidomain reverse transcriptase protein, which interact with each other to mediate splicing and mobility reactions. A wealth of biochemical, cross-linking, and X-ray crystal structure studies have helped to reveal how the two components cooperate to carry out the splicing and mobility reactions. In addition to the standard retroelement form, group II introns have evolved into derivative forms by either losing specific splicing or mobility characteristics, or becoming functionally specialized. Of particular interest are the eukaryotic derivatives-the spliceosome, spliceosomal introns, and non-LTR retroelements-which together make up approximately half of the human genome. On a practical level, the properties of group II introns have been exploited to develop group II intron-based biotechnological tools. WIREs RNA 2016, 7:341-355. doi: 10.1002/wrna.1339 For further resources related to this article, please visit the WIREs website.

Group II introns: versatile ribozymes and retroelements.

https://doi.org/10.1016/j.isci.2022.105054

Cameron Semper

Papers

Evolution of group II introns

Structural characterization of Nonstructural protein 1 from SARS-CoV-2.

A reverse transcriptase-related protein mediates phage resistance and polymerizes untemplated DNA in vitro

Group II introns: versatile ribozymes and retroelements.

Recombinant production of growth factors for application in cell culture