Diverse functions of restriction-modification systems in addition to cellular defense

Abstract

Restriction-modification (R-M) systems are ubiquitous and are often considered primitive immune systems in bacteria. Their diversity and prevalence across the prokaryotic kingdom are an indication of their success as a defense mechanism against invading genomes. However, their cellular defense function does not adequately explain the basis for their immaculate specificity in sequence recognition and nonuniform distribution, ranging from none to too many, in diverse species. The present review deals with new developments which provide insights into the roles of these enzymes in other aspects of cellular function. In this review, emphasis is placed on novel hypotheses and various findings that have not yet been dealt with in a critical review. Emerging studies indicate their role in various cellular processes other than host defense, virulence, and even controlling the rate of evolution of the organism. We also discuss how R-M systems could have successfully evolved and be involved in additional cellular portfolios, thereby increasing the relative fitness of their hosts in the population.

Fig 1

Restriction-modification (R-M) systems as defense mechanisms. R-M systems recognize the methylation status of incoming foreign DNA, e.g., phage genomes. Methylated sequences are recognized as self, while recognition sequences on the incoming DNA lacking methylation are recognized as nonself and are cleaved by the restriction endonuclease (REase). The methylation status at the genomic recognition sites is maintained by the cognate methyltransferase (MTase) of the R-M system.

Fig 2

Distribution of R-M systems. (A) Genome-wide analysis for the presence of conserved MTase genes among bacteria with genome sizes ranging from 0.5 to 13 Mbp. The plot shows the median value of the distribution of the number of MTase genes with the specified class interval of genome size. A correlation of an increase in the number of modification systems with an increase in the genome size can be observed. The anomalous decrease (in the 1- to 1.5-Mbp genome size class) in the distribution of R-M systems is because of many Brucella species harboring a single R-M system per chromosome. The presence of multiple R-M systems among Helicobacter and Campylobacter species brings an anomalous increase in the 1.5- to 2-Mbp genome size class. (B) A linear correlation can be observed when the above-mentioned bacterial species are omitted from the 1- to 1.49-Mbp and the 1.5- to 1.99-Mbp classes.

Fig 3

Abundance of R-M systems in naturally competent bacteria. Whole-genome sequence analyses of some of the naturally competent bacteria show that they are rich in R-M genes (5 to 34 genes) compared to other noncompetent bacteria (e.g., a single R-M system in many Bacillus anthracis strains).

Fig 4

Role of R-M systems in recombination. R-M systems effectively restrict incoming DNA. (A) Restriction of incoming DNA from a closely related bacterium (harboring similar Chi sequences) generates DNA fragments which can be utilized as substrates for homologous recombination by the RecBCD pathway. (B) In contrast, the fragments generated by the restriction of phage DNA (lacking the Chi sequence) are recognized as nonself and subjected to further degradation by the RecBCD pathway.

Fig 5

Postsegregational cell killing. Plasmid-harbored R-M gene complexes tend to propagate as selfish genetic elements to promote their own survival. The R-M system present in a cell expresses both REase and MTase: the REase restricts the foreign DNA, and the MTase protects the host genome against cleavage by the cognate REase. The postsegregational loss of the R-M gene complex results in the loss of methylation. The REase, owing to its higher level of stability, attacks the unmodified host genome, resulting in cell death (see “Selfish Genes”). The R-M gene complex thus propagates in the clonal population, resulting in the addiction of the host cell.

Fig 6

Role of R-M systems in the evolution of new strains. The horizontal transfer of DNA in bacteria increases the genetic diversity among them. A bacterial cell which acquires a new R-M gene complex (right) becomes genetically isolated from its clonal population (left). The MTase component of the newly acquired R-M system modifies the genome. Owing to this change in the methylation pattern, the REase prevents the genetic exchange of alleles between closely related strains. Furthermore, mutations acquired in these populations would facilitate genetic diversity, resulting in different genotypes. These populations would further evolve into different strains.

Fig 7

Distribution of R-M systems in RecBC⁻ organisms. Shown are data from a genome-wide analysis of the presence of conserved methyltransferase genes among bacteria with genome sizes ranging from 0.5 to 13 Mb (see the supplemental material). The plot shows the mean values for the distributions of numbers of R-M systems with the specified class intervals of genome size. The list of organisms lacking RecBC was taken from data reported previously (229, 230). A correlation of an increase in the number of R-M systems in RecBC⁻ organisms compared to the total distribution of R-M systems can be observed.

Fig 8

Role of R-M systems in genome evolution. The probable role of defense systems in the evolution of genomes is depicted. (A) Initially, RNA viruses coexisted with bacteria containing RNA genomes. With the evolution of uridine-containing DNA (U-DNA) genomes in bacteria and the acquisition of RNA-dependent endonucleases, a primitive R-M system could have ensured the restriction of the RNA viruses. (B) Such a selection pressure would enforce the evolution of a U-DNA genome in viruses to evade this primitive R-M system. This in turn would result in the evolution of thymidine-containing DNA (T-DNA) genomes in bacteria to evade phage infection. (C) The phage adapts to the host defense strategy by evolving a T-DNA genome. (D) Continuous selection would result in an “arms race” between bacteria and viruses, resulting in the utilization of modified DNA bases in phage and bacterial genomes.