Removal of a frameshift between the hsdM and hsdS genes of the EcoKI Type IA DNA restriction and modification system produces a new type of system and links the different families of Type I systems

Roberts, Gareth A.; Chen, Kai; Cooper, Laurie P.; White, John H.; Blakely, Garry W.; Dryden, David T. F.

doi:10.1093/nar/gks876

Cited by 9 publications

(10 citation statements)

References 51 publications

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“…For clarity we designate the upstream MTase subunit containing the NPPF motif as ‘M1’, and the downstream MTase subunit containing the NPPY motif IV as ‘M2.’ As is common with Type I systems, some of these systems have additional open reading frames interspersed between the R-M system component genes; most commonly between the two MTase genes (6 of 10 characterized systems), but also between the M2.MTase and the S gene (two systems), or between the S gene and the R gene (one system). In one case, the M2 MTase gene is fused to the specificity subunit: M2.Dac11109IV contains both the M2 subunit and the S subunit by sequence similarity analysis, similar to the previous demonstration that the hsdM and hsdS genes of the EcoKI system could be fused into a single orf that maintained function (16). This is also consistent with the observation that the M2 (NPPY) MTase is located next to the S subunit.…”

Section: Resultssupporting

confidence: 73%

Novel m4C modification in type I restriction-modification systems

et al. 2016

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We identify a new subgroup of Type I Restriction-Modification enzymes that modify cytosine in one DNA strand and adenine in the opposite strand for host protection. Recognition specificity has been determined for ten systems using SMRT sequencing and each recognizes a novel DNA sequence motif. Previously characterized Type I systems use two identical copies of a single methyltransferase (MTase) subunit, with one bound at each half site of the specificity (S) subunit to form the MTase. The new m4C-producing Type I systems we describe have two separate yet highly similar MTase subunits that form a heterodimeric M1M2S MTase. The MTase subunits from these systems group into two families, one of which has NPPF in the highly conserved catalytic motif IV and modifies adenine to m6A, and one having an NPPY catalytic motif IV and modifying cytosine to m4C. The high degree of similarity among their cytosine-recognizing components (MTase and S) suggest they have recently evolved, most likely from the far more common m6A Type I systems. Type I enzymes that modify cytosine exclusively were formed by replacing the adenine target recognition domain (TRD) with a cytosine-recognizing TRD. These are the first examples of m4C modification in Type I RM systems.

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Section: Resultssupporting

confidence: 73%

Novel m4C modification in type I restriction-modification systems

et al. 2016

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“…S1, see Supporting Information). Precedent exists for this type of genomic organization in the bacterial EcoKI DNA methyltransferase (Roberts et al ., ). This trimeric protein comprises two modification subunits (M) and one sequence specificity subunit (S).…”

Section: Resultsmentioning

confidence: 97%

“…This trimeric protein comprises two modification subunits (M) and one sequence specificity subunit (S). The 3′ end of the gene encoding the M subunit overlaps the 5′ end of the S subunit by one nucleotide where translation from the two different open reading frames is translationally coupled (Roberts et al ., ). Further investigation will be required to determine whether these RrCEPs are similarly translationally coupled, and whether or not both are produced as full‐length functional proteins.…”

Section: Resultsmentioning

confidence: 97%

Functional C‐TERMINALLY ENCODED PEPTIDE (CEP) plant hormone domains evolved de novo in the plant parasite Rotylenchulus reniformis

Akker

Lilley

Yusup

et al. 2016

Molecular Plant Pathology

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SummarySedentary plant‐parasitic nematodes (PPNs) induce and maintain an intimate relationship with their host, stimulating cells adjacent to root vascular tissue to re‐differentiate into unique and metabolically active ‘feeding sites’. The interaction between PPNs and their host is mediated by nematode effectors. We describe the discovery of a large and diverse family of effector genes, encoding C‐TERMINALLY ENCODED PEPTIDE (CEP) plant hormone mimics (RrCEPs), in the syncytia‐forming plant parasite Rotylenchulus reniformis. The particular attributes of RrCEPs distinguish them from all other CEPs, regardless of origin. Together with the distant phylogenetic relationship of R. reniformis to the only other CEP‐encoding nematode genus identified to date (Meloidogyne), this suggests that CEPs probably evolved de novo in R. reniformis. We have characterized the first member of this large gene family (RrCEP1), demonstrating its significant up‐regulation during the plant–nematode interaction and expression in the effector‐producing pharyngeal gland cell. All internal CEP domains of multi‐domain RrCEPs are followed by di‐basic residues, suggesting a mechanism for cleavage. A synthetic peptide corresponding to RrCEP1 domain 1 is biologically active and capable of up‐regulating plant nitrate transporter (AtNRT2.1) expression, whilst simultaneously reducing primary root elongation. When a non‐CEP‐containing, syncytia‐forming PPN species (Heterodera schachtii) infects Arabidopsis in a CEP‐rich environment, a smaller feeding site is produced. We hypothesize that CEPs of R. reniformis represent a two‐fold adaptation to sustained biotrophy in this species: (i) increasing host nitrate uptake, whilst (ii) limiting the size of the syncytial feeding site produced.

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“…Of note is that extra HsdM was NOT required for MS fus to be active. This is in contrast to the MS fus protein constructed from the EcoKI Type I RM system which needed additional HsdM to be added to the reaction ( 42 ). This is consistent with the identification of a fragment of the fusion protein remaining associated with CC398-1 MS fus throughout protein purification.…”

Section: Resultsmentioning

confidence: 99%

“…Roberts et al. were able to remove this frameshift from the MTase genes of the EcoKI Type I enzyme to create a fusion of the M and S subunits ( 42 ). This protein product showed full RM activity in vivo and was also successfully over-expressed and purified.…”

Section: Introductionmentioning

confidence: 99%

A model for the evolution of prokaryotic DNA restriction-modification systems based upon the structural malleability of Type I restriction-modification enzymes

Bower

Cooper

Roberts

et al. 2018

Nucleic Acids Research

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Restriction Modification (RM) systems prevent the invasion of foreign genetic material into bacterial cells by restriction and protect the host's genetic material by methylation. They are therefore important in maintaining the integrity of the host genome. RM systems are currently classified into four types (I to IV) on the basis of differences in composition, target recognition, cofactors and the manner in which they cleave DNA. Comparing the structures of the different types, similarities can be observed suggesting an evolutionary link between these different types. This work describes the ‘deconstruction’ of a large Type I RM enzyme into forms structurally similar to smaller Type II RM enzymes in an effort to elucidate the pathway taken by Nature to form these different RM enzymes. Based upon the ability to engineer new enzymes from the Type I ‘scaffold’, an evolutionary pathway and the evolutionary pressures required to move along the pathway from Type I RM systems to Type II RM systems are proposed. Experiments to test the evolutionary model are discussed.

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Removal of a frameshift between the hsdM and hsdS genes of the EcoKI Type IA DNA restriction and modification system produces a new type of system and links the different families of Type I systems

Cited by 9 publications

References 51 publications

Novel m4C modification in type I restriction-modification systems

Novel m4C modification in type I restriction-modification systems

Functional C‐TERMINALLY ENCODED PEPTIDE (CEP) plant hormone domains evolved de novo in the plant parasite Rotylenchulus reniformis

A model for the evolution of prokaryotic DNA restriction-modification systems based upon the structural malleability of Type I restriction-modification enzymes

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