Identification and biosynthesis of thymidine hypermodifications in the genomic DNA of widespread bacterial viruses

Lee, Yan‐Jiun; Dai, Nan; Walsh, Shannon E.; Müller, Stephanie; Fraser, Morgan E; Kauffman, Kathryn M.; Guan, Chaowen; Corrêa, Ivan R.; Weigele, Peter

doi:10.1073/pnas.1714812115

Cited by 53 publications

(55 citation statements)

References 67 publications

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“…5hmdU DNA Kinase Activity: Phosphorylation of Phage SP8 DNA to Become More Resistant to Type II Restrictions 5hmdU DNA kinase can phosphorylate the 5-hydroxymethyl group in the 5hmdU in a sequence-specific manner (Lee et al, 2018) making the base more negatively charged. The 5hmdU DNA kinase has been shown to block NcoI restriction after the kinase reaction 2 .…”

Section: Restriction Of Phage Sp8 Dnamentioning

confidence: 99%

“…The companion methyltransferase (MTase) usually modifies the host genome rendering it refractory to the endonuclease and thereby preventing self-restriction (reviewed in Pingoud and Jeltsch, 2001;Pingoud et al, 2016). To counteract host restriction systems, many phages synthesize heavily modified (aka hypermodified) bases in their genomes by a combination of biosynthetic activities acting before and/or after DNA replication (Gold and Schweiger, 1969;Weigele and Raleigh, 2016;Lee et al, 2018). Additionally, some phage or prophage genomes encode multispecificity C5 methyltransferases (Schumann et al, 1996;Xu et al, 2012), Dam methylase (Hattman et al, 1985), or frequent adenine MTase (Drozdz et al, 2012) to similarly protect phage genome from Type II restriction.…”

Section: Introductionmentioning

confidence: 99%

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In vitro Type II Restriction of Bacteriophage DNA With Modified Pyrimidines

et al. 2020

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To counteract host-encoded restriction systems, bacteriophages (phages) incorporate modified bases in their genomes. For example, phages carry in their genomes modified pyrimidines such as 5-hydroxymethyl-cytosine (5hmC) in T4gt deficient in αand β-glycosyltransferases, glucosylated-5-hydroxymethylcytosine (5gmC) in T4, 5methylcytosine (5mC) in Xp12, and 5-hydroxymethyldeoxyuridine (5hmdU) in SP8. In this work we sequenced phage Xp12 and SP8 genomes and examined Type II restriction of T4gt, T4, Xp12, and SP8 phage DNAs. T4gt, T4, and Xp12 genomes showed resistance to 81.9% (186 out of 227 enzymes tested), 94.3% (214 out of 227 enzymes tested), and 89.9% (196 out of 218 enzymes tested), respectively, commercially available Type II restriction endonucleases (REases). The SP8 genome, however, was resistant to only ∼8.3% of these enzymes (17 out of 204 enzymes tested). SP8 DNA could be further modified by adenine DNA methyltransferases (MTases) such as M.Dam and M.EcoGII as well as a number of cytosine DNA MTases, such as CpG methylase. The 5hmdU base in SP8 DNA was phosphorylated by treatment with a 5hmdU DNA kinase to achieve ∼20% phosphorylated 5hmdU, resulting resistance or partially resistant to more Type II restriction. This work provides a convenient reference for molecular biologists working with modified pyrimidines and using REases. The genomic sequences of phage Xp12 and SP8 lay the foundation for further studies on genetic pathways for 5mC and 5hmdU DNA base modifications and for comparative phage genomics.

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Section: Restriction Of Phage Sp8 Dnamentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

In vitro Type II Restriction of Bacteriophage DNA With Modified Pyrimidines

et al. 2020

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“…Well-studied phage DNA modifications include 5-methylcytosine in phage XP12 (Feng, et al, 1978;Huang et al, 1982); 5-hydroxymethylcytosine in phage T4gt, β-glucosyl-5-hydroxymethylcytosine in phage T4 (Cao et al, 1983), N6-(1-acetamido)-adenine in bacteriophage Mu (Hattman, 1979) and 5-hydroxymethyluracil in Bacillus phage SP8 (Vilpo and Vilpo, 1995). In addition, recently discovered 5-(2-aminoethoxy)methyluridine in Salmonella phage VII, 5-(2-aminoethyl)uridine in Pseudomonas phage M6 (Lee et al, 2018) and phosphorothioate (PT) modification of the DNA backbone (Wang et al, 2007;Chen et al, 2017) expanded the list of naturally-occurring DNA modifications.…”

Section: Introductionmentioning

confidence: 99%

Identification of the minimal bacterial 2′‐deoxy‐7‐amido‐7‐deazaguanine synthesis machinery

Yuan

Hutinet

Valera

et al. 2018

Molecular Microbiology

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The 7-deazapurine derivatives, 2'-deoxy-7-cyano-7-deazaguanosine (dPreQ ) and 2'-deoxy-7-amido-7-deazaguanosine (dADG) are recently discovered DNA modifications encoded by the dpd cluster found in a diverse set of bacteria. Here we identify the genes required for the formation of dPreQ and dADG in DNA and propose a biosynthetic pathway. The preQ base is a precursor that in Salmonella Montevideo, is synthesized as an intermediate in the pathway of the tRNA modification queuosine. Of the 11 genes (dpdA - dpdK) found in the S. Montevideo dpd cluster, dpdA and dpdB are necessary and sufficient to synthesize dPreQ , while dpdC is additionally required for dADG synthesis. Among the rest of the dpd genes, dpdE, dpdG, dpdI, dpdK, dpdD and possibly dpdJ appear to be involved in a restriction-like phenotype. Indirect competition for preQ base led to a model for dADG synthesis in which DpdA inserts preQ into DNA with the help of DpdB, and then DpdC hydrolyzes dPreQ to dADG. The role of DpdB is not entirely clear as it is dispensable in other dpd clusters. Our discovery of a minimal gene set for introducing 7-deazapurine derivatives in DNA provides new tools for biotechnology applications and demonstrates the interplay between the DNA and RNA modification machineries.

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“…This is also true for the hydroxymethyluracil synthase ( Figure 3 ). Interestingly, there is some evidence that other members of the Ackermannviridae family may possess modified bases derived from HmUra [ 67 , 68 ]. However, the only deduced ΦW-14 protein remotely related to a polyamine biosynthetic protein is a 434-amino acid polypeptide that is a putative bifunctional glutathionylspermidine synthetase/amidase (gp91).…”

Section: Resultsmentioning

confidence: 99%

The Sequence of Two Bacteriophages with Hypermodified Bases Reveals Novel Phage-Host Interactions

Kropinski

Turner

Nash

et al. 2018

Viruses

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Bacteriophages SP-15 and ΦW-14 are members of the Myoviridae infecting Bacillus subtilis and Delftia (formerly Pseudomonas) acidovorans, respectively. What links them is that in both cases, approximately 50% of the thymine residues are replaced by hypermodified bases. The consequence of this is that the physico-chemical properties of the DNA are radically altered (melting temperature (Tm), buoyant density and susceptibility to restriction endonucleases). Using 454 pyrosequencing technology, we sequenced the genomes of both viruses. Phage ΦW-14 possesses a 157-kb genome (56.3% GC) specifying 236 proteins, while SP-15 is larger at 222 kb (38.6 mol % G + C) and encodes 318 proteins. In both cases, the phages can be considered genomic singletons since they do not possess BLASTn homologs. While no obvious genes were identified as being responsible for the modified base in ΦW-14, SP-15 contains a cluster of genes obviously involved in carbohydrate metabolism.

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Identification and biosynthesis of thymidine hypermodifications in the genomic DNA of widespread bacterial viruses

Cited by 53 publications

References 67 publications

In vitro Type II Restriction of Bacteriophage DNA With Modified Pyrimidines

In vitro Type II Restriction of Bacteriophage DNA With Modified Pyrimidines

Identification of the minimal bacterial 2′‐deoxy‐7‐amido‐7‐deazaguanine synthesis machinery

The Sequence of Two Bacteriophages with Hypermodified Bases Reveals Novel Phage-Host Interactions

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