Sequences and studies of bacteriophage T4 rII mutants

Shinedling, Sidney T.; Singer, Britta Swebilius; Gayle, Margit; Pribnow, David; Jarvis, Erin; Edgar, Bruce A.; Gold, Larry

doi:10.1016/0022-2836(87)90176-8

Cited by 26 publications

(13 citation statements)

References 47 publications

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“…Sequence conversions between similar but ectopic sequences seem to have generated some of the classic rIIB mutants as well (Shinedling et al 1987). One of the base substitutions alters a newly proposed secondary structure element called P9.0, which is in- We propose that either unequal crossing over during recombination or slipped mispairing during replication deleted these 476 bp and one of the 15-bp repeats, to introduce the 491-bp deletion seen in T4.…”

Section: : T G C a C G C G A T G A A C A G C T T C A C C T T A A mentioning

confidence: 99%

The phage T4 nrdB intron: a deletion mutant of a version found in the wild.

Eddy¹,

Gold²

1991

Genes Dev.

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Bacteriophage T4 possesses three self-splicing group I introns. Two of the three introns are mobile elements; the third, in the gene encoding a subunit of the phage nucleotide reductase (nrdB), is not mobile. Because intron mobility offers a reasonable explanation for the paradoxical occurrence of large intervening sequences in a space-efficient eubacterial phage, it is puzzling that the nrdB intron is not mobile like its compatriots. We have discovered a larger nrdB intron in a closely related phage, and we infer from comparative sequence data that the T4 intron is a deletion mutant derived from this larger intron. This larger nrdB intron encodes an open reading frame of 269 codons, which we have cloned and overexpressed. The overexpressed protein shows a dsDNA endonuclease activity specific for the intronless nrdB gene, typical of mobile introns. Thus, we believe that all three introns of T4 are or were mobile "infectious introns" and that they have entered into and been maintained in the phage population by virtue of this efficient mobility.[Key Words: Bacteriophage T4; nrdB intron; endonuclease mobility]

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Section: : T G C a C G C G A T G A A C A G C T T C A C C T T A A mentioning

confidence: 99%

The phage T4 nrdB intron: a deletion mutant of a version found in the wild.

Eddy¹,

Gold²

1991

Genes Dev.

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“…Lynn Ripley (1982) described two models for templated mutagenesis based on imperfect palindromic repeats, in one of which the ectopic template was in the other parental strand and in the other of which the ectopic template was in the daughter strand itself. Examination of other mutations added imperfect direct repeats as mutagenic templates (Ripley et al 1986;Shinedling et al 1987). Templated mutations are usually invoked when they simultaneously acquire multiple changes for which a plausible donor template can be found.…”

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confidence: 99%

Templated Mutagenesis in Bacteriophage T4 Involving Imperfect Direct or Indirect Sequence Repeats

Schultz

Drake

2008

Genetics

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Some mutations arise in association with a potential sequence donor that consists of an imperfect direct or reverse repeat. Many such mutations are complex; that is, they consist of multiple close sequence changes. Current models posit that the primer terminus of a replicating DNA molecule dissociates, reanneals with an ectopic template, extends briefly, and then returns to the cognate template, bringing with it a locally different sequence; alternatively, a hairpin structure may form the mutational intermediate when processed by mismatch repair. This process resembles replication repair, in which primer extension is blocked by a lesion in the template; in this case, the ectopic template is the other daughter strand, and the result is errorfree bypass of the lesion. We previously showed that mutations that impair replication repair can enhance templated mutagenesis. We show here that the intensity of templated mutation can be exquisitely sensitive to its local sequence, that the donor and recipient arms of an imperfect inverse repeat can exchange roles, and that double mutants carrying two alleles, each affecting both templated mutagenesis and replication repair, can have unexpected phenotypes. We also record an instance in which the mutation rates at two particular sites change concordantly with a distant sequence change, but in a manner that appears unrelated to templated mutagenesis. T EMPLATED mutations are initiated when a DNA primer strand dissociates from its cognate template and anneals with a fragment of complementary sequence in an ectopic template. The relocated primer may be extended, acquiring a short sequence that is not fully complementary to the original template, and then reanneal with its cognate template. If the acquired noncomplementary bases escape correctly oriented proofreading and mismatch repair, mutations will result. Lynn Ripley (1982) described two models for templated mutagenesis based on imperfect palindromic repeats, in one of which the ectopic template was in the other parental strand and in the other of which the ectopic template was in the daughter strand itself. Examination of other mutations added imperfect direct repeats as mutagenic templates (Ripley et al. 1986;Shinedling et al. 1987). Templated mutations are usually invoked when they simultaneously acquire multiple changes for which a plausible donor template can be found. Despite the intrinsic fascination of templated complex mutations and their potential importance for certain evolutionary paths and some human genetic disorders, the enzymology that creates them has not been characterized.Template switching also occurs in a mutation-avoiding mode called replication repair. In the canonical model for replication repair (Fujiwara and Tatsumi 1976;Higgins et al. 1976), a primer strand whose extension has been blocked by DNA damage switches to the other daughter strand as a template, extends briefly, and then switches back to its cognate template, thus accurately bypassing the damaged site.In 1987, a mechanism for su...

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“…T4 strain BAS4 is a double rII mutant, carrying the UV294 allele in rIIA (deletion of an extra T in run of T's at position 469-473 bp of the T4 genome) and the UV232 allele in rIIB (insertion of extra T in run of T's at position 168,468-168,469 bp). T4 strain FC11 harbors a frameshift mutation in rIIB resulting from deletion of one T in a run of T's at position 168,875-168,879 (Shinedling et al 1987). T4 strains used for plasmid DNA replication experiments are derivatives of strain K10, which carries the following mutations: amB262 (gene 38), amS29 (gene 51), nd28 (denA), and rIIPT8 (denB-rII D ) (Selick et al 1988).…”

Section: Strainsmentioning

confidence: 99%

Coordination and Processing of DNA Ends During Double-Strand Break Repair: The Role of the Bacteriophage T4 Mre11/Rad50 (MR) Complex

et al. 2013

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The in vivo functions of the bacteriophage T4 Mre11/Rad50 (MR) complex (gp46/47) in double-strand-end processing, double-strand break repair, and recombination-dependent replication were investigated. The complex is essential for T4 growth, but we wanted to investigate the in vivo function during productive infections. We therefore generated a suppressed triple amber mutant in the Rad50 subunit to substantially reduce the level of complex and thereby reduce phage growth. Growth-limiting amounts of the complex caused a concordant decrease in phage genomic recombination-dependent replication. However, the efficiencies of doublestrand break repair and of plasmid-based recombination-dependent replication remained relatively normal. Genetic analyses of linked markers indicated that double-strand ends were less protected from nuclease erosion in the depleted infection and also that end coordination during repair was compromised. We discuss models for why phage genomic recombination-dependent replication is more dependent on Mre11/Rad50 levels when compared to plasmid recombination-dependent replication. We also tested the importance of the conserved histidine residue in nuclease motif I of the T4 Mre11 protein. Substitution with multiple different amino acids (including serine) failed to support phage growth, completely blocked plasmid recombination-dependent replication, and led to the stabilization of double-strand ends. We also constructed and expressed an Mre11 mutant protein with the conserved histidine changed to serine. The mutant protein was found to be completely defective for nuclease activities, but retained the ability to bind the Rad50 subunit and double-stranded DNA. These results indicate that the nuclease activity of Mre11 is critical for phage growth and recombination-dependent replication during T4 infections.

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Sequences and studies of bacteriophage T4 rII mutants

Cited by 26 publications

References 47 publications

The phage T4 nrdB intron: a deletion mutant of a version found in the wild.

The phage T4 nrdB intron: a deletion mutant of a version found in the wild.

Templated Mutagenesis in Bacteriophage T4 Involving Imperfect Direct or Indirect Sequence Repeats

Coordination and Processing of DNA Ends During Double-Strand Break Repair: The Role of the Bacteriophage T4 Mre11/Rad50 (MR) Complex

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