The Bacillus subtilis AddAB helicase/nuclease is regulated by its cognate Chi sequence in vitro

Chédin, Frédéric; Ehrlich, S. Dusko; Kowalczykowski, Stephen C.

doi:10.1006/jmbi.2000.3556

Cited by 77 publications

(82 citation statements)

References 48 publications

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“…We asked whether replisomes were also required for RecA-GFP focus formation in response to DSBs. It has been shown in vitro that the B. subtilis AddAB enzyme can process a DSB, resulting in ssDNA that might be used for RecA filament formation (36)(37)(38). Because, unlike E. coli, B. subtilis contains five epistasis groups for homologous recombination, other enzymes likely contribute to DSB processing (21).…”

Section: The Subcellular Position Of Reca-gfp Foci Is Consistent Withmentioning

confidence: 99%

Replication is required for the RecA localization response to DNA damage in Bacillus subtilis

Simmons

Grossman

Walker

2007

Proc. Natl. Acad. Sci. U.S.A.

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In both prokaryotes and eukaryotes, proteins involved in DNA repair often organize into multicomponent complexes that can be visualized as foci in living cells. We used a RecA-GFP fusion to examine the subcellular cues that direct RecA-GFP to assemble as foci in response to DNA damage. We used two different methods to inhibit initiation of DNA replication and determined that DNA replication is required for the cell to establish RecA-GFP foci after exposure to DNA-damaging agents. Furthermore, use of endonuclease cleavage to generate a site-specific double-strand break demonstrated that the replication machinery (replisome) and DNA synthesis are required for assembly of RecA-GFP foci during repair of a double-strand break. We monitored the cellular levels of RecA and found that focus formation does not require further induction of protein levels, suggesting that foci result from a redistribution of existing protein to sites of damage encountered by the replisome. Taken together, our results support the model that existing RecA protein is recruited to ssDNA generated by the replisome at sites of DNA damage. These results provide insight into the mechanisms that the cell uses to recruit repair proteins to damaged DNA in living cells.replisome ͉ focus ͉ double-strand break

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Section: The Subcellular Position Of Reca-gfp Foci Is Consistent Withmentioning

confidence: 99%

Replication is required for the RecA localization response to DNA damage in Bacillus subtilis

Simmons

Grossman

Walker

2007

Proc. Natl. Acad. Sci. U.S.A.

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“…-like sites have been identified in two Gram-positive bacteria, Lactococcus lactis and Bacillus subtilis. Similarly to Ec , Ll and Bs modify the activities of RecBCD-like enzymes (namely RexAB and AddAB, respectively in L. lactis and B. subtilis), although it has not been shown that they constitute recombination hotspots in these species (16,17). Interestingly, both Ll (5Ј-GCGCGTG-3Ј) and Bs (5Ј-AGCGG-3Ј) are also GC-rich sequences.…”

Section: Structure Of Njs: Presence Of Short Stretches Of Identity Atmentioning

confidence: 99%

Homologous recombination at the border: Insertion-deletions and the trapping of foreign DNA in Streptococcus pneumoniae

Prudhomme

Libante

Claverys

2002

Proc. Natl. Acad. Sci. U.S.A.

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Integration of foreign DNA was observed in the Gram-positive human pathogen Streptococcus pneumoniae (pneumococcus) after transformation with DNA from a recombinant Escherichia coli bacteriophage carrying a pneumococcal insert. Segments of DNA replaced chromosomal sequences adjacent to the region homologous with the pneumococcal insert, whence the name insertion-deletion. Here we report that a pneumococcal insert was absolutely required for insertion-deletion formation, but could be as short as 153 bp; that the sizes of foreign DNA insertions (289 -2,474 bp) and concomitant chromosomal deletions (45-1,485 bp) were not obviously correlated; that novel joints clustered preferentially within segments of high GC content; and that the crossovers in 29 independent novel joints were located 1 bp from the border or within short (3-10 nt long) stretches of identity (microhomology) between resident and foreign DNA. The data are consistent with a model in which the insert serving as a homologous recombination anchor favors interaction and subsequent illegitimate recombination events at microhomologies between foreign and resident sequences. The potential of homologydirected illegitimate recombination for genome evolution was illustrated by the trapping of functional heterologous genes. G enetic transformation, which was discovered in the Grampositive Streptococcus pneumoniae (pneumococcus) (1), is believed to play a central role in the biology of this human pathogen through its contribution to genetic plasticity (see ref.2 for a review). Transformation with naked DNA allows intraspecies and interspecies gene transfer. As such exchanges involve homologous recombination, they are generally ''conservative,'' i.e., they do not result in the creation of novel sequences but simply in a redistribution of previously existing genes. However, a pre-existing gene also can be modified when the two interacting sequences are partly divergent. Homologous recombination then leads to the production of mosaic genes, as exemplified in the case of the pbp genes of S. pneumoniae that encode altered penicillin-binding proteins with decreased affinity for ␤-lactam antibiotics (see ref. 3 for a review).Besides these conservative facets of transformation, is there any potential for the creation of novel sequence combinations or genes? The observation that transformation with chimeric donor DNA is mutagenic (4) suggested that shuffling and reassembly of previously unrelated sequences could readily occur in S. pneumoniae. The chimeric DNA extracted from a recombinant Escherichia coli bacteriophage carrying a pneumococcal insert produced illegitimate recombinants at a frequency of about 0.5% that of homologous recombinants (4). Illegitimate recombinants resulted from simultaneous insertion of heterologous vector (i.e., ) sequences and deletion of chromosomal sequences adjacent to the region homologous to the insert. These illegitimate events were therefore termed insertion-deletions (InsDels). As the pneumococcal insert in the chimeric donor seemed r...

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“…In Bacillus subtilis the recombination genes, other than recA, which is central to all pathways of recombinational repair, have been placed into six different epistatic groups: a [comprising recF, recL, recO, and recR (recFLOR) and recN], b (addA and addB), g (recP and recH), e (ruvA, ruvB, recD, and recU), z (recS), and h (recG) (Alonso et al 1993;Fernandez et al 1998Fernandez et al , 1999Fernandez et al , 2000Chedin et al 2000;Ayora et al 2004;Carrasco et al 2004). Throughout this article, unless stated otherwise, the indicated genes and products refer to those of B. subtilis origin.…”

mentioning

confidence: 99%

“…In wild-type cells, the loading of RecA protein onto SSBcoated ssDNA (presynaptic step) relies on AddAB or RecN-RecFLOR proteins (Chedin et al 2000;Kidane et al 2004). Recently it has been shown that: (i) 35% of the cells in a DrecA and 5% in a DrecU mutant contain unrepaired DSBs under normal growth conditions (Kidane et al 2004); (ii) the ruvA gene complements the defect of the recB2 mutation classified within the e epistatic group [hence recB2 was renamed ruvA2 ]; (iii) purified RecU protein binds preferentially to three-and four-strand junctions and cleaves Holliday junction substrates to produce nicked duplexes and (iv) in the absence of the RuvAB or RecG branch-migration activities, RecU and the poorly characterized RecD bias exchange toward crossovers (CO) .…”

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confidence: 99%

The RuvAB Branch Migration Translocase and RecU Holliday Junction Resolvase Are Required for Double-Stranded DNA Break Repair in Bacillus subtilis

et al. 2005

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In models of Escherichia coli recombination and DNA repair, the RuvABC complex directs the branch migration and resolution of Holliday junction DNA. To probe the validity of the E. coli paradigm, we examined the impact of mutations in DruvAB and DrecU (a ruvC functional analog) on DNA repair. Under standard transformation conditions we failed to construct DruvAB DrecG, DrecU DruvAB, DrecU DrecG, or DrecU DrecJ strains. However, DruvAB could be combined with addAB (recBCD), recF, recH, DrecS, DrecQ, and DrecJ mutations. The DruvAB and DrecU mutations rendered cells extremely sensitive to DNA-damaging agents, although less sensitive than a DrecA strain. When damaged cells were analyzed, we found that RecU was recruited to defined double-stranded DNA breaks (DSBs) and colocalized with RecN. RecU localized to these centers at a later time point during DSB repair, and formation was dependent on RuvAB. In addition, expression of RecU in an E. coli ruvC mutant restored full resistance to UV light only when the ruvAB genes were present. The results demonstrate that, as with E. coli RuvABC, RuvAB targets RecU to recombination intermediates and that all three proteins are required for repair of DSBs arising from lesions in chromosomal DNA.

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The Bacillus subtilis AddAB helicase/nuclease is regulated by its cognate Chi sequence in vitro

Cited by 77 publications

References 48 publications

Replication is required for the RecA localization response to DNA damage in Bacillus subtilis

Replication is required for the RecA localization response to DNA damage in Bacillus subtilis

Homologous recombination at the border: Insertion-deletions and the trapping of foreign DNA in Streptococcus pneumoniae

The RuvAB Branch Migration Translocase and RecU Holliday Junction Resolvase Are Required for Double-Stranded DNA Break Repair in Bacillus subtilis

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