Academic journal article Genetics

Recombination and Annealing Pathways Compete for Substrates in Making Rrn Duplications in Salmonella Enterica

Academic journal article Genetics

Recombination and Annealing Pathways Compete for Substrates in Making Rrn Duplications in Salmonella Enterica

Article excerpt

THE most frequent tandem gene duplications in the Salmonella chromosome arise between directly repeated ribosomal RNA (rrn) loci (Anderson and Roth 1981; Reams et al. 2010). Duplications formed between such extensive repeats (5.5 kb) are generally thought to arise by unequal recombination between copies of the repeat in different sister chromosomes (Roth et al. 1996; Romero and Palacios 1997) (see Figure 1). Such exchanges are expected to depend heavily on the homologous recombination pathways that rely on the strand exchange enzyme RecA. In keeping with this expectation, duplications between chromosomal repeats of the lac operon form at high rates, comparable to those of rrn duplication, but depend heavily on RecA (A. Reams, M. Carter, and J. Roth unpublished results). In contrast, recent assays in two other situations suggest that RecA can be dispensable for formation of duplications between long repeats that are subject to frequent nicks or breaks (Reams et al. 2010, 2012). Duplication of regions not flanked by extensive repeats are also RecA independent, but arise at rates two to three orders of magnitude lower than those for rrn-mediated duplications (Reams and Neidle 2003, 2004; Kugelberg et al. 2006; Reams et al. 2010). Here we describe duplications arising by exchanges between directly repeated rrn loci that are separated by .40 kb and are normal residents of the Salmonella chromosome. The goal is to understand how these duplications arise at such high rates with or without RecA.

Seven rrn loci are scattered around the Salmonella enterica chromosome (Figure 2) and have nearly identical base sequences. Recombination between these loci generates a variety of chromosome rearrangements including duplications, inversions, and translocations (Anderson and Roth 1981; Liu and Sanderson 1998; Helm et al. 2003). Tandem duplications between rrn loci form at very high rates. For example, the argH gene, between rrnA and rrnB (see Figure 2), duplicates at 1.9 3 1023/cell/division (Reams et al. 2010), and this rate is essentially unaltered in recA mutants (Reams et al. 2010). These duplications of argH are lost at a 10-fold higher rate (1022/cell/division) by heavily RecA-dependent recombination between the extensive duplicated regions (.150 kb). Because of their high loss rate and fitness cost, argH duplications are carried as stable polymorphisms in unselected cultures and are maintained at a steady-state frequency of ^1% (Reams et al. 2010). These high steadystate frequencies are likely to be a general property of all duplications in all organisms.

The surprising RecA independence of rrn-mediated duplication formation is examined here using the purHD locus. This region is flanked by the most closely spaced rrn cistrons, rrnB and rrnE (separated by 40 kb), and is held at the highest duplication steady-state frequency (^3%) of any tested point in the chromosome (Anderson and Roth 1981). It is suggested that the high rate and apparent recombination independence of rrn duplications may reflect two features of rrn sequences. First, rrn cistrons are the most highly transcribed genes in the Salmonella and Escherichia coli chromosomes (Dennis 2004). Second, rrn sequences include many stem-loop structures that are responsible for folding of the ribosomal RNAs (16S, 23S, 5S) produced from each locus. The palindromic features of rrn DNA may allow untranscribed strands to form secondary structures that are subject to cutting or breakage. These unusual features may make rrn sequences prone to frequent gaps and breaks.

It is proposed here that blockage of early recombination steps (RecBCDA and RecFORA) can activate a single-strand annealing pathway of duplication formation that compensates for loss of recombination. When frequent DNA damage within rrn sequences remains unrepaired by recombination, these lesions can accumulate sufficiently that two different rrn loci can provide ends. Duplications can form by annealing when two single-strand ends are provided and neither strand is coated with inhibitory RecA protein. …

Search by... Author
Show... All Results Primary Sources Peer-reviewed

Oops!

An unknown error has occurred. Please click the button below to reload the page. If the problem persists, please try again in a little while.