Cassette-like variation of restriction enzyme genes in Escherichia coli C and relatives

Abstract

A surprising result of comparative bacterial genomics has been the large amount of DNA found to be present in one strain but not in another of the same species. We examine in detail one location where gene content varies extensively, the restriction cluster in Escherichia coli. This region is designated the Immigration Control Region (ICR) for the density and variability of restriction functions found there. To better define the boundaries of this variable locus, we determined the sequence of the region from a restrictionless strain, E.coli C. Here we compare the 13.7 kb E.coli C sequence spanning the site of the ICR with corresponding sequences from five E.coli strains and Salmonella typhimurium LT2. To discuss this variation, we adopt the term 'framework' to refer to genes that are stable components of genomes within related lineages, while 'migratory' genes are transient inhabitants of the genome. Strikingly, seven different migratory DNA segments, encoding different sets of genes and gene fragments, alternatively occupy a single well-defined location in the seven strains examined. The flanking framework genes, yjiS and yjiA, display approximately normal patterns of conservation. The patterns observed are consistent with the action of a site-specific recombinase. Since no nearby gene codes for a likely recombinase of known families, such a recombinase must be of a new family or unlinked.

Figure 1

Genetic structure surrounding the ICR in E.coli. Conserved framework genes (striped boxes, approximately to scale) encode hypothetical proteins of unknown function, between 98.4 and 98.9 min on the K-12 chromosome. The variable region (dashed line) found between yjiS and yjiA contains distinct gene sets in different strains. A part of this region has been designated the Immigration Control Region (ICR) in E.coli K-12.

Figure 2

Gene content of the ICR in six strains of E.coli and in Salmonella enterica sv typhimurium LT2. Boxes (not to scale) represent gene coding sequence, even if the gene is truncated. Similarity was assessed at the DNA level; if similarity was >65%, genes were judged orthologous, and named according to the earliest genome sequence. Boxes of the same color represent orthologous blocks of genes that appear to be moving together. Orange boxes: type IA restriction genes (hsdRMS) and mrr. Blue boxes: type IB restriction genes (hsdRMS) and two ORFs of unknown function, Z5949 and Z5950; these are fused in C and CFT073 with a 10 amino acid linker relative to the two in O157:H7. Yellow boxes: yjiW. Green boxes: unknowns Z5943 and Z5944. White boxes: genes unique in this dataset. These are: yjiT, yjiU and mcrD, all of unknown function, in K-12; restriction genes mcrB and mcrC; penicillin acylase (pac) in W; and unknowns STM4522, 4528 and 4529 in S.typhimurium.

Figure 3

DNA sequence alignments at the right (yjiA) border of the ICR. O157:H7 was taken as the reference sequence (black letters) to minimize the amount of color. Differences from this are coded in color hierarchically, and the main goal is to highlight the sudden transition to diversity at the yjiA stop codon. Where K1 differs from O157:H7, the K1 allele is blue (even in other strains); additional differences found in C are in green; further differences found in S.typhimurium are in red; then differences found in W are in yellow and finally remaining differences in K-12 are in pink.

Figure 4

DNA sequence alignments at the left (yjiS) border of the ICR. O157:H7 was taken as the reference sequence (black letters) to minimize the amount of color. Differences from this are coded in color hierarchically as in Figure 3. The sequence of the hsdR fragment found in C is boxed.

Figure 5

Dyad symmetry element in DNA sequence around the proposed empty ‘attachment site’. Dyad symmetry elements can serve as binding sites or sites of action for proteins. Conserved sequence to the left of the yjiS stop codon was joined with sequence to the right of the yjiA stop codon for each strain, and aligned. Stop codons are boxed. The consensus sequence convention is: capital, all sequences identical; lower case: a majority of sequences identical. Mismatches to the consensus are lettered in red. Nucleotides involved in an interrupted dyad symmetry element are marked with a vertical line; mismatches within the dyad with dots.