Inversion of Genes in Salmonella
Page 818 Figure 19.18 Proposed molecular pathway for transposition and chromosome rearrangements. Donor DNA, including the transposon, shown in red, recipient DNA contains small light green. The pathway has four steps, beginning with staggered, singlestrand cleavages (Step 1a) at each end of the transposable element and at each end of the "target" nucleotide sequence to be duplicated. The cleavages expose (Step 1b) the DNA strand ends involved in the next step: the joining of DNA strands from donor and recipient molecules in such a way that the doublestranded transposable element has a DNA replication fork at each end (Step 2). DNA synthesis (Step 3) replicates transposon (red bars) and target sequence (light green squares), accounting for the observed duplication. This step forms two new complete doublestranded molecules; each copy of the transposable element joins a segment of the donor molecule and a segment of the recipient molecules. (Copies of the element serve as linkers for the recombination of two unrelated DNA molecules.) In the final Step 4, reciprocal recombination between copies of the transposable element inserts the element at a new genetic site and regenerates the donor molecule. Redrawn from Cohen, S. N., and Shapiro, J. A. Sci. Am. 252:40, 1980. W. H. Freeman and Company, Copyright © 1980. The result is that more and more pathogenic bacterial strains become resistant to an increasing number of antibiotics. 19.7— Inversion of Genes in Salmonella A different mechanism of differential gene regulation has been discovered for one set of genes in Salmonella. Similar control mechanisms exist for the expression of other genes in other prokaryotes (e.g., a bacteriophage called ). Bacteria move by waving their flagella that are composed predominantly of subunits of a protein called flagellin. Many Salmonella species possess two different flagellin genes and express only one of these genes at a time. Bacteria are said to be in phase 1 if they are expressing the H1 flagellin gene and in phase 2 if they are expressing the H2 flagellin gene. A bacterial clone in one phase switches to the other phase about once every 1000 divisions. This switch is called phase variation, and its occurrence is controlled at the level of transcription of H1 and H2 genes. Organization of the flagellin genes and their regulatory elements are shown in Figure 19.19. A 995bp segment of DNA flanked by 14bp repeats is adjacent to the H2 gene and a rhl gene that codes for a repressor of H1. The H2 and Page 819 Figure 19.19 Organization of the flagellin genes of Salmonella. Orientation of a 995bp DNA segment flanked by 14bp inverted repeats (IRL and IRR) controls the expression of H1 and H2 flagellin genes. In phase 2, transcription initiates at promoter P within the invertible segment and continues through H2 and rh1 genes. In phase 1, the orientation is reversed so that transcription of H2 and rh1 genes does not occur. rhl genes are coordinately transcribed. Therefore, when H2 is expressed, the repressor is also made and turns off H1 expression. When H2 protein and the repressor are not made, the H1 gene is derepressed and H1 synthesis occurs. The promoter for the operon containing H2 and rhl lies near one end of the 995bp segment, just inside one copy of the 14bp repeats. This segment can undergo inversions between the 14bp repeats. In one orientation of the segment, the promoter is upstream of the H2–rhl transcription unit; in the other orientation it points toward the opposite direction so that H2 and rhl are not transcribed. In addition to containing this promoter, the invertible segment of DNA possesses the hin gene whose product is an enzyme that catalyzes the inversion event itself. The hin gene seems to be transcribed constitutively at a low rate. Mutations in hin reduce the rate of inversion by 10,000fold. Therefore phase variation is controlled by physical inversion of the segment of DNA that removes a promoter from its position in front of the H2–rhl operon. When the promoter is in the opposite direction, it presumably still initiates transcription, but the fate of that RNA is unknown. It does not initiate transcription of the H1 that maps in this direction. That gene apparently has its own promoter controlled directly by the rhl repressor. Inversion of the hin segment probably occurs via recombination between the 14bp inverted repeats that is similar to recombination events involved in the transposition of a transposon. In fact, transposons do invert relative to their flanking sequences in a fashion exactly analogous to the hin inversion. Furthermore, the amino acid sequence of the hin product shows considerable similarity to that of the tnpR product of the Tn3 transposon, which participates in the integration of the transposon into a new site. Thus it is possible, and even likely, that the two processes are evolutionarily related.