Chapter 14 : Transposition in Mycobacteria

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Recombination by transposition occurs without homology between the sequences of the transposon and the target sequence. A large number of different mobile genetic elements have been discovered in prokaryotes. The most basic is known as insertion sequences (ISs). In the cases of Tn5 and Tn10, only one of the ISs can mediate transposition. The chapter focuses on mycobacterial transposable elements. Work on constructing transposons from ISs for use as genetic tools in the mycobacteria has also been initiated. The analysis of cointegrates isolated from different strains each having one copy of Tn670 inserted has demonstrated that transposition occurs randomly with no specific target sites. Transposable elements from mycobacteria are potentially useful sources of genetic tools for the manipulation of mycobacteria in general and for the investigation of virulence mechanisms in pathogenic strains. Useful insertion elements for the development of mutagenesis systems for mycobacteria should (i) have a high frequency of transposition, (ii) not be present in the bacterial strains in which they will be used for mutagenesis, and (iii) exhibit no site or regional specificity. In mycobacteria, nonreplicative vectors have been used to demonstrate the transposition of IS6100, IS900, and IS6110 in . Transposons can be used for purposes other than mutagenesis. Genes of interest can be cloned in them and introduced as a stable single copy into the chromosome. Most ISs isolated from mycobacteria are species specific. Primers have been derived from these sequences and used for identification in polymerase chain reaction tests.

Citation: McAdam R, Guilhot C, Gicquel B. 1994. Transposition in Mycobacteria, p 199-216. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch14
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Image of Figure 1.
Figure 1.

Model (according to Shapiro [1979]) of two different mechanisms of transposition. The three possible outcomes are illustrated. (I) The transposon, represented by a black arrow, is cleaved at its 3' extremities. A staggered cleavage of the target site leads to 5' protruding ends. (The specific cuts are indicated by small arrows.) (II) Joining the 3' ends of the transposon to the 5' ends of the target forms a transposition intermediate called the Shapiro intermediate. This structure can be resolved in the following two ways: (III) specific cleavage of the 5' ends of the transposon and filling in of the target site (white box) leads to a simple insertion of the transposon in the recipient by “conservative transposition,” or (IV) replication of the target site and the transposon by using the recipient as primer (white boxes and dashed arrows) leads to the formation of a cointegrate. This is called “replicative transposition.” (V) This structure can be resolved either by a site-specific resolvase or by the general homologous recombination pathway of the host.

Citation: McAdam R, Guilhot C, Gicquel B. 1994. Transposition in Mycobacteria, p 199-216. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch14
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Image of Figure 2.
Figure 2.

Main ORFs of mycobacterial insertion sequences. Capital letters indicate the putative transposases of the IS. In the elements of the IS3 family (i.e., IS, IS, and ISthe transposase is possibly the result of a fusion of ORFA and ORFB. In the case of IS, ORFA encodes the putative resolvase, and ORFC encodes a putative transposase.

Citation: McAdam R, Guilhot C, Gicquel B. 1994. Transposition in Mycobacteria, p 199-216. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch14
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Image of Figure 3.
Figure 3.

Alignment of the ORFBs of IS, IS1137, IS3, and This alignment was realized by using the Higgins and Sharp (1989) method (PILEUP; GCG, University of Wisconsin). Highly conserved amino acids related to retroviral integrase are underlined. The consensus is indicated when the amino acid is conserved in at least three sequences.

Citation: McAdam R, Guilhot C, Gicquel B. 1994. Transposition in Mycobacteria, p 199-216. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch14
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Image of Figure 4.
Figure 4.

Alignment of the putative transposases of IS, IS, Iand 6. This alignment was realized by using the Higgins and Sharp (1989) method (PILEUP; GCG, University of Wisconsin). The consensus is indicated when the amino acid is conserved in at least three sequences.

Citation: McAdam R, Guilhot C, Gicquel B. 1994. Transposition in Mycobacteria, p 199-216. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch14
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Generic image for table
Table 1.

Main features of mycobacterial ISs

Citation: McAdam R, Guilhot C, Gicquel B. 1994. Transposition in Mycobacteria, p 199-216. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch14
Generic image for table
Table 2.

Auxotrophic types

Citation: McAdam R, Guilhot C, Gicquel B. 1994. Transposition in Mycobacteria, p 199-216. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch14

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