Conjugative transposition of Tn916: the transposon int gene is required only in the donor

Abstract

Conjugative transposition of transposon Tn916 has been shown to proceed by excision of the transposon in the donor strain and insertion of this element in the recipient. This process requires the product of the transposon int gene. We report here the surprising finding that the int gene is required only in the donor during conjugative transposition. We find that Tn916 int-l, whose int gene has been inactivated by an insertion mutation, transposes when a complementing wild-type int gene is present only in the donor during mating. When the int+ gene is present in a plasmid and is expressed from the spac promoter, conjugative transposition is very inefficient. However, when the Int+ function is supplied from a coresident distantly linked Tn916 tra-641 mutant, which is defective in a function required for conjugation, efficient conjugative transposition of Tn916 int-i occurs. This suggests either that Int is not required for integration of Tn916 in gram-positive bacteria or that the protein is transferred from the donor to the transconjugant during the mating event. When the nonconjugative plasmid pAT145 was present in the donor, it was rarely cotransferred with Tn916. This suggests that complete fusion of mating cells is not common during conjugative transposition.Conjugative transposons promote bacterial conjugation, during which they excise from a donor and then insert into a new target in the recipient genome (recent reviews include references 6, 21, 22, and 29). All such transposons share regions of DNA homology and carry the tetM gene, a determinant of tetracyline resistance which is expressed in bacteria of many different genera (6). The promiscuity of conjugative transposons is an important clinical problem, since they are transferred among unrelated gram-positive bacteria and serve as vectors for the spread of antibiotic resistance. Although these elements do not promote conjugation between Escherichia coli strains (7, 11), they do transpose in this organism. These elements integrate into many different sites in the genome of their hosts, and multiple transposons are often found in transconjugants (10). Furthermore, the presence of a conjugative transposon in the recipient strain in mating does not affect the frequency of integration of a second such element (8, 16). These properties make conjugative transposons valuable for mutagenesis, especially in gram-positive bacteria.The best-studied conjugative transposons are the 16.4-kb Tn916 (9) and TnI545, a 25.3-kb transposon. In addition to tetM, the latter also carries antibiotic resistance determinants for erythromycin and kanamycin (3). The terminal 250 bp of these two elements are essentially identical, and they have similar restriction maps.

Control of SXT Integration and Excision

“…The requirement for SetCD in recipient cells to mediate integration of these plasmids suggests that SetCDactivated de novo expression of int from SXT transferred into recipient cells is required for SXT integration into the chromosome. Since it was previously shown that int is required in both donor and recipient cells for SXT transfer (17), such a requirement for int expression in recipient cells differs from the requirement for Tn916, where Int is thought to be transported along with Tn916 DNA through the mating pore into recipient cells (8). Our findings may suggest that setC and setD are transcribed very soon after SXT enters recipient cells.…”

NBU1, a mobilizable site-specific integrated element from Bacteroides spp., can integrate nonspecifically in Escherichia coli

“…Bringel et al (4) reported that Tn916 integrase produced in the donor was transferred to the recipient with the transposon and was sufficient for integration in the recipient, although the frequency of integration was over 1,000-fold lower than it was if the integrase was provided in the recipient. In this case, an integrase-defective derivative of Tn916Kan was inserted in the chromosome and the Tn916 integrase was provided in trans on a plasmid or on a wild-type Tn916(Tc).…”

Xis Protein Binding to the Left Arm Stimulates Excision of Conjugative Transposon Tn916

“…The transcription of the Tn916 Xis and Tn916 Int proteins is coordinately regulated (11), suggesting that within the donor, equal amounts of Tn916 Xis and Tn916 Int could function to promote excision, while the presence of Tn916 Xis could prevent reintegration. Since the Tn916 Int protein is only required to be produced in the donor cell (5) and is presumably transferred to the new recipient cell along with the transposon, the absence of Tn916 Xis in the recipient would then enable the integrative pathway to dominate.…”