Transposon-mediated mobilization of chromosomally located catabolic operons of the CAM plasmid by TOL plasmid transposon Tn4652 and CAM plasmid transposon Tn3614.

The CAM (camphor degradation) plasmid is integrated into the chromosome of Pseudomonas putida PaW-line strains and is not self-transferable as a plasmid via conjugation. Our results show that the mobilization of chromosomally located CAM and the integration of cam-operons into the chromosome of the new Cam+ transconjugants is a recA-independent process mediated by transposons Tn4652 (17 kbp) and Tn3614 (7.2 kbp). Transposon Tn3614 is apparently identical to the left-hand and the right-hand sequences of the TOL plasmid pWW0 transposon Tn4654. The insertion of Tn401 inside the left-hand terminal IR of Tn4652 completely inhibited the mobilization of CAM. According to our data transposons Tn4652 and Tn3614 together with CAM plasmid catabolic operons are integrated into the chromosome. We propose that in pseudomonads the transposons Tn4652 and Tn3614 play a key role in the evolution and spread of new catabolic plasmids in nature.

Degradation of phenol and m-toluate in Pseudomonas sp. strain EST1001 and its Pseudomonas putida transconjugants is determined by a multiplasmid system.

The utilization of phenol, m-toluate, and salicylate (Phe+, mTol+, and Sal+ characters, respectively) in Pseudomonas sp. strain EST1001 is determined by the coordinated expression of genes placed in different plasmids, i.e., by a multiplasmid system. The natural multiplasmid strain EST1001 is phenotypically unstable. In its Phe-, mTol-, and Sal- segregants, the plasmid DNA underwent structural rearrangements without a marked loss of plasmid DNA, and the majority of segregants gave revertants. The genes specifying the degradation of phenol and m-toluate were transferable to P. putida PaW340, and in this strain a new multiplasmid system with definite structural changes was formed. The 17-kilobase transposable element, a part of the TOL plasmid pWWO present in the chromosome of PaW340, was inserted into the plasmid DNA in transconjugants. In addition, transconjugant EST1020 shared pWWO-like structures. Enzyme assays demonstrated that ortho-fission reactions were used by bacteria that grew on phenol, whereas m-toluate was catabolized by a meta-fission reaction. Salicylate was a functional inducer of the enzymes of both pathways. The expression of silent metabolic pathways of phenol or m-toluate degradation has been observed in EST1001 Phe- mTol+ and Phe+ mTol- transconjugants. The switchover of phenol degradation from the ortho- to the meta-pathway in EST1033 also showed the flexibility of genetic material in EST1001 transconjugants.

Molecular relationships of degradative plasmids determined by in situ hybridisation of their endonuclease-generated fragments.

Plasmid inter-relationships were studied by hybridisation of a radioactively labelled DNA probe to endonuclease-derived fragmentation patterns of plasmids bound to a nitrocellulose filter. The degradative plasmids SAL and NAH were found to be very closely related, but probably one did not give rise to the other by just a single deletion or insertion. Relationships between SAL and other degradative plasmids are complex; substantial homology was found with TOL and other plasmids encoding toluate dissimilation and significant homology was found with OCT.