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Journal of Bacteriology, April 2000, p. 2184-2190, Vol. 182, No. 8
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Type IV Pilus Genes pilA and pilC of Pseudomonas stutzeri Are Required for Natural Genetic Transformation, and pilA Can Be Replaced by Corresponding Genes from Nontransformable Species

Stefan Graupner,1 Verena Frey,1 Rozita Hashemi,1 Michael G. Lorenz,2 Gudrun Brandes,3 and Wilfried Wackernagel1,*

AG Genetik, Fachbereich Biologie, Universität Oldenburg, D-26111 Oldenburg,1 Marine Mikrobiologie, Fachbereich Biologie/Chemie, Zentrum für Umweltforschung und Technologie, Universität Bremen, D-28359 Bremen,2 and Medizinische Hochschule Hannover, Abteilung Zellbiologie, D-30625 Hannover,3 Germany

Received 5 October 1999/Accepted 27 January 2000

Pseudomonas stutzeri lives in terrestrial and aquatic habitats and is capable of natural genetic transformation. After transposon mutagenesis, transformation-deficient mutants were isolated from a P. stutzeri JM300 strain. In one of them a gene which coded for a protein with 75% amino acid sequence identity to PilC of Pseudomonas aeruginosa, an accessory protein for type IV pilus biogenesis, was inactivated. The presence of type IV pili was demonstrated by susceptibility to the type IV pilus-dependent phage PO4, by occurrence of twitching motility, and by electron microscopy. The pilC mutant had no pili and was defective in twitching motility. Further sequencing revealed that pilC is clustered in an operon with genes homologous to pilB and pilD of P. aeruginosa, which are also involved in pilus formation. Next to these genes but transcribed in the opposite orientation a pilA gene encoding a protein with high amino acid sequence identity to pilin, the structural component of type IV pili, was identified. Insertional inactivation of pilA abolished pilus formation, PO4 plating, twitching motility, and natural transformation. The amounts of 3H-labeled P. stutzeri DNA that were bound to competent parental cells and taken up were strongly reduced in the pilC and pilA mutants. Remarkably, the cloned pilA genes from nontransformable organisms like Dichelobacter nodosus and the PAK and PAO strains of P. aeruginosa fully restored pilus formation and transformability of the P. stutzeri pilA mutant (along with PO4 plating and twitching motility). It is concluded that the type IV pili of the soil bacterium P. stutzeri function in DNA uptake for transformation and that their role in this process is not confined to the species-specific pilin.

* Corresponding author. Mailing address: AG Genetik, Fachbereich Biologie, Universität Oldenburg, Postfach 2503, D-26111 Oldenburg, Germany. Phone: 49-(0)441-7983298. Fax: 49-(0)441-7985606. E-mail: genetics{at}biologie.uni-oldenburg.de.

Journal of Bacteriology, April 2000, p. 2184-2190, Vol. 182, No. 8
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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