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Journal of Bacteriology, March 2000, p. 1541-1548, Vol. 182, No. 6
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

TraG from RP4 and TraG and VirD4 from Ti Plasmids Confer Relaxosome Specificity to the Conjugal Transfer System of pTiC58

Claire M. Hamilton,^1, Hyewon Lee,² Pei-Li Li,¹ David M. Cook,^1, Kevin R. Piper,³ Susanne Beck von Bodman,^1,§ Erich Lanka,⁴ Walt Ream,² and Stephen K. Farrand^1,3,*

Departments of Crop Sciences¹ and Microbiology,³ University of Illinois at Urbana-Champaign, Urbana, Illinois 61801; Department of Microbiology, Oregon State University, Corvallis, Oregon 97331²; and Max-Planck-Institut für Molekulare Genetik, Dahlem, D-14195 Berlin, Germany⁴

Received 26 August 1999/Accepted 21 December 1999

Plasmid conjugation systems are composed of two components, the DNA transfer and replication system, or Dtr, and the mating pair formation system, or Mpf. During conjugal transfer an essential factor, called the coupling protein, is thought to interface the Dtr, in the form of the relaxosome, with the Mpf, in the form of the mating bridge. These proteins, such as TraG from the IncP1 plasmid RP4 (TraG_RP4) and TraG and VirD4 from the conjugal transfer and T-DNA transfer systems of Ti plasmids, are believed to dictate specificity of the interactions that can occur between different Dtr and Mpf components. The Ti plasmids of Agrobacterium tumefaciens do not mobilize vectors containing the oriT of RP4, but these IncP1 plasmid derivatives lack the trans-acting Dtr functions and TraG_RP4. A. tumefaciens donors transferred a chimeric plasmid that contains the oriT and Dtr genes of RP4 and the Mpf genes of pTiC58, indicating that the Ti plasmid mating bridge can interact with the RP4 relaxosome. However, the Ti plasmid did not mobilize transfer from an IncQ relaxosome. The Ti plasmid did mobilize such plasmids if TraG_RP4 was expressed in the donors. Mutations in traG_RP4 with defined effects on the RP4 transfer system exhibited similar phenotypes for Ti plasmid-mediated mobilization of the IncQ vector. When provided with VirD4, the tra system of pTiC58 mobilized plasmids from the IncQ relaxosome. However, neither TraG_RP4 nor VirD4 restored transfer to a traG mutant of the Ti plasmid. VirD4 also failed to complement a traG_RP4 mutant for transfer from the RP4 relaxosome or for RP4-mediated mobilization from the IncQ relaxosome. TraG_RP4-mediated mobilization of the IncQ plasmid by pTiC58 did not inhibit Ti plasmid transfer, suggesting that the relaxosomes of the two plasmids do not compete for the same mating bridge. We conclude that TraG_RP4 and VirD4 couples the IncQ but not the Ti plasmid relaxosome to the Ti plasmid mating bridge. However, VirD4 cannot couple the IncP1 or the IncQ relaxosome to the RP4 mating bridge. These results support a model in which the coupling proteins specify the interactions between Dtr and Mpf components of mating systems.

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^* Corresponding author. Mailing address: Department of Crop Sciences, University of Illinois at Urbana-Champaign, 240 Edward R. Madigan Laboratory (ERML), 1201 West Gregory Dr., Urbana, IL 61801. Phone: (217) 333-1524. Fax: (217) 244-7830. E-mail: stephenf{at}uiuc.edu.

Present address: Clontech, Inc., Palo Alto, CA 94303.

Present address: MJ Research, Inc., Indian Village, NV 89451.

^§ Present address: Department of Plant Science, University of Connecticut, Storrs, CT 06269.

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Journal of Bacteriology, March 2000, p. 1541-1548, Vol. 182, No. 6
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

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