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Genes involved in conjugative DNA processing of plasmid R6K.
Mol Microbiol 1997; 24(6):1157-68MM

Abstract

The conjugative transfer region of the IncX plasmid R6K (TRA(x)) was analysed by transposon mutagenesis and DNA sequencing. Tn5tac1 insertional mutations localized TRA(x) to a 14.8 kb segment containing the alpha origin of transfer (oriT alpha), genes involved in conjugative DNA-processing (Dtr(x)) and genes involved in pilus synthesis and assembly (Mpf(x)). A second functional oriT, oriTbeta, was located at a distance of 5.3 kb from oriT alpha and was outside TRA(x). Mpf(x) occupied a segment of 10kb, as judged by the location of insertions conferring resistance to infection by the X pilus-specific phage X-2. At both sides of Mpf(x) there were insertions that were Tra but X-2 sensitive, suggesting that the mutations were in Dtr(x). This region was sequenced and three genes were identified: taxA, taxB, and taxC. The overall organization was oriT alpha-taxA-taxC-Mpf(x)-taxB. taxC coded for a oriT-relaxase that belongs to the VirD2 family. taxA coded for a protein of 181 amino acids that showed similarity to TraY of F-like plasmids and to the Arc-repressor superfamily. TaxB showed similarity to TraG-like proteins, a protein superfamily probably involved in coupling the relaxosome to the DNA-transport apparatus. TaxA and TaxC are required for oriT nicking in vivo. The nicking reaction was mistakenly assumed by Flashner et al. (1996) to represent a feature of the vegetative replication origins. However, insertions or deletions disrupting taxA and taxC affected conjugation but not replication of R6K. Conversely, protein pi, which is absolutely required for replication of R6K, was not required for conjugative transfer. In addition, protein DDP3, which is also assumed to have a role in replication, was found to be a positive modulator of bacterial conjugation. Taken together, these results rule out a direct and essential involvement of conjugation proteins in R6K vegetative replication, and also rule out the requirement of replication protein pi for conjugation.

Authors+Show Affiliations

Departamento de Biología Molecular, Universidad de Cantabria, Santander, Spain.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

9218765

Citation

Núñez, B, et al. "Genes Involved in Conjugative DNA Processing of Plasmid R6K." Molecular Microbiology, vol. 24, no. 6, 1997, pp. 1157-68.
Núñez B, Avila P, de la Cruz F. Genes involved in conjugative DNA processing of plasmid R6K. Mol Microbiol. 1997;24(6):1157-68.
Núñez, B., Avila, P., & de la Cruz, F. (1997). Genes involved in conjugative DNA processing of plasmid R6K. Molecular Microbiology, 24(6), pp. 1157-68.
Núñez B, Avila P, de la Cruz F. Genes Involved in Conjugative DNA Processing of Plasmid R6K. Mol Microbiol. 1997;24(6):1157-68. PubMed PMID: 9218765.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Genes involved in conjugative DNA processing of plasmid R6K. AU - Núñez,B, AU - Avila,P, AU - de la Cruz,F, PY - 1997/6/1/pubmed PY - 1997/6/1/medline PY - 1997/6/1/entrez SP - 1157 EP - 68 JF - Molecular microbiology JO - Mol. Microbiol. VL - 24 IS - 6 N2 - The conjugative transfer region of the IncX plasmid R6K (TRA(x)) was analysed by transposon mutagenesis and DNA sequencing. Tn5tac1 insertional mutations localized TRA(x) to a 14.8 kb segment containing the alpha origin of transfer (oriT alpha), genes involved in conjugative DNA-processing (Dtr(x)) and genes involved in pilus synthesis and assembly (Mpf(x)). A second functional oriT, oriTbeta, was located at a distance of 5.3 kb from oriT alpha and was outside TRA(x). Mpf(x) occupied a segment of 10kb, as judged by the location of insertions conferring resistance to infection by the X pilus-specific phage X-2. At both sides of Mpf(x) there were insertions that were Tra but X-2 sensitive, suggesting that the mutations were in Dtr(x). This region was sequenced and three genes were identified: taxA, taxB, and taxC. The overall organization was oriT alpha-taxA-taxC-Mpf(x)-taxB. taxC coded for a oriT-relaxase that belongs to the VirD2 family. taxA coded for a protein of 181 amino acids that showed similarity to TraY of F-like plasmids and to the Arc-repressor superfamily. TaxB showed similarity to TraG-like proteins, a protein superfamily probably involved in coupling the relaxosome to the DNA-transport apparatus. TaxA and TaxC are required for oriT nicking in vivo. The nicking reaction was mistakenly assumed by Flashner et al. (1996) to represent a feature of the vegetative replication origins. However, insertions or deletions disrupting taxA and taxC affected conjugation but not replication of R6K. Conversely, protein pi, which is absolutely required for replication of R6K, was not required for conjugative transfer. In addition, protein DDP3, which is also assumed to have a role in replication, was found to be a positive modulator of bacterial conjugation. Taken together, these results rule out a direct and essential involvement of conjugation proteins in R6K vegetative replication, and also rule out the requirement of replication protein pi for conjugation. SN - 0950-382X UR - https://www.unboundmedicine.com/medline/citation/9218765/Genes_involved_in_conjugative_DNA_processing_of_plasmid_R6K_ L2 - https://onlinelibrary.wiley.com/resolve/openurl?genre=article&sid=nlm:pubmed&issn=0950-382X&date=1997&volume=24&issue=6&spage=1157 DB - PRIME DP - Unbound Medicine ER -