Cargando…

Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster

Lactococcus lactis MG1363 is an important gram-positive model organism. It is a plasmid-free and phage-cured derivative of strain NCDO712. Plasmid-cured strains facilitate studies on molecular biological aspects, but many properties which make L. lactis an important organism in the dairy industry ar...

Descripción completa

Detalles Bibliográficos
Autores principales: Tarazanova, Mariya, Beerthuyzen, Marke, Siezen, Roland, Fernandez-Gutierrez, Marcela M., de Jong, Anne, van der Meulen, Sjoerd, Kok, Jan, Bachmann, Herwig
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5152845/
https://www.ncbi.nlm.nih.gov/pubmed/27941999
http://dx.doi.org/10.1371/journal.pone.0167970
_version_ 1782474631046758400
author Tarazanova, Mariya
Beerthuyzen, Marke
Siezen, Roland
Fernandez-Gutierrez, Marcela M.
de Jong, Anne
van der Meulen, Sjoerd
Kok, Jan
Bachmann, Herwig
author_facet Tarazanova, Mariya
Beerthuyzen, Marke
Siezen, Roland
Fernandez-Gutierrez, Marcela M.
de Jong, Anne
van der Meulen, Sjoerd
Kok, Jan
Bachmann, Herwig
author_sort Tarazanova, Mariya
collection PubMed
description Lactococcus lactis MG1363 is an important gram-positive model organism. It is a plasmid-free and phage-cured derivative of strain NCDO712. Plasmid-cured strains facilitate studies on molecular biological aspects, but many properties which make L. lactis an important organism in the dairy industry are plasmid encoded. We sequenced the total DNA of strain NCDO712 and, contrary to earlier reports, revealed that the strain carries 6 rather than 5 plasmids. A new 50-kb plasmid, designated pNZ712, encodes functional nisin immunity (nisCIP) and copper resistance (lcoRSABC). The copper resistance could be used as a marker for the conjugation of pNZ712 to L. lactis MG1614. A genome comparison with the plasmid cured daughter strain MG1363 showed that the number of single nucleotide polymorphisms that accumulated in the laboratory since the strains diverted more than 30 years ago is limited to 11 of which only 5 lead to amino acid changes. The 16-kb plasmid pSH74 was found to contain a novel 8-kb pilus gene cluster spaCB-spaA-srtC1-srtC2, which is predicted to encode a pilin tip protein SpaC, a pilus basal subunit SpaB, and a pilus backbone protein SpaA. The sortases SrtC1/SrtC2 are most likely involved in pilus polymerization while the chromosomally encoded SrtA could act to anchor the pilus to peptidoglycan in the cell wall. Overexpression of the pilus gene cluster from a multi-copy plasmid in L. lactis MG1363 resulted in cell chaining, aggregation, rapid sedimentation and increased conjugation efficiency of the cells. Electron microscopy showed that the over-expression of the pilus gene cluster leads to appendices on the cell surfaces. A deletion of the gene encoding the putative basal protein spaB, by truncating spaCB, led to more pilus-like structures on the cell surface, but cell aggregation and cell chaining were no longer observed. This is consistent with the prediction that spaB is involved in the anchoring of the pili to the cell.
format Online
Article
Text
id pubmed-5152845
institution National Center for Biotechnology Information
language English
publishDate 2016
publisher Public Library of Science
record_format MEDLINE/PubMed
spelling pubmed-51528452016-12-28 Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster Tarazanova, Mariya Beerthuyzen, Marke Siezen, Roland Fernandez-Gutierrez, Marcela M. de Jong, Anne van der Meulen, Sjoerd Kok, Jan Bachmann, Herwig PLoS One Research Article Lactococcus lactis MG1363 is an important gram-positive model organism. It is a plasmid-free and phage-cured derivative of strain NCDO712. Plasmid-cured strains facilitate studies on molecular biological aspects, but many properties which make L. lactis an important organism in the dairy industry are plasmid encoded. We sequenced the total DNA of strain NCDO712 and, contrary to earlier reports, revealed that the strain carries 6 rather than 5 plasmids. A new 50-kb plasmid, designated pNZ712, encodes functional nisin immunity (nisCIP) and copper resistance (lcoRSABC). The copper resistance could be used as a marker for the conjugation of pNZ712 to L. lactis MG1614. A genome comparison with the plasmid cured daughter strain MG1363 showed that the number of single nucleotide polymorphisms that accumulated in the laboratory since the strains diverted more than 30 years ago is limited to 11 of which only 5 lead to amino acid changes. The 16-kb plasmid pSH74 was found to contain a novel 8-kb pilus gene cluster spaCB-spaA-srtC1-srtC2, which is predicted to encode a pilin tip protein SpaC, a pilus basal subunit SpaB, and a pilus backbone protein SpaA. The sortases SrtC1/SrtC2 are most likely involved in pilus polymerization while the chromosomally encoded SrtA could act to anchor the pilus to peptidoglycan in the cell wall. Overexpression of the pilus gene cluster from a multi-copy plasmid in L. lactis MG1363 resulted in cell chaining, aggregation, rapid sedimentation and increased conjugation efficiency of the cells. Electron microscopy showed that the over-expression of the pilus gene cluster leads to appendices on the cell surfaces. A deletion of the gene encoding the putative basal protein spaB, by truncating spaCB, led to more pilus-like structures on the cell surface, but cell aggregation and cell chaining were no longer observed. This is consistent with the prediction that spaB is involved in the anchoring of the pili to the cell. Public Library of Science 2016-12-12 /pmc/articles/PMC5152845/ /pubmed/27941999 http://dx.doi.org/10.1371/journal.pone.0167970 Text en © 2016 Tarazanova et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Tarazanova, Mariya
Beerthuyzen, Marke
Siezen, Roland
Fernandez-Gutierrez, Marcela M.
de Jong, Anne
van der Meulen, Sjoerd
Kok, Jan
Bachmann, Herwig
Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster
title Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster
title_full Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster
title_fullStr Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster
title_full_unstemmed Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster
title_short Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster
title_sort plasmid complement of lactococcus lactis ncdo712 reveals a novel pilus gene cluster
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5152845/
https://www.ncbi.nlm.nih.gov/pubmed/27941999
http://dx.doi.org/10.1371/journal.pone.0167970
work_keys_str_mv AT tarazanovamariya plasmidcomplementoflactococcuslactisncdo712revealsanovelpilusgenecluster
AT beerthuyzenmarke plasmidcomplementoflactococcuslactisncdo712revealsanovelpilusgenecluster
AT siezenroland plasmidcomplementoflactococcuslactisncdo712revealsanovelpilusgenecluster
AT fernandezgutierrezmarcelam plasmidcomplementoflactococcuslactisncdo712revealsanovelpilusgenecluster
AT dejonganne plasmidcomplementoflactococcuslactisncdo712revealsanovelpilusgenecluster
AT vandermeulensjoerd plasmidcomplementoflactococcuslactisncdo712revealsanovelpilusgenecluster
AT kokjan plasmidcomplementoflactococcuslactisncdo712revealsanovelpilusgenecluster
AT bachmannherwig plasmidcomplementoflactococcuslactisncdo712revealsanovelpilusgenecluster