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Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C

Rickettsia bellii is an obligate intracellular bacterium that is one of the few rickettsiae that encode a complete set of conjugative transfer (tra) genes involved in bacterial conjugation and has been shown to exhibit pili-like structures. The reductive genomes of rickettsiae beg the question wheth...

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Autores principales: Heu, Chan C., Kurtti, Timothy J., Nelson, Curtis M., Munderloh, Ulrike G.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4564193/
https://www.ncbi.nlm.nih.gov/pubmed/26352829
http://dx.doi.org/10.1371/journal.pone.0137214
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author Heu, Chan C.
Kurtti, Timothy J.
Nelson, Curtis M.
Munderloh, Ulrike G.
author_facet Heu, Chan C.
Kurtti, Timothy J.
Nelson, Curtis M.
Munderloh, Ulrike G.
author_sort Heu, Chan C.
collection PubMed
description Rickettsia bellii is an obligate intracellular bacterium that is one of the few rickettsiae that encode a complete set of conjugative transfer (tra) genes involved in bacterial conjugation and has been shown to exhibit pili-like structures. The reductive genomes of rickettsiae beg the question whether the tra genes are nonfunctional or functioning to enhance the genetic plasticity and biology of rickettsiae. We characterized the transcriptional dynamics of R. bellii tra genes in comparison to genes transcribed stably and above the background level to understand when and at what levels the tra genes are active or whether the tra genes are degenerative. We determined that the best reference genes, out of 10 tested, were methionyl tRNA ligase (metG) or a combination of metG and ribonucleoside diphosphate reductase 2 subunit beta (nrdF), using statistical algorithms from two different programs: Normfinder and BestKeeper. To validate the use of metG with other rickettsial genes exhibiting variable transcriptional patterns we examined its use with sca2 and rickA, genes involved in actin based motility. Both were shown to be up-regulated at different times of replication in Vero cells, showing variable and stable transcription levels of rickA and sca2, respectively. traA (Ti) was up-regulated at 72 hours post inoculation in the tick cell line ISE6, but showed no apparent changes in the monkey cell line Vero and mouse cell line L929. The transcription of tra genes was positively correlated with one another and up-regulated from 12 to 72 hours post inoculation (HPI) when compared to RBE_0422 (an inactivated transposase-derivative found within the tra cluster). Thus, the up-regulation of the tra genes indicated that the integrity and activity of each gene were intact and may facilitate the search for the optimal conditions necessary to demonstrate conjugation in rickettsiae.
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spelling pubmed-45641932015-09-17 Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C Heu, Chan C. Kurtti, Timothy J. Nelson, Curtis M. Munderloh, Ulrike G. PLoS One Research Article Rickettsia bellii is an obligate intracellular bacterium that is one of the few rickettsiae that encode a complete set of conjugative transfer (tra) genes involved in bacterial conjugation and has been shown to exhibit pili-like structures. The reductive genomes of rickettsiae beg the question whether the tra genes are nonfunctional or functioning to enhance the genetic plasticity and biology of rickettsiae. We characterized the transcriptional dynamics of R. bellii tra genes in comparison to genes transcribed stably and above the background level to understand when and at what levels the tra genes are active or whether the tra genes are degenerative. We determined that the best reference genes, out of 10 tested, were methionyl tRNA ligase (metG) or a combination of metG and ribonucleoside diphosphate reductase 2 subunit beta (nrdF), using statistical algorithms from two different programs: Normfinder and BestKeeper. To validate the use of metG with other rickettsial genes exhibiting variable transcriptional patterns we examined its use with sca2 and rickA, genes involved in actin based motility. Both were shown to be up-regulated at different times of replication in Vero cells, showing variable and stable transcription levels of rickA and sca2, respectively. traA (Ti) was up-regulated at 72 hours post inoculation in the tick cell line ISE6, but showed no apparent changes in the monkey cell line Vero and mouse cell line L929. The transcription of tra genes was positively correlated with one another and up-regulated from 12 to 72 hours post inoculation (HPI) when compared to RBE_0422 (an inactivated transposase-derivative found within the tra cluster). Thus, the up-regulation of the tra genes indicated that the integrity and activity of each gene were intact and may facilitate the search for the optimal conditions necessary to demonstrate conjugation in rickettsiae. Public Library of Science 2015-09-09 /pmc/articles/PMC4564193/ /pubmed/26352829 http://dx.doi.org/10.1371/journal.pone.0137214 Text en © 2015 Heu 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, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Heu, Chan C.
Kurtti, Timothy J.
Nelson, Curtis M.
Munderloh, Ulrike G.
Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C
title Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C
title_full Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C
title_fullStr Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C
title_full_unstemmed Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C
title_short Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C
title_sort transcriptional analysis of the conjugal transfer genes of rickettsia bellii rml 369-c
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4564193/
https://www.ncbi.nlm.nih.gov/pubmed/26352829
http://dx.doi.org/10.1371/journal.pone.0137214
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