Genomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase

ABSTRACT DNA methylation in prokaryotes is widespread. The most common modification of the genome is the methylation of adenine at the N-6 position. In Escherichia coli K-12 and many gammaproteobacteria, this modification is catalyzed by DNA adenine methyltransferase (Dam) at the GATC consensus sequ...

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Autores principales:	Lacey L. Westphal, Peter Sauvey, Matthew M. Champion, Ian M. Ehrenreich, Steven E. Finkel
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2016
Materias:	Dam methyltransferase long-term stationary phase SMRT epigenetics long-term survival methylation Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/db0632fe30b545d98833ffc1ea0fbb54
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spelling	oai:doaj.org-article:db0632fe30b545d98833ffc1ea0fbb542021-12-02T19:45:29ZGenomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase10.1128/mSystems.00130-162379-5077https://doaj.org/article/db0632fe30b545d98833ffc1ea0fbb542016-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSystems.00130-16https://doaj.org/toc/2379-5077ABSTRACT DNA methylation in prokaryotes is widespread. The most common modification of the genome is the methylation of adenine at the N-6 position. In Escherichia coli K-12 and many gammaproteobacteria, this modification is catalyzed by DNA adenine methyltransferase (Dam) at the GATC consensus sequence and is known to modulate cellular processes including transcriptional regulation of gene expression, initiation of chromosomal replication, and DNA mismatch repair. While studies thus far have focused on the motifs associated with methylated adenine (meA), the frequency of meA across the genome, and temporal dynamics during early periods of incubation, here we conduct the first study on the temporal dynamics of adenine methylation in E. coli by Dam throughout all five phases of the bacterial life cycle in the laboratory. Using single-molecule real-time sequencing, we show that virtually all GATC sites are significantly methylated over time; nearly complete methylation of the chromosome was confirmed by mass spectroscopy analysis. However, we also detect 66 sites whose methylation patterns change significantly over time within a population, including three sites associated with sialic acid transport and catabolism, suggesting a potential role for Dam regulation of these genes; differential expression of this subset of genes was confirmed by quantitative real-time PCR. Further, we show significant growth defects of the dam mutant during long-term stationary phase (LTSP). Together these data suggest that the cell places a high premium on fully methylating the chromosome and that alterations in methylation patterns may have significant impact on patterns of transcription, maintenance of genetic fidelity, and cell survival. IMPORTANCE While it has been shown that methylation remains relatively constant into early stationary phase of E. coli, this study goes further through death phase and long-term stationary phase, a unique time in the bacterial life cycle due to nutrient limitation and strong selection for mutants with increased fitness. The absence of methylation at GATC sites can influence the mutation frequency within a population due to aberrant mismatch repair. Therefore, it is important to investigate the methylation status of GATC sites in an environment where cells may not prioritize methylation of the chromosome. This study demonstrates that chromosome methylation remains a priority even under conditions of nutrient limitation, indicating that continuous methylation at GATC sites could be under positive selection.Lacey L. WestphalPeter SauveyMatthew M. ChampionIan M. EhrenreichSteven E. FinkelAmerican Society for MicrobiologyarticleDam methyltransferaselong-term stationary phaseSMRTepigeneticslong-term survivalmethylationMicrobiologyQR1-502ENmSystems, Vol 1, Iss 6 (2016)
institution	DOAJ
collection	DOAJ
language	EN
topic	Dam methyltransferase long-term stationary phase SMRT epigenetics long-term survival methylation Microbiology QR1-502
spellingShingle	Dam methyltransferase long-term stationary phase SMRT epigenetics long-term survival methylation Microbiology QR1-502 Lacey L. Westphal Peter Sauvey Matthew M. Champion Ian M. Ehrenreich Steven E. Finkel Genomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase
description	ABSTRACT DNA methylation in prokaryotes is widespread. The most common modification of the genome is the methylation of adenine at the N-6 position. In Escherichia coli K-12 and many gammaproteobacteria, this modification is catalyzed by DNA adenine methyltransferase (Dam) at the GATC consensus sequence and is known to modulate cellular processes including transcriptional regulation of gene expression, initiation of chromosomal replication, and DNA mismatch repair. While studies thus far have focused on the motifs associated with methylated adenine (meA), the frequency of meA across the genome, and temporal dynamics during early periods of incubation, here we conduct the first study on the temporal dynamics of adenine methylation in E. coli by Dam throughout all five phases of the bacterial life cycle in the laboratory. Using single-molecule real-time sequencing, we show that virtually all GATC sites are significantly methylated over time; nearly complete methylation of the chromosome was confirmed by mass spectroscopy analysis. However, we also detect 66 sites whose methylation patterns change significantly over time within a population, including three sites associated with sialic acid transport and catabolism, suggesting a potential role for Dam regulation of these genes; differential expression of this subset of genes was confirmed by quantitative real-time PCR. Further, we show significant growth defects of the dam mutant during long-term stationary phase (LTSP). Together these data suggest that the cell places a high premium on fully methylating the chromosome and that alterations in methylation patterns may have significant impact on patterns of transcription, maintenance of genetic fidelity, and cell survival. IMPORTANCE While it has been shown that methylation remains relatively constant into early stationary phase of E. coli, this study goes further through death phase and long-term stationary phase, a unique time in the bacterial life cycle due to nutrient limitation and strong selection for mutants with increased fitness. The absence of methylation at GATC sites can influence the mutation frequency within a population due to aberrant mismatch repair. Therefore, it is important to investigate the methylation status of GATC sites in an environment where cells may not prioritize methylation of the chromosome. This study demonstrates that chromosome methylation remains a priority even under conditions of nutrient limitation, indicating that continuous methylation at GATC sites could be under positive selection.
format	article
author	Lacey L. Westphal Peter Sauvey Matthew M. Champion Ian M. Ehrenreich Steven E. Finkel
author_facet	Lacey L. Westphal Peter Sauvey Matthew M. Champion Ian M. Ehrenreich Steven E. Finkel
author_sort	Lacey L. Westphal
title	Genomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase
title_short	Genomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase
title_full	Genomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase
title_fullStr	Genomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase
title_full_unstemmed	Genomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase
title_sort	genomewide dam methylation in <named-content content-type="genus-species">escherichia coli</named-content> during long-term stationary phase
publisher	American Society for Microbiology
publishDate	2016
url	https://doaj.org/article/db0632fe30b545d98833ffc1ea0fbb54
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Genomewide Dam Methylation in <named-content content-type="genus-species">Escherichia coli</named-content> during Long-Term Stationary Phase

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