Abstract
Despite the large interest in the human microbiome in recent years, there are no reports of bacterial DNA methylation in the microbiome. Here metagenomic sequencing using the Pacific Biosciences platform allowed for rapid identification of bacterial GATC methylation status of a bacterial species in human stool samples. For this work, two stool samples were chosen that were dominated by a single species, Bacteroides dorei. Based on 16S rRNA analysis, this species represented over 45% of the bacteria present in these two samples. The B. dorei genome sequence from these samples was determined and the GATC methylation sites mapped. The Bacteroides dorei genome from one subject lacked any GATC methylation and lacked the DNA adenine methyltransferase genes. In contrast, B. dorei from another subject contained 20,551 methylated GATC sites. Of the 4970 open reading frames identified in the GATC methylated B. dorei genome, 3184 genes were methylated as well as 1735 GATC methylations in intergenic regions. These results suggest that DNA methylation patterns are important to consider in multi-omic analyses of microbiome samples seeking to discover the diversity of bacterial functions and may differ between disease states.
Highlights
DNA adenosine methylation (Dam methylation) of the 5 -GATC3 motif in bacterial genomes, in Escherichia coli, has been well characterized for decades (Marinus, 1987; Barras and Marinus, 1989; Marinus and Casadesus, 2009)
Of all of the DNA methylation systems known in bacteria, GATC methylation appears to have the highest impact on gene expression (Barras and Marinus, 1989)
PacBio sequencing and assembly from 105 and 439 metagenomic DNA resulted in two closed Bacteroides dorei genomes of 5,726,633 and 5,243,219 bp, respectively, The 105 and 439 genomes assembled to an average coverage across each genome of 306.72- and 249.47-fold, respectively
Summary
DNA adenosine methylation (Dam methylation) of the 5 -GATC3 motif in bacterial genomes, in Escherichia coli, has been well characterized for decades (Marinus, 1987; Barras and Marinus, 1989; Marinus and Casadesus, 2009). Of all of the DNA methylation systems known in bacteria, GATC methylation appears to have the highest impact on gene expression (Barras and Marinus, 1989). The biochemistry of the adenosine methylation reaction catalyzed by bacterial DNA adenine methyltransferase (DamMT) is well characterized (Thielking et al, 1997; Urig et al, 2002). As DamMT mutants are not lethal, the phenotype of DamMT mutants in several bacteria have shown that methylation of the GATC motif affects gene expression of many processes including chromosome replication, mismatch repair, and nucleoid structure (Løbner-Olesen et al, 2005). Significant efforts have been made in recent years to design antibiotics that inhibit DamMT (Mashhoon et al, 2004, 2006; Hobley et al, 2012; McKelvie et al , 2013)
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