Richard Roberts

Bacterial Methylomes


Abstract

We have become accustomed to describing genomes as strings of A’s, C’s, G’s and T’s. This is how DNA sequencing results are reported and then stored in the sequence databases. But we know that almost universally DNA carries modifications, such as methylation, which play a key role in many biological events. In humans, m5CpG and its oxidized product hm5C are important epigenetic elements that can affect embryonic development and differentiation. Prokaryotic genomes are also methylated, but with just a few exceptions – such as protection for restriction enzymes, marking parental strand during mismatch repair and involvement in cell cycle control – rather little is known of their function. A major reason for this is that the complete analysis of methylation patterns in bacterial genomes has been extremely difficult if not impossible. Bisulfite sequencing can be used, with difficulty, to probe m5C modification, but m4C and m6A modifications, which are promiscuous in bacterial genomes, have proved almost impossible to investigate.

Within the last two years, however, a new technology, SMRT™ sequencing, has been developed by Pacific Biosciences that permits both the DNA sequence and its methylation pattern to be determined simultaneously. This has provided a breakthrough in analyzing bacterial methylation by enabling DNA methyltransferase (MTase) recognition specificities to be determined in an almost trivial fashion. It provides a unique insight into prokaryotic biology and reveals extremely interesting patterns of methylation that await biological function determination. Whole genomes can be scanned in a single experiment with ease. Many exciting new findings are emerging and complete methylomes for more than 500 bacterial and archaeal genomes are now known. Some MTases are part of restriction-modification systems, which protect bacteria from phage infections and prevent foreign DNA from entering the cells. For Type I and Type III restriction systems this also means that the restriction enzyme recognition sequences will also become known because the specificity is determined by the methylase in the case of the Type III enzymes and by a common specificity subunit in the case of the Type I enzymes. This analysis used to take months for one enzyme and was rarely undertaken. Some new types of MTases have been found and because many MTases are not associated with restriction systems it is probable that new functions await discovery. A door has been cracked open that promises a new dimension in the study of genomes.

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2. Murray, I.A., Clark, T.A., Morgan, R.D., Boitano, M., Anton, B.P., Luong, K., Fomenkov, A., Turner, S.W., Korlach, J., Roberts, R.J. Nucleic Acids Res. 40: 11450-11462 (2012). The methylomes of six bacteria.


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