Glucosylated 5-Hydroxymethylpyrimidines as Epigenetic DNA Bases Regulating Transcription and Restriction Cleavage
Abstrakt
5-(β-D-Glucopyranosylmmethyl)-2'-demuridine and -cytidine 5'-O-triphosphates were prepared and used for polymerase-mediated (primer extension or PCR) synthesis of DNA containing glucosylated 5-hydroxymethyluracil (5hmU) or 5-hydroxymethyluracil (5hmC). The presence of any glucosylated pyrimidines fully protected DNA from cleavage by type II restriction endonucleases.
On the other hand, while the presence of glucosylated 5hmU completely inhibited transcription by bacterial (Escherichia coli) RNA polymerase, the DNA containing the corresponding glucosylated 5hmC allowed a similar level of transcription as natural DNA. This suggests different roles of these hypermodified bases in the epigenetic regulation of transcription in bacteriophages or kinetoplastid parasites.
Consequently, enzymatic glucosylation of 5hmC-containing DNA can be used for tuning of transcription activity.