Structural requirements of the RNA precursor for the biosynthesis of the branched RNA-linked multicopy single-stranded DNA of Myxococcus xanthus

Abstract

A precursor RNA molecule (pre-msdRNA) of approximately 375 bases is considered to form a stable secondary structure which serves as a primer as well as a template to synthesize the branched RNA-linked multicopy single-stranded DNA (msDNA) of Myxococcus xanthus. When 3-base mismatches were introduced into the stem structure immediately upstream of the branched rG residue to which msDNA is linked by a 2',5'-phosphodiester linkage, the production of msDNA was almost completely blocked. However, if additional 3-base substitutions were made on the other strand to resume the complementary base pairing, msDNA production was restored, being consistent with the proposed model of msDNA synthesis. We also found that the branched rG residue of pre-msdRNA could not be replaced with either rC or rA, while the 5' end (dC) of msDNA which is linked to the branched rG could be substituted with a dG residue. Together with several other mutations, the structural requirements of pre-msdRNA are discussed with respect to the mechanism of msDNA biosynthesis.