Inwon Park

Mutational Analysis of RNA Structures and Sequences Postulated to Affect 3′ Processing of M1 RNA, the RNA Component of Escherichia coli RNase P

When the rnpB gene encoding M1 RNA, the RNA component of Escherichia coli RNase P, is transcribed, the primary M1 RNA transcript (pM1 RNA) is produced and subsequently processed at the 3′ end to generate the mature M1 RNA. To study features of pM1 RNA thought to be involved in RNA processing, systematic mutations were introduced in sequence elements and secondary structures surrounding the processing site using p23 RNA, a truncated pM1 RNA transcribed from the internally deleted rnpB gene, as a model substrate and the processing of its mutant derivatives was analyzed in vivo and in vitro. Neither the alteration of two bases forming the processing site nor the disruption of secondary structures surrounding the site significantly affected the processing efficiency although the secondary structures were required for maintaining RNA stability. In contrast, mutations at the rne-dependent site, GAUUU, immediately 3′ to the processing site inhibited the processing and the extent of the inhibition varied with the altered sequences. Furthermore, the processing of the mutants of the rne-dependent site as well as wild-type p23 RNA was inhibited in an E. coli rnets strain at the nonpermissive temperature.

Identification of a structural motif of 23S rRNA interacting with 5S rRNA.

To identify RNA motifs interacting with 5S rRNA, a systematic evolution of ligands by exponential enrichment experiment was applied. Some of the resulting RNA aptamers contained a consensus sequence similar to the sequence in the loop region of helix 89 of 23S rRNA. We show that the synthetic helix 89 RNA motif indeed interacted with 5S rRNA and that the region around loop B of 5S rRNA was involved in this interaction. These results suggest the presence of a novel RNA–RNA interaction between 23S rRNA and 5S rRNA which may play an important role in the ribosome function.