FUNCTION OF RNA SECONDARY STRUCTURES IN TRANSCRIPTIONAL ATTENUATION OF THE BACILLUS-SUBTILIS PYR OPERON

The Bacillus subtilis pyr operon is regulated by exogenous pyrimidines by a transcriptional attenuation mechanism. Transcription in vitro fr om pyr DNA templates specifying attenuation regions yielded terminated and read-through transcripts of the expected lengths. Addition of the PyrR regulatory protein plus UMP led to greatly increased termination . Synthetic antisense deoxyoligonucleotides were used to probe possibl e secondary structures in the pyr mRNA that were proposed to play role s in controlling attenuation. Oligonucleotides predicted to disrupt te rminator structures suppressed termination, whereas oligonucleotides p redicted to disrupt the stem of antiterminator stem-loops strongly pro moted termination at the usual termination site. Oligonucleotides that disrupt a previously unrecognized stem-loop structure, called the ant i-antiterminator, the formation of which interferes with formation of the downstream antiterminator, suppressed termination. We propose that transcriptional attenuation of the pyr operon is governed by switchin g between alternative antiterminator versus anti-antiterminator plus t erminator structures, and that PyrR acts by UMP dependent binding to a nd stabilization of the anti-antiterminator.