Nonsense polarity, RNA processing and decay in phage f1.

Nonsense polarity in most cases depends on activation of cryptic transcription terminators. We found that the strong polar effect observed in the nonsense polar mutant R4 of phage f1, mapping in the 5’ proximal region of gene III, instead depends on enhanced instability of mutant mRNAs, whose pattern can be restored by reduction of RNase E activity. rne -(ts) E. coli strains allowed to explore the mechanisms underlying f1 mRNA processing and degradation. The major gene III species, a 1.8 Kb long molecule, appeared to be a secondary transcript, whose decay is modulated by a REP, located at its 3' end. The RNA pool of a mutagenized phage unable to form that structure, lacks completely that transcript. Infection of RNaseE-(ts) cells resulted in the accumulation of very large transcripts (3.9-6.4 Kb) and the reduction of the level of the 1.8 Kb transcript. The pool of gene III messages found at non permissive temperature includes transcripts composed uniquely by coding sequences (present also in RNase E+ cells) and mRNAs containing, in their internal portion, the untranslated sequences of the IG region. Thus it appears that in wild type cells these sequences are transcribed and promptly cut by RNase E. Cloning of the IG sequences together with last nt of gene IV into a plasmid, maintaining the natural translation frame, confirmed the presence of RNase E cutting sites into the IG, whose translation in suppressor cells makes them resistant to this ribonuclease. Thus ribosome activity appears to challenge that of the RNase E in mRNA processing.