Translation initiation of IS50R read-through transcripts

Abstract

IS50R (and Tn5) normally transposes at a low frequency, partly because cells containing this insertion sequence synthesize low levels of the transposase protein. Since the 5' end of the transposase gene is located next to the outer end of IS50R (and thus close to flanking host sequences), transposition into actively transcribed genes could result in the production of read-through transcripts that would encode the transposase. We have found that these read-through transcripts are made, but are translated poorly. We isolated mutations that increase translation initiation of transposase from read-through transcripts. Most of these mutations destabilize a potential RNA secondary structure in the ribosome binding site that could form in read-through transcripts, but not in normal transcripts. In vitro RNA secondary structure analysis has confirmed the predicted RNA secondary structure and the effects of mutations. We have shown that RNA secondary structure is the major factor limiting transposase expression from read-through transcripts.