Transcription termination directed by heteroduplex thr attenuator templates: Evidence that the transcript stem and loop structure is the termination signal

Abstract

We have constructed both homoduplex and heteroduplex restriction fragment templates that contained mutations in the G + C-rich region of dyad symmetry in the Escherichia coli thr operon attenuator and have studied their termination properties in an in vitro transcription system. We found that the DNA sequence of the template strand determines the transcription termination efficiency. We also constructed two double mutant heteroduplex templates to examine the role of possible complementary base pairs between the nontranscribed DNA strand and the nascent RNA in the transcription bubble. There was no indication that the potential DNA-RNA interactions in the G + C-rich region are involved in transcription termination. Taken together, our results are consistent with and extend those obtained by Ryan and Chamberlin (Ryan, T., and Chamberlin, J. (1983) J. Biol. Chem. 258, 4690-4693) for the trp operon attenuator and argue strongly against models that propose that DNA hairpin formation in the nontranscribed DNA strand or pairing of the nascent RNA with the nontranscribed DNA strand are important for transcription termination.