RNA sequence and secondary structure requirements for rho-dependent transcription termination

The interaction of E. coli termination factor rho with the nascent RNA transcript appears to be a central feature of the rho-dependent transcription termination process. Based on in vitro studies of the rho-dependent termination of the transcript initiated at the PR promoter of bacteriophage lambda, and on earlier studies, Morgan, Bear and von Hippel (J. Biol. Chem. 258, 9565-9574, 1983) proposed a model defining the features of a potential binding site for rho protein on transcripts subject to rho-dependent termination. This model suggested that an effective rho binding site on a nascent RNA transcript should be: (i) greater than 70-80 nucleotide residues in length; (ii) essentially unencumbered with stable secondary structure; (iii) relatively sequence non-specific; and (iv) located within a few hundred nucleotide residues upstream of the potential rho-dependent terminus. In this paper we examine the sequences and secondary structures of several transcripts that exhibit rho-dependent termination to test this hypothesis further. Unstructured regions of approximately the expected size and location were found on all the transcripts examined. Though several short specific sequence elements were found to occur in a very similar arrangement on the lambda PR- and lambda PL-initiated transcripts of lambda phage, no such elements of sequence regularity were found on any of the other rho-dependent transcripts. The results of the sequence comparisons reported here strongly support the generality of the "unstructured binding site" hypothesis for rho-dependent termination.