A potential stem-loop structure and the sequence CAAUCAA in the transcript are insufficient to signal ϱ-dependent transcription termination at λtR1

It has been suggested that a sequence in the RNA transcript that can form a stem and loop structure, followed by the sequence CAAUCAA, is the signal for rho-dependent transcription termination. We tested this hypothesis by synthesizing a DNA duplex whose sequence corresponds to a region of the lambda tR1 terminator that contains these structural features. We cloned this synthetic DNA fragment under the control of the lacUV5 promoter, and showed that it does not cause rho-dependent termination in vitro. RNA polymerase pauses during in vitro transcription across the synthetic sequence, although less efficiently than at the corresponding sequence on the lambda template. No rho-mediated termination was detected even under conditions that prolonged transcriptional pausing at the synthetic site, indicating that the synthetic sequence is defective as a transcript release site. We suggest that unlike rho-independent terminators, rho-dependent terminators require sequences in addition to those immediately before the sites of termination.