TRANSCRIPTION-FREQUENCY-DEPENDENT MODULATION OF AN ATTENUATOR IN A RIBOSOMAL-PROTEIN RNA-POLYMERASE OPERON REQUIRES AN UPSTREAM SITE

Although the attenuator located between the ribosomal protein and RNA polymerase gene domains of the Escherichia coli rplKAJLrpoBC operon ha s a maximum termination efficiency of 80%, the level of termination is diminished with decreasing transcription frequency. In this report, t he use of transcriptional fusions to further investigate the mechanism of transcription-frequency-dependent regulation is described. The ter mination efficiency of two other weak terminators was assayed over a w ide range of transcription frequencies programmed by different strengt h promoters. The results indicated that a decrease in termination effi ciency with decreasing transcription frequency is not an inherent prop erty of weak terminators. Deletion of the 165 bp segment located 439-2 74 bp upstream of the attenuator abrogated the difference in terminati on efficiency normally seen between high and low levels of transcripti on. This suggests that a cis-acting site located in this upstream regi on is necessary for transcription-frequency-dependent modulation of th e attenuator's function. However, this site apparently works only in c ombination with the attenuator, since it did not cause transcription-f requency-dependent modulation when placed upstream of two other weak t erminators. Analysis of the readthrough frequencies of single or tande m copies of the attenuator indicated that the transcription complexes which pass through the attenuator have not been converted to terminati on-resistant complexes in a manner analogous to the N-mediated antiter mination system of lambda. Finally, an examination of termination effi ciency in three nusA mutants suggested that although NusA increases re adthrough at the attenuator it is not directly involved in transcripti on-frequency-dependent modulation.