Bacterial transcription terminators: the RNA 3'-end chronicles

Abstract

The process of transcription termination is essential to proper expression of bacterial genes and, in many cases, to the regulation of bacterial gene expression. Two types of bacterial transcriptional terminators are known to control gene expression. Intrinsic terminators dissociate transcription complexes without the assistance of auxiliary factors. Rho-dependent terminators are sites of dissociation mediated by an RNA helicase called Rho. Despite decades of study, the molecular mechanisms of both intrinsic and Rho-dependent termination remain uncertain in key details. Most knowledge is based on the study of a small number of model terminators. The extent of sequence diversity among functional terminators and the extent of mechanistic variation as a function of sequence diversity are largely unknown. In this review, we consider the current state of knowledge about bacterial termination mechanisms and the relationship between terminator sequence and steps in the termination mechanism.

Fig. 1

Structure of the EC. (a) Model of EC based on a Thermus thermophilus EC crystal structure (Protein Data Bank ID 2o5i). (b) Cutaway view of EC model showing locations of open and closed conformations of the TL (Protein Data Bank IDs 1iw7 and 2o5j, respectively)^, and the location of the lid separating the RNA exit and main channels. (c) Close-up view of the RNA exit channel with the flap tip removed and the nucleotides numbered for an EC in the pretranslocated register.

Fig. 2

Mechanisms of intrinsic termination. The major intermediates in the intrinsic termination pathway are depicted in schematic form. Three alternative routes to EC disruption by hairpin completion are depicted. The version of hairpin invasion depicted corresponds to the specific conformational change model of Epshtein et al. The changes in TL conformation in different intermediates are emphasized by color changes but remain speculative.

Fig. 3

Sequence features of intrinsic terminators. Canonical RNA sequences (red) are depicted paired to a DNA scaffold with paired and unpaired nucleotides depicted as filled circles and lines. The sequence of λt_R2 is used to depict canonical features in the RNA, except the hairpin loop, which is shown as filled circles and lines. (a) Configuration at the pause step, with portions of the DNA scaffold omitted. The extent of conservation in the RNA 3′ stem and in the U-tract is shown above the DNA as information content using WebLogo (www.weblogo.berkeley.edu).^, The %AT for downstream DNA of 50 near-perfect U-tract terminators (blue) and 50 imperfect U-tract terminators (red) selected as described in the text is shown. The variations in %AT at positions 11–13 and 19–21 (numbered relative to the U-tract) are significantly different from a randomized sequence (p≤0.05; t-test). (b) RNA/DNA configuration after hairpin nucleation. (c) RNA/DNA configuration during EC disruption, with canonical terminator features labeled.

Fig. 4

Steps prior to Rho termination. The blue panel (left) depicts a model in which Rho binds only to RNA, whereas the green panel (right) depicts an alternate model in which Rho binds directly to RNAP. rut RNA may remain bound to the Rho-NTD during translocation (tethered tracking model), forming a loop between the primary and secondary RNA-binding sites (shown as partially transparent RNA), or rut RNA may be released during translocation (simple translocation model, shown as opaque RNA). Both RNA extraction and conformational change models are possible in either pathway, as depicted in the darker panels (bottom).

Fig. 5

Crystal structures of Rho. (a) The open form of Rho (1pvo) bound to AMPPNP (phosphoaminophosphonic acid-adenylate ester, an ATP analogue) and RNA. (b) The closed asymmetric form of Rho (3ice) bound to ADP-BeF (adenosine-5′-diphosphate, an ATP analogue; BeF, a phosphate mimic) and RNA.

Fig. 6

Two-checkpoint mechanism to suppress Rho termination in protein-coding genes (polarity). Competition can occur for recruitment of rut RNA sequestered in the ribosome and for the NusG-CTD, which binds ribosomal protein S10 or Rho. Because NusG affects Rho dissociation from ECs but not recruitment, the two mechanisms may operate as sequential checks to determine whether an mRNA can be translated.