Structural Organization of Transcription Termination Factor Rho

Abstract

Escherichia coli has two known modes for termination of RNA transcription (1-4). One is intrinsic to the function of RNA polymerase, which can spontaneously terminate transcription in response to certain, limited sequences. The other mode is dependent upon the action of an essential protein factor called Rho and occurs at sequences that are specific for its function but that are less constrained than the sequences for intrinsic termination.Rho protein functions as a hexamer of a single polypeptide chain with 419 residues, which is the product of the rho gene (5). It is an RNA-binding protein with the capacity to hydrolyze ATP and other nucleoside triphosphates. Rho acts to cause termination by first binding to a site on the nascent transcript and by subsequently using its ATP hydrolysis activity as a source of energy to mediate dissociation of the transcript from RNA polymerase and the DNA template (6). In the cell, the ability of Rho to act at several terminators is dependent upon the presence of an essential 21-kDa protein called NusG (7) that binds both RNA polymerase and Rho itself (8). In vitro the dependence on NusG became apparent only at proximal terminators (at sites Ͻ300 base pairs from the promoter) and under conditions when the RNA molecules are being elongated at the in vivo rate of ϳ40 -50 nucleotides/s (9). The requirement for NusG when RNA chain growth is fast suggests that the NusG is acting to overcome a kinetic limitation of Rho to act alone, perhaps through mediating earlier access to the nascent RNA by the formation of a complex of Rho with RNA polymerase (10).The mechanism of how Rho acts to dissociate the transcription complex is unknown. One important approach to elucidating how interactions with RNA mediate termination of transcription is to determine the structure of the protein. Until a good crystal structure becomes available the properties of its structure will have to be inferred from other, less direct methods, such as biochemical characterization of the protein, phylogenetic comparative analyses, and the functional properties of mutants with known amino acid changes. This review summarizes our current understanding of the structure and function of transcription termination factor Rho based on these indirect approaches.

Nucleotide Binding Induces Conformational Changes in Escherichia coli Transcription Termination Factor Rho

“…7). It is known that the RNA wraps around and binds with a high affinity to the primary RNA-binding sites in the N-terminal domain of Rho (3,(27)(28)(29). Our results indicate that RNA binding to the primary sites causes a conformational change in the quaternary structure of Rho that exposes the nucleotide-binding sites at the subunit interface and renders them more accessible to the incoming and outgoing nucleotides (Fig.…”

“…The Escherichia coli transcription termination factor Rho is a hexameric protein that has the ability to unwind RNA/DNA heteroduplexes (1)(2)(3)(4)(5). Although the exact mechanism by which Rho causes transcription termination is not known, Rho protein binds nascent mRNA at specific loading sites, and it is believed that Rho translocates along RNA until it reaches the transcription complex, where it disrupts the transcription ternary complex (3,6).…”

“…Although the exact mechanism by which Rho causes transcription termination is not known, Rho protein binds nascent mRNA at specific loading sites, and it is believed that Rho translocates along RNA until it reaches the transcription complex, where it disrupts the transcription ternary complex (3,6). Translocation along nucleic acid is, therefore, a basic activity of Rho similar to helicases (7,8), and understanding it requires the knowledge of how nucleotide binding and hydrolysis events at the multiple nucleotide-binding sites on the Rho hexamer are coordinated to the mechanics of protein translocation.…”

The Transcription Elongation Factor CA150 Interacts with RNA Polymerase II and the Pre-mRNA Splicing Factor SF1

“…The transcription termination factor Rho interacts with the NusG protein to mediate termination. Rho also has an RNA binding domain that recognizes the nascent transcript, while NusG binds directly to the polymerase (71). In addition, the phage lambda N protein, an antitermination factor, affects elongation through a similar pathway.…”

Preparation and Characterization of Bacterial Protein Complexes for Structural Analysis

“…These mechanisms, commonly referred to as Rhoindependent and Rho-dependent termination, are essential for the regulation of bacterial gene expression (Richardson and Greenblatt, 1996). Rho-dependent termination requires the presence of a hexameric helicase, Rho (Brown et al, 1981), an essential transcription factor that binds nucleic acids at specific termination sites (rut), and translocates along the RNA until it reaches the transcription complex (Geiselmann et al, 1993;Platt, 1994;Richardson, 1996). One of the Rho-specific inhibitors of transcription is the product of the yaeO gene, which reduces termination in the Rho-dependent bacteriophage terminator tL1, and upstream the autogenously regulated gene rho (Pichoff et al, 1998).…”