Rho-dependent transcription termination

Richardson, John P.

doi:10.1016/0167-4781(90)90048-7

Cited by 51 publications

(44 citation statements)

References 53 publications

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“…ATP hydrolysis is required for termination by vaccinia VTF (17,18), E. coli rho (11)(12)(13), and RNA polymerase III termina- …”

Section: Discussionmentioning

confidence: 99%

“…Instead, VTF is required to recognize a special sequence in the nascent RNA and transduce the signal to an activity named factor X, which catalyzed the ATP hydrolysis required for transcript release (10). Rho-dependent transcription termination in Escherichia coli requires ATP hydrolysis catalyzed by rho, an RNA-dependent ATPase (11)(12)(13). Rho binds to RNA and translocates along nascent transcripts to track elongating complexes and catalyzes transcript release by hydrolyzing ATP.…”

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confidence: 99%

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Drosophila Factor 2, an RNA Polymerase II Transcript Release Factor, Has DNA-dependent ATPase Activity

Zhi

Price

1997

Journal of Biological Chemistry

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Factor 2 demonstrated a strong DNA-dependent ATPase activity that correlated with its transcript release activity. At 20 g/ml DNA, the ATPase activity of factor 2 had an apparent K m(ATP) of 28 M and an estimated K cat of 140 min ؊1 . Factor 2 caused the release of nascent transcripts associated with elongation complexes generated by RNA polymerase II on a dC-tailed template. Therefore, no other protein cofactors are required for the transcript release activity of factor 2. Using the dC-tailed template assay, it was found that renaturation of the template was required for factor 2 function.Control of transcription elongation has been recognized as an important mechanism regulating eucaryotic gene expression (1-3). The strategy to achieve the elongation control is to first apply a negative elongation potential to block and ultimately terminate RNA polymerase II transcription. The premature termination is then regulated by positive factors which enable RNA polymerase II to read through the block to transcription elongation. Based on our in vitro studies of Drosophila RNA polymerase II transcription, we proposed a model for the control of elongation by RNA polymerase II that incorporated the function of both negative and positive factors (4,5). According to the model, all RNA polymerase II molecules that initiate from a promoter are destined to produce only short transcripts because of the action of negative transcription elongation factor (N-TEF).1 Escape from this abortive elongation into productive elongation requires the action of positive transcription elongation factor (P-TEF). P-TEFb, one of the components of P-TEF, has been purified and was found to phosphorylate the C-terminal domain of the largest subunit of RNA polymerase II (6, 7). Factor 2, the first identified component of N-TEF, has been purified to apparent homogeneity as a monomer with a molecular mass of 154 kDa (8). Factor 2 associates with early elongation complexes and causes premature termination by releasing RNA polymerase II transcripts generated from early elongation complexes in an ATP-dependent manner (8).ATP has been found to be required for factor-dependent termination in several transcription systems. Autoantigen La, an RNA polymerase III transcription termination factor, hydrolyzes ATP to provide energy for the termination reaction (9). The ATPase activity of La was dependent on RNA binding and was only activated by double-stranded RNA or DNA-RNA hybrid (9). In vaccinia virus, transcription termination mediated by virus-encoded termination factor (VTF) also requires ATP hydrolysis, though VTF itself is not directly responsible for the ATP hydrolysis. Instead, VTF is required to recognize a special sequence in the nascent RNA and transduce the signal to an activity named factor X, which catalyzed the ATP hydrolysis required for transcript release (10). Rho-dependent transcription termination in Escherichia coli requires ATP hydrolysis catalyzed by rho, an RNA-dependent ATPase (11-13). Rho binds to RNA and translocates along nascent tra...

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“…ATP hydrolysis is required for termination by vaccinia VTF (17,18), E. coli rho (11)(12)(13), and RNA polymerase III termina- …”

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Drosophila Factor 2, an RNA Polymerase II Transcript Release Factor, Has DNA-dependent ATPase Activity

Zhi

Price

1997

Journal of Biological Chemistry

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“…In strain CU1675, the deletion did not extend that far but did apparently extend far enough to have strongly reduced the polar effect. Recall that rhodependent termination is believed to be dependent on a region of unstructured as well as untranslated message upstream of the termination site (44).…”

Section: Discussionmentioning

confidence: 99%

“…It has been repeatedly observed in all experiments of this kind that the derepression obtained with ,-galactosidase when leucine is litniting is less than that seen (4,57). The endpoint of the other deletion, AilvG2216, is about 130 bp upstream of the first of the three rho-dependent termination sites, a distance that may interfere with the rho-dependent termination (44). It is of interest, nevertheless, that the expression in the construct with AilvG2216 is only about one-half that in the construct with the three rho-dependent terminators deleted.…”

Section: Methodsmentioning

confidence: 99%

Specificity of attenuation control in the ilvGMEDA operon of Escherichia coli K-12

1991

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Three different approaches were used to examine the regulatory effects of the amino acids specified by the peptide-coding region of the leader transcript of the ilvGMEDA operon of Escherichia coli K-12. Gene expression was examined in strains carrying an ilvGMED'-lac operon fusion. In one approach, auxotrophic derivatives were starved of single amino acids for brief periods, and the burst of j-galactosidase synthesis upon adding the missing amino acid was determined. Auxotrophic derivatives were also grown for brief periods with a limited supply of one amino acid (derepression experiments). Finally, prototrophic strains were grown in minimal medium supplemented with single and multiple supplements of the chosen amino acids. Although codons for arginine, serine, and proline are interspersed among the codons for the three branched-chain (regulatory) amino acids, they appeared to have no effect when added in excess to prototrophs or when supplied in restricted amounts to auxotrophs. Deletions removing the terminator stem from the leader removed all ilvspecific control, indicating that the attenuation mechanism is the sole mechanism for ilv-specific control.The specific control of expression of the ilvGMEDA operon in the enteric bacteria is characterized by repression of the operon when the three branched-chain amino acids are in ample supply but derepression when the supply of any one of the three is limited (14,30). This ilv-specific regulation appears to be exclusively a consequence of an attenuation mechanism ( Fig. 1) (4,32,38). In principle, the attenuation mechanism controlling the ilv operon appears to be quite similar to that found to control operons involved in tryptophan (35), leucine (19), phenylalanine (61), histidine (3), and threonine (17) biosynthesis: the leader region specifies a short peptide containing a disproportionate frequency of the amino acids that are found to regulate the operon. Thus, the rate at which the leader can be translated serves to sense whether the regulatory amino acid is in short or ample supply.As emphasized by Landick and Yanofsky (31), an important feature of the attenuation mechanism is the pausing of RNA polymerase at a certain site in the leader region. The pause allows time for the ribosome to initiate translation, and as translation proceeds, the base pairing in the protector (1:2 stem, Fig. 2) is progressively disrupted. Should ribosome movement be retarded, presumably by stalling at a regulatory codon, long enough for the bases in the upstream arm of the terminator (bases 157 to 164) to emerge on the nascent transcript, these bases can pair with those in the downstream arm of the protector (bases 98 to 105) in the antiterminator (2:3 stem, Fig. 3). It is assumed that, in the absence of ribosome stalling (repressing conditions), translation proceeds rapidly enough that the bases in the downstream arm of the protector will also be covered by the ribosome or otherwise be sterically prevented from antiterminator formation and the terminator will be formed instead. Altho...

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“…The Escherichia coli protein rho is required for correct transcript termination at numerous specific (rho-dependent) termination sites on the DNA template (for reviews see PIatt & Bear, 1983;von Hippel et al, 1984; Platt, 1986; Richardson, 1990). At other specific (intrinsic or factor-independent) sites, RNA polymerase can terminate transcription without the participation of protein cofactors.…”

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confidence: 99%

Functional interactions of ligand cofactors with Escherichia coli transcription termination factor rho. II. Binding of RNA

1992

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The rho protein of Escherichia coli interacts with the nascent RNA transcript while RNA polymerase is paused at specific rho-dependent termination sites on the DNA template, and (in a series of steps that are still largely undefined) brings about transcript termination at these sites. In this paper we characterize the interactions of rho with RNA and relate these interactions to the quaternary structure of the functional form of rho. We use CD spectroscopy and analytical ultracentrifugation to determine the binding interactions of rho with RNA ligands of defined length ([rC], where n 2 6). Rho binds to long RNA chains as a hexamer characterized by D3 symmetry. Each hexamer binds -70 residues of RNA. We show by ultracentrifugation and dynamic laser light scattering that, in the presence of RNA ligands less than 22 nucleotide residues in length, rho changes its quaternary structure and becomes a homogeneous dodecamer. The dodecamer contains six strong binding sites for short RNA ligands: i.e., one site for every two rho protomers. The measured association constant of these short RNAs to rho increases with increasing (rC), length, up to n = 9, suggesting that the binding site of each rho protomer interacts with 9 RNA nucleotide residues. Oligo(rC) ligands bound to the strong RNA binding sites on the rho dodecamer do not significantly stimulate the RNA-dependent ATPase activity of rho. Based on these features of the rho-RNA interaction and other experimental data we propose a molecular model of the interaction of rho with its cofactors.

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Rho-dependent transcription termination

Cited by 51 publications

References 53 publications

Drosophila Factor 2, an RNA Polymerase II Transcript Release Factor, Has DNA-dependent ATPase Activity

Drosophila Factor 2, an RNA Polymerase II Transcript Release Factor, Has DNA-dependent ATPase Activity

Specificity of attenuation control in the ilvGMEDA operon of Escherichia coli K-12

Functional interactions of ligand cofactors with Escherichia coli transcription termination factor rho. II. Binding of RNA

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