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Found: the elusive ANTAR transcription antiterminator.

Stewart V, van Tilbeurgh H - PLoS Genet. (2012)

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of California Davis, Davis, California, USA. vjstewart@ucdavis.edu

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Intrinsic terminators, which consist of an RNA stem-loop followed by a poly-U tract, catalyze termination by disrupting the RNA polymerase elongation complex... Both leaders encode two obvious stem-loop secondary structures: the distal intrinsic terminator (including a poly-U tract), and P1, a proximal structure essential for antitermination (Figure 1A)... Moreover, the distal stem of P2 overlaps part of the terminator proximal stem, , and thereby forms an antiterminator structure (Figure 1B)... This suggests a general model for ANTAR-mediated transcription termination, in which the dimeric ANTAR protein binds simultaneously to structures P1 and P2, stabilizing the P2 antiterminator to enable transcription readthrough. (Indeed, restriction sites introduced into the nasF operon leader destroy the P2 structure, explaining the resultant uninducible phenotype.) To test this model, Ramesh et al. first constructed a variety of site-specific alterations in the eutP leader, and confirmed the subelements important for EutV binding and antitermination: the P1 and P2 structures; their relative spacing; and the P1 and P2 loop residues A1 and G4... Presumably, each ANTAR monomer binds one of the two structures, P1 or P2... Finally, Ramesh et al. conducted bioinformatic analyses to identify ANTAR target leaders in bacterial genomes... Examples were found in a broad range of species from the Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria... Some of the leaders are adjacent to genes of unknown or uncertain function, whereas many others are adjacent to genes involved in inorganic nitrogen acquisition (ammonium, nitrate, or dinitrogen) or assimilation (glutamine synthetase and associated regulators)... The CAT domain is a homodimer of four-stranded β-sheets... Phosphorylation results in massive structural changes that bring the two CAT monomers into proper alignment to interact with a distorted minor grove in the antiterminator hairpin stem... The ANTAR domain adopts a helix-turn-helix conformation, but how it binds RNA is unknown... RNA-binding helix-turn-helix proteins include σ region 4.2, which binds the −35 region of promoter DNA as well as 6S RNA, and Ffh, which binds 4.5 S RNA in the signal recognition particle... Future progress in understanding ANTAR-mediated antitermination requires knowledge of ANTAR domain conformation and interaction with its RNA target; conformational shifts mediated through the coiled-coil; and kinetics of RNA binding in relation to transcription-driven formation of the P1, P2, and terminator structures.

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RNA-binding proteins stabilize antiterminator structures.Alternative stem-loop structures in transcribed leader regions from representative operons are shown. Terminator stem regions that participate in alternative structures are in blue, whereas antiterminator regions are in red. (A) NasR-responsive nasF operon leader in termination conformation. Dots indicate the critical residues A1 and G4 in the P1 and P2 loops. (B) Each ANTAR monomer is hypothesized to bind one of the loops, P1 and P2. Stabilizing the P2 antiterminator structure permits transcription readthrough [6] by shortening the terminator stem and separating it from the poly-U tract [2]. (C) LicT-responsive bglP operon leader in termination conformation. RAT is the ribonucleic antiterminator. (D) Each CAT (co-antiterminator) monomer binds to the antiterminator stem. Unusual base-pairing within the antiterminator stem is depicted schematically [25]. Stablilizing the antiterminator structure permits transcription readthrough [22] by shortening the terminator stem and separating it from the poly-U tract.
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pgen-1002773-g001: RNA-binding proteins stabilize antiterminator structures.Alternative stem-loop structures in transcribed leader regions from representative operons are shown. Terminator stem regions that participate in alternative structures are in blue, whereas antiterminator regions are in red. (A) NasR-responsive nasF operon leader in termination conformation. Dots indicate the critical residues A1 and G4 in the P1 and P2 loops. (B) Each ANTAR monomer is hypothesized to bind one of the loops, P1 and P2. Stabilizing the P2 antiterminator structure permits transcription readthrough [6] by shortening the terminator stem and separating it from the poly-U tract [2]. (C) LicT-responsive bglP operon leader in termination conformation. RAT is the ribonucleic antiterminator. (D) Each CAT (co-antiterminator) monomer binds to the antiterminator stem. Unusual base-pairing within the antiterminator stem is depicted schematically [25]. Stablilizing the antiterminator structure permits transcription readthrough [22] by shortening the terminator stem and separating it from the poly-U tract.

Mentions: AmiR and NasR target the transcribed leader RNA upstream of the amiE and nasF operons, respectively [12], [14]. Both leaders encode two obvious stem-loop secondary structures: the distal intrinsic terminator (including a poly-U tract) [2], and P1, a proximal structure essential for antitermination (Figure 1A). Further analysis of the nasF operon leader identified three subelements essential for NasR binding and antitermination: the P1 stem; residues A1 and G4 in the P1 loop; and part of the linker region that connects the P1 and terminator stem-loop structures [16]. However, the RNA secondary structure(s) formed during antitermination remained a mystery.


Found: the elusive ANTAR transcription antiterminator.

Stewart V, van Tilbeurgh H - PLoS Genet. (2012)

RNA-binding proteins stabilize antiterminator structures.Alternative stem-loop structures in transcribed leader regions from representative operons are shown. Terminator stem regions that participate in alternative structures are in blue, whereas antiterminator regions are in red. (A) NasR-responsive nasF operon leader in termination conformation. Dots indicate the critical residues A1 and G4 in the P1 and P2 loops. (B) Each ANTAR monomer is hypothesized to bind one of the loops, P1 and P2. Stabilizing the P2 antiterminator structure permits transcription readthrough [6] by shortening the terminator stem and separating it from the poly-U tract [2]. (C) LicT-responsive bglP operon leader in termination conformation. RAT is the ribonucleic antiterminator. (D) Each CAT (co-antiterminator) monomer binds to the antiterminator stem. Unusual base-pairing within the antiterminator stem is depicted schematically [25]. Stablilizing the antiterminator structure permits transcription readthrough [22] by shortening the terminator stem and separating it from the poly-U tract.
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pgen-1002773-g001: RNA-binding proteins stabilize antiterminator structures.Alternative stem-loop structures in transcribed leader regions from representative operons are shown. Terminator stem regions that participate in alternative structures are in blue, whereas antiterminator regions are in red. (A) NasR-responsive nasF operon leader in termination conformation. Dots indicate the critical residues A1 and G4 in the P1 and P2 loops. (B) Each ANTAR monomer is hypothesized to bind one of the loops, P1 and P2. Stabilizing the P2 antiterminator structure permits transcription readthrough [6] by shortening the terminator stem and separating it from the poly-U tract [2]. (C) LicT-responsive bglP operon leader in termination conformation. RAT is the ribonucleic antiterminator. (D) Each CAT (co-antiterminator) monomer binds to the antiterminator stem. Unusual base-pairing within the antiterminator stem is depicted schematically [25]. Stablilizing the antiterminator structure permits transcription readthrough [22] by shortening the terminator stem and separating it from the poly-U tract.
Mentions: AmiR and NasR target the transcribed leader RNA upstream of the amiE and nasF operons, respectively [12], [14]. Both leaders encode two obvious stem-loop secondary structures: the distal intrinsic terminator (including a poly-U tract) [2], and P1, a proximal structure essential for antitermination (Figure 1A). Further analysis of the nasF operon leader identified three subelements essential for NasR binding and antitermination: the P1 stem; residues A1 and G4 in the P1 loop; and part of the linker region that connects the P1 and terminator stem-loop structures [16]. However, the RNA secondary structure(s) formed during antitermination remained a mystery.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of California Davis, Davis, California, USA. vjstewart@ucdavis.edu

AUTOMATICALLY GENERATED EXCERPT
Please rate it.

Intrinsic terminators, which consist of an RNA stem-loop followed by a poly-U tract, catalyze termination by disrupting the RNA polymerase elongation complex... Both leaders encode two obvious stem-loop secondary structures: the distal intrinsic terminator (including a poly-U tract), and P1, a proximal structure essential for antitermination (Figure 1A)... Moreover, the distal stem of P2 overlaps part of the terminator proximal stem, , and thereby forms an antiterminator structure (Figure 1B)... This suggests a general model for ANTAR-mediated transcription termination, in which the dimeric ANTAR protein binds simultaneously to structures P1 and P2, stabilizing the P2 antiterminator to enable transcription readthrough. (Indeed, restriction sites introduced into the nasF operon leader destroy the P2 structure, explaining the resultant uninducible phenotype.) To test this model, Ramesh et al. first constructed a variety of site-specific alterations in the eutP leader, and confirmed the subelements important for EutV binding and antitermination: the P1 and P2 structures; their relative spacing; and the P1 and P2 loop residues A1 and G4... Presumably, each ANTAR monomer binds one of the two structures, P1 or P2... Finally, Ramesh et al. conducted bioinformatic analyses to identify ANTAR target leaders in bacterial genomes... Examples were found in a broad range of species from the Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria... Some of the leaders are adjacent to genes of unknown or uncertain function, whereas many others are adjacent to genes involved in inorganic nitrogen acquisition (ammonium, nitrate, or dinitrogen) or assimilation (glutamine synthetase and associated regulators)... The CAT domain is a homodimer of four-stranded β-sheets... Phosphorylation results in massive structural changes that bring the two CAT monomers into proper alignment to interact with a distorted minor grove in the antiterminator hairpin stem... The ANTAR domain adopts a helix-turn-helix conformation, but how it binds RNA is unknown... RNA-binding helix-turn-helix proteins include σ region 4.2, which binds the −35 region of promoter DNA as well as 6S RNA, and Ffh, which binds 4.5 S RNA in the signal recognition particle... Future progress in understanding ANTAR-mediated antitermination requires knowledge of ANTAR domain conformation and interaction with its RNA target; conformational shifts mediated through the coiled-coil; and kinetics of RNA binding in relation to transcription-driven formation of the P1, P2, and terminator structures.

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