Unknown

Dataset Information

0

HK022 Nun Requires Arginine-Rich Motif Residues Distinct from ? N.


ABSTRACT: Bacteriophage ? N protein binds boxB RNA hairpins in the nut (N utilization) sites of immediate early ? transcripts and interacts with host factors to suppress transcriptional termination at downstream terminators. In opposition to ? N, the Nun protein of HK022 binds the boxBs of coinfecting ? transcripts, interacts with a similar or identical set of host factors, and terminates transcription to suppress ? replication. Comparison of N-boxB and Nun-boxB nuclear magnetic resonance (NMR) structural models suggests similar interactions, though limited mutagenesis of Nun is available. Here, libraries of Nun's arginine-rich motif (ARM) were screened for the ability to exclude ? coinfection, and mutants were assayed for Nun termination with a boxB plasmid reporter system. Several Nun ARM residues appear to be immutable: Asp26, Arg28, Arg29, Arg32, Trp33, and Arg36. Asp26 and Trp33 appear to be unable to contact boxB and are not found at equivalent positions in ? N ARM. To understand if the requirement of Asp26, Trp33, and Arg36 indicated differences between HK022 Nun termination and ? N antitermination complexes, the same Nun libraries were fused to the activation domain of ? N and screened for clones able to complement N-deficient ?. Mutants were assayed for N antitermination. Surprisingly, Asp26 and Trp33 were still essential when Nun ARM was fused to N. Docking suggests that Nun ARM contacts a hydrophobic surface of the NusG carboxy-terminal domain containing residues necessary for Nun function. These findings indicate that Nun ARM relies on distinct contacts in its ternary complex and illustrate how protein-RNA recognition can evolve new regulatory functions.? N protein interacts with host factors to allow ? nut-containing transcripts to elongate past termination signals. A competing bacteriophage, HK022, expresses Nun protein, which causes termination of ? nut transcripts. ? N and HK022 Nun use similar arginine-rich motifs (ARMs) to bind the same boxB RNAs in nut transcripts. Screening libraries of Nun ARM mutants, both in HK022 Nun and in a ? N fusion, revealed amino acids essential to Nun that could bind one or more host factors. Docking suggests that NusG, which is present in both Nun termination and N antitermination, is a plausible partner. These findings could help understand how transcription elongation is regulated and illustrate how subtle differences allow ARMs to evolve new regulatory functions.

SUBMITTER: Tawk CS 

PROVIDER: S-EPMC4621093 | BioStudies | 2015-01-01

REPOSITORIES: biostudies

Similar Datasets

2008-01-01 | S-EPMC2597491 | BioStudies
2008-01-01 | S-EPMC2583623 | BioStudies
2009-01-01 | S-EPMC2763281 | BioStudies
2014-01-01 | S-EPMC4060646 | BioStudies
2008-01-01 | S-EPMC2446750 | BioStudies
2005-01-01 | S-EPMC1276712 | BioStudies
2008-01-01 | S-EPMC2677985 | BioStudies
2017-01-01 | S-EPMC5386594 | BioStudies
2011-01-01 | S-EPMC3177189 | BioStudies
2018-01-01 | S-EPMC6256539 | BioStudies