Essential roles for imuA'- and imuB-encoded accessory factors in DnaE2-dependent mutagenesis in Mycobacterium tuberculosis.

Medical Research Council/National Health Laboratory Service/University of the Witwatersrand Molecular Mycobacteriology Research Unit and Department of Science and Technology/National Research Foundation Centre of Excellence for Biomedical Tuberculosis Research, Faculty of Health Sciences, University of the Witwatersrand and the National Health Laboratory Service, Johannesburg 2000, South Africa.
Proceedings of the National Academy of Sciences (Impact Factor: 9.81). 07/2010; 107(29):13093-8. DOI: 10.1073/pnas.1002614107
Source: PubMed

ABSTRACT In Mycobacterium tuberculosis (Mtb), damage-induced mutagenesis is dependent on the C-family DNA polymerase, DnaE2. Included with dnaE2 in the Mtb SOS regulon is a putative operon comprising Rv3395c, which encodes a protein of unknown function restricted primarily to actinomycetes, and Rv3394c, which is predicted to encode a Y-family DNA polymerase. These genes were previously identified as components of an imuA-imuB-dnaE2-type mutagenic cassette widespread among bacterial genomes. Here, we confirm that Rv3395c (designated imuA') and Rv3394c (imuB) are individually essential for induced mutagenesis and damage tolerance. Yeast two-hybrid analyses indicate that ImuB interacts with both ImuA' and DnaE2, as well as with the beta-clamp. Moreover, disruption of the ImuB-beta clamp interaction significantly reduces induced mutagenesis and damage tolerance, phenocopying imuA', imuB, and dnaE2 gene deletion mutants. Despite retaining structural features characteristic of Y-family members, ImuB homologs lack conserved active-site amino acids required for polymerase activity. In contrast, replacement of DnaE2 catalytic residues reproduces the dnaE2 gene deletion phenotype, strongly implying a direct role for the alpha-subunit in mutagenic lesion bypass. These data implicate differential protein interactions in specialist polymerase function and identify the split imuA'-imuB/dnaE2 cassette as a compelling target for compounds designed to limit mutagenesis in a pathogen increasingly associated with drug resistance.

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