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Title: Tn-seq of Caulobacter crescentus under uranium stress reveals genes essential for detoxification and stress tolerance

Ubiquitous aquatic bacterium Caulobacter crescentus is highly resistant to uranium (U) and facilitates U biomineralization and thus holds promise as an agent of U bioremediation. In order to gain an understanding of how C. crescentus tolerates U, we employed transposon (Tn) mutagenesis paired with deep sequencing (Tn-seq) in a global screen for genomic elements required for U resistance. Of the 3,879 annotated genes in the C. crescentus genome, 37 were found to be specifically associated with fitness under U stress, 15 of which were subsequently tested through mutational analysis. Systematic deletion analysis revealed that mutants lacking outer membrane transporters (rsaFa and rsaFb), a stress-responsive transcription factor (cztR), or a ppGpp synthetase/hydrolase (spoT) exhibited a significantly lower survival rate under U stress. RsaFa and RsaFb, which are homologues of TolC in Escherichia coli, have previously been shown to mediate S-layer export. Transcriptional analysis revealed upregulation of rsaFa and rsaFb by 4- and 10-fold, respectively, in the presence of U. We additionally show that rsaFa mutants accumulated higher levels of U than the wild type, with no significant increase in oxidative stress levels. These results suggest a function for RsaFa and RsaFb in U efflux and/or maintenance of membrane integrity during Umore » stress. In addition, we present data implicating CztR and SpoT in resistance to U stress. Together, our findings reveal novel gene targets that are key to understanding the molecular mechanisms of U resistance in C. crescentus.« less
Authors:
 [1] ;  [1] ;  [1] ;  [2] ;  [2] ;  [3] ;  [4] ;  [5] ;  [6] ;  [7] ;  [1]
  1. Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
  2. DOE Joint Genome Inst., Walnut Creek, CA (United States)
  3. Univ. of British Columbia, Vancouver, BC (Canada)
  4. California State Univ., Northridge, CA (United States)
  5. Stanford Univ. School of Medicine, Stanford, CA (United States)
  6. ETH Zurich (Switzerland)
  7. Univ. of Wyoming, Laramie, WY (United States)
Publication Date:
OSTI Identifier:
1234593
Report Number(s):
LLNL-JRNL--670205
Journal ID: ISSN 0021-9193; TRN: US1600611
Grant/Contract Number:
AC52-07NA27344
Type:
Accepted Manuscript
Journal Name:
Journal of Bacteriology
Additional Journal Information:
Journal Volume: 197; Journal Issue: 19; Journal ID: ISSN 0021-9193
Publisher:
American Society for Microbiology
Research Org:
Lawrence Livermore National Laboratory (LLNL), Livermore, CA (United States)
Sponsoring Org:
USDOE
Country of Publication:
United States
Language:
English
Subject:
63 RADIATION, THERMAL, AND OTHER ENVIRONMENTAL POLLUTANT EFFECTS ON LIVING ORGANISMS AND BIOLOGICAL MATERIALS; 59 BASIC BIOLOGICAL SCIENCES