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Applied and Environmental Microbiology
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Environmental Microbiology

Meta-Analyses of Dehalococcoides mccartyi Strain 195 Transcriptomic Profiles Identify a Respiration Rate-Related Gene Expression Transition Point and Interoperon Recruitment of a Key Oxidoreductase Subunit

Cresten B. Mansfeldt, Annette R. Rowe, Gretchen L. W. Heavner, Stephen H. Zinder, Ruth E. Richardson
F. E. Löffler, Editor
Cresten B. Mansfeldt
aSchool of Civil and Environmental Engineering, Cornell University, Ithaca, New York, USA
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Annette R. Rowe
bDepartment of Microbiology, Cornell University, Ithaca, New York, USA
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Gretchen L. W. Heavner
aSchool of Civil and Environmental Engineering, Cornell University, Ithaca, New York, USA
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Stephen H. Zinder
bDepartment of Microbiology, Cornell University, Ithaca, New York, USA
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Ruth E. Richardson
aSchool of Civil and Environmental Engineering, Cornell University, Ithaca, New York, USA
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F. E. Löffler
Roles: Editor
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DOI: 10.1128/AEM.02130-14
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ABSTRACT

A cDNA-microarray was designed and used to monitor the transcriptomic profile of Dehalococcoides mccartyi strain 195 (in a mixed community) respiring various chlorinated organics, including chloroethenes and 2,3-dichlorophenol. The cultures were continuously fed in order to establish steady-state respiration rates and substrate levels. The organization of array data into a clustered heat map revealed two major experimental partitions. This partitioning in the data set was further explored through principal component analysis. The first two principal components separated the experiments into those with slow (1.6 ± 0.6 μM Cl−/h)- and fast (22.9 ± 9.6 μM Cl−/h)-respiring cultures. Additionally, the transcripts with the highest loadings in these principal components were identified, suggesting that those transcripts were responsible for the partitioning of the experiments. By analyzing the transcriptomes (n = 53) across experiments, relationships among transcripts were identified, and hypotheses about the relationships between electron transport chain members were proposed. One hypothesis, that the hydrogenases Hup and Hym and the formate dehydrogenase-like oxidoreductase (DET0186-DET0187) form a complex (as displayed by their tight clustering in the heat map analysis), was explored using a nondenaturing protein separation technique combined with proteomic sequencing. Although these proteins did not migrate as a single complex, DET0112 (an FdhB-like protein encoded in the Hup operon) was found to comigrate with DET0187 rather than with the catalytic Hup subunit DET0110. On closer inspection of the genome annotations of all Dehalococcoides strains, the DET0185-to-DET0187 operon was found to lack a key subunit, an FdhB-like protein. Therefore, on the basis of the transcriptomic, genomic, and proteomic evidence, the place of the missing subunit in the DET0185-to-DET0187 operon is likely filled by recruiting a subunit expressed from the Hup operon (DET0112).

FOOTNOTES

    • Received 27 June 2014.
    • Accepted 17 July 2014.
    • Accepted manuscript posted online 25 July 2014.
  • Supplemental material for this article may be found at http://dx.doi.org/10.1128/AEM.02130-14.

  • Copyright © 2014, American Society for Microbiology. All Rights Reserved.
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Meta-Analyses of Dehalococcoides mccartyi Strain 195 Transcriptomic Profiles Identify a Respiration Rate-Related Gene Expression Transition Point and Interoperon Recruitment of a Key Oxidoreductase Subunit
Cresten B. Mansfeldt, Annette R. Rowe, Gretchen L. W. Heavner, Stephen H. Zinder, Ruth E. Richardson
Applied and Environmental Microbiology Sep 2014, 80 (19) 6062-6072; DOI: 10.1128/AEM.02130-14

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Meta-Analyses of Dehalococcoides mccartyi Strain 195 Transcriptomic Profiles Identify a Respiration Rate-Related Gene Expression Transition Point and Interoperon Recruitment of a Key Oxidoreductase Subunit
Cresten B. Mansfeldt, Annette R. Rowe, Gretchen L. W. Heavner, Stephen H. Zinder, Ruth E. Richardson
Applied and Environmental Microbiology Sep 2014, 80 (19) 6062-6072; DOI: 10.1128/AEM.02130-14
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