TY - JOUR
T1 - Regulation of nitrous oxide reductase genes by NasT-mediated transcription antitermination in Bradyrhizobium diazoefficiens
AU - Sanchez Gomez, Cristina
AU - Mitsui, Hisayuki
AU - Minamisawa, Kiwamu
N1 - Funding Information:
We thank M. Hidaka and T. Uchida (Graduate School of Agricultural Science, Tohoku University, Sendai, Japan) for kindly providing purified GST and GST–NasT proteins and for discussion regarding the manuscript. This work was supported by a Grant-in-Aid for Scientific Research (A) 26252065 from the Ministry of Education, Culture, Sports, Science and Technology of Japan, and by a grant from Science and Technology Research Promotion Program for Agriculture, Forestry, Fisheries and Food Industry. All authors have approved the manuscript and declare no conflict of interest.
Publisher Copyright:
© 2017 Society for Applied Microbiology and John Wiley & Sons Ltd
PY - 2017/8
Y1 - 2017/8
N2 - In Bradyrhizobium diazoefficiens, maximal expression of the nitrous oxide reductase gene (nosZ) requires oxygen limitation and the presence of a nitrogen oxide. The putative transcription antiterminator NasT is a positive regulator of nosZ; but in the absence of nitrate, NasT is counteracted by the nitrate sensor NasS. Here, we examined the NasT-mediated mechanism of nosRZDFYLX gene cluster expression. We mapped two transcription start sites of nosR and identified two potential hairpins, H1 and H2, within the 5′-leader of nosR transcripts. Electrophoretic mobility shift assay showed that NasT specifically bound the nosR-leader RNA and deletion of H1 abolished such binding. Under aerobic nitrate-deficient conditions, deletion of H1 or H2 increased the level of nosRZD transcripts. Under denitrifying conditions (anaerobiosis with nitrate supply), the level of nosRZD transcripts was severely impaired in the nasT mutant; in the nasT background, deletions of either hairpin led to increased level of nosRZD transcripts. In contrast to nosRZD coding region, nosR-leader transcript level was not affected by nasS or nasT mutations under aerobic or denitrifying conditions respectively. These results suggest that the two-hairpin RNA structure acts for transcription termination upstream of nosR and the binding of NasT to H1 facilitates read-through transcription to induce nos expression.
AB - In Bradyrhizobium diazoefficiens, maximal expression of the nitrous oxide reductase gene (nosZ) requires oxygen limitation and the presence of a nitrogen oxide. The putative transcription antiterminator NasT is a positive regulator of nosZ; but in the absence of nitrate, NasT is counteracted by the nitrate sensor NasS. Here, we examined the NasT-mediated mechanism of nosRZDFYLX gene cluster expression. We mapped two transcription start sites of nosR and identified two potential hairpins, H1 and H2, within the 5′-leader of nosR transcripts. Electrophoretic mobility shift assay showed that NasT specifically bound the nosR-leader RNA and deletion of H1 abolished such binding. Under aerobic nitrate-deficient conditions, deletion of H1 or H2 increased the level of nosRZD transcripts. Under denitrifying conditions (anaerobiosis with nitrate supply), the level of nosRZD transcripts was severely impaired in the nasT mutant; in the nasT background, deletions of either hairpin led to increased level of nosRZD transcripts. In contrast to nosRZD coding region, nosR-leader transcript level was not affected by nasS or nasT mutations under aerobic or denitrifying conditions respectively. These results suggest that the two-hairpin RNA structure acts for transcription termination upstream of nosR and the binding of NasT to H1 facilitates read-through transcription to induce nos expression.
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U2 - 10.1111/1758-2229.12543
DO - 10.1111/1758-2229.12543
M3 - Article
C2 - 28474433
AN - SCOPUS:85024830527
VL - 9
SP - 389
EP - 396
JO - Environmental Microbiology Reports
JF - Environmental Microbiology Reports
SN - 1758-2229
IS - 4
ER -