Regulation and Characterization of Mutants of fixABCX in Rhizobium leguminosarum

Webb, Isabel U. C., Xu, Jiabao, Sánchez-Cañizares, Carmen, Karunakaran, Ramakrishnan, Ramachandran, Vinoy K., Rutten, Paul J., East, Alison K., Huang, Wei E., Watmough, Nicholas J. and Poole, Philip S. (2021) Regulation and Characterization of Mutants of fixABCX in Rhizobium leguminosarum. Molecular Plant-Microbe Interactions, 34 (10). ISSN 0894-0282

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Abstract

Symbiosis between Rhizobium leguminosarum and Pisum sativum requires tight control of redox balance in order to maintain respiration under the microaerobic conditions required for nitrogenase while still producing the eight electrons and sixteen molecules of ATP needed for nitrogen fixation. FixABCX, a cluster of electron transfer flavoproteins essential for nitrogen fixation, is encoded on the Sym plasmid (pRL10), immediately upstream of nifA, which encodes the general transcriptional regulator of nitrogen fixation. There is a symbiotically regulated NifA-dependent promoter upstream of fixA (P nifA1), as well as an additional basal constitutive promoter driving background expression of nifA (P nifA2). These were confirmed by 59-end mapping of transcription start sites using differential RNA-seq. Complementation of polar fixAB and fixX mutants (Fix - strains) confirmed expression of nifA from P nifA1 in symbiosis. Electron microscopy combined with single-cell Raman microspectroscopy characterization of fixAB mutants revealed previously unknown heterogeneity in bacteroid morphology within a single nodule. Two morphotypes of mutant fixAB bacteroids were observed. One was larger than wild-type bacteroids and contained high levels of polyhydroxy-3-butyrate, a complex energy/reductant storage product. A second bacteroid phenotype was morphologically and compositionally different and resembled wild-type infection thread cells. From these two characteristic fixAB mutant bacteroid morphotypes, inferences can be drawn on the metabolism of wild-type nitrogen-fixing bacteroids.

Item Type: Article
Additional Information: Funding Information: Funding: This research was funded by Engineering and Physical Sciences Research Council grants EP/M002403/1 and EP/M02833X/1, Biotechnology and Biological Sciences Research Council grants BB/ J014524/1, BB/M011224/1, and BB/N003608/1, and Natural Environment Research Council grant NE/M002934/1. Publisher Copyright: Copyright © 2021 The Author(s).
Uncontrolled Keywords: bacteroid morphology,drna-seq,electron transfer flavoproteins,fixabcx,nifa regulation,nitrogen fixation,phb,polyhydroxy-3-butyrate,redox balance,rhizobium leguminosarum bv. viciae,single-cell raman microspectroscopy (scrm),physiology,agronomy and crop science ,/dk/atira/pure/subjectarea/asjc/1300/1314
Faculty \ School: Faculty of Science > School of Biological Sciences
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Depositing User: LivePure Connector
Date Deposited: 13 Nov 2021 01:51
Last Modified: 22 Nov 2021 01:37
URI: https://ueaeprints.uea.ac.uk/id/eprint/82089
DOI: 10.1094/MPMI-02-21-0037-R

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