Regulation and characterization of mutants of fixABCX in Rhizobium leguminosarum

Webb, I., Xu, J. , Sanchez-Canizares, C., Karunakaran, R., Ramachandran, V., Rutten, P., East, A., Huang, W., Watmough, N. and Poole, P. (2021) Regulation and characterization of mutants of fixABCX in Rhizobium leguminosarum. Molecular Plant-Microbe Interactions, 34(10), pp. 1167-1180. (doi: 10.1094/mpmi-02-21-0037-r)

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 (PnifA1), as well as an additional basal constitutive promoter driving background expression of nifA (PnifA2). These were confirmed by 5′-end mapping of transcription start sites using differential RNA-seq. Complementation of polar fixAB and fixX mutants (Fix− strains) confirmed expression of nifA from PnifA1 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: Articles
• Additional 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.
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Xu, Dr Jiabao
• Authors: Webb, I., Xu, J., Sanchez-Canizares, C., Karunakaran, R., Ramachandran, V., Rutten, P., East, A., Huang, W., Watmough, N., and Poole, P.
• College/School: College of Science and Engineering > School of Engineering > Biomedical Engineering
• Journal Name: Molecular Plant-Microbe Interactions
• Publisher: American Phytopathological Society
• ISSN: 0894-0282
• ISSN (Online): 1943-7706
• Published Online: 28 October 2021
• Copyright Holders: Copyright © 2021 The Author(s).
• First Published: First published in Molecular Plant-Microbe Interactions 34(10):1167-1180
• Publisher Policy: Reproduced under a Creative Commons license