Kruse, I., MacLean, A. E., Hill, L. and Balk, J. (2018) Genetic dissection of cyclic pyranopterin monophosphate biosynthesis in plant mitochondria. Biochemical Journal, 475(2), pp. 495-509. (doi: 10.1042/BCJ20170559) (PMID:29247140) (PMCID:PMC5791162)
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Abstract
Mitochondria play a key role in the biosynthesis of two metal cofactors, iron–sulfur (FeS) clusters and molybdenum cofactor (Moco). The two pathways intersect at several points, but a scarcity of mutants has hindered studies to better understand these links. We screened a collection of sirtinol-resistant Arabidopsis thaliana mutants for lines with decreased activities of cytosolic FeS enzymes and Moco enzymes. We identified a new mutant allele of ATM3 (ABC transporter of the mitochondria 3), encoding the ATP-binding cassette transporter of the mitochondria 3 (systematic name ABCB25), confirming the previously reported role of ATM3 in both FeS cluster and Moco biosynthesis. We also identified a mutant allele in CNX2, cofactor of nitrate reductase and xanthine dehydrogenase 2, encoding GTP 3′,8-cyclase, the first step in Moco biosynthesis which is localized in the mitochondria. A single-nucleotide polymorphism in cnx2-2 leads to substitution of Arg88 with Gln in the N-terminal FeS cluster-binding motif. cnx2-2 plants are small and chlorotic, with severely decreased Moco enzyme activities, but they performed better than a cnx2-1 knockout mutant, which could only survive with ammonia as a nitrogen source. Measurement of cyclic pyranopterin monophosphate (cPMP) levels by LC–MS/MS showed that this Moco intermediate was below the limit of detection in both cnx2-1 and cnx2-2, and accumulated more than 10-fold in seedlings mutated in the downstream gene CNX5. Interestingly, atm3-1 mutants had less cPMP than wild type, correlating with previous reports of a similar decrease in nitrate reductase activity. Taken together, our data functionally characterize CNX2 and suggest that ATM3 is indirectly required for cPMP synthesis.
Item Type: | Articles |
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Additional Information: | Funding: I.K. was funded by a Dean’s studentship from the University of East Anglia and an Institute Development Grant from the John Innes Centre [CX410-J11A]. A.M. was funded by a John Innes Foundation studentship. The LC–MS/MS analysis was supported by a Biotechnology and Biological Sciences Research Council (BBSRC) Institute Strategic Grant [BB/P012523/1]. |
Status: | Published |
Refereed: | Yes |
Glasgow Author(s) Enlighten ID: | MacLean, Dr Andrew |
Authors: | Kruse, I., MacLean, A. E., Hill, L., and Balk, J. |
College/School: | College of Medical Veterinary and Life Sciences > School of Infection & Immunity |
Journal Name: | Biochemical Journal |
Publisher: | Portland Press |
ISSN: | 0264-6021 |
ISSN (Online): | 1470-8728 |
Published Online: | 15 December 2017 |
Copyright Holders: | Copyright © 2018 The Authors |
First Published: | First published in Biochemical Journal 475(2):495-509 |
Publisher Policy: | Reproduced under a Creative Commons license |
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