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J. Biol. Chem., Vol. 283, Issue 9, 5708-5718, February 29, 2008
A Temperature-sensitive Mutation in the Arabidopsis thaliana Phosphomannomutase Gene Disrupts Protein Glycosylation and Triggers Cell Death*![]() 1![]() 12![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() 17![]() 1
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Eukaryotic phosphomannomutases (PMMs) catalyze the interconversion of mannose 6-phosphate to mannose 1-phosphate and are essential to the biosynthesis of GDP-mannose. As such, plant PMMs are involved in ascorbic acid (AsA) biosynthesis and N-glycosylation. We report on the conditional phenotype of the temperature-sensitive Arabidopsis thaliana pmm-12 mutant. Mutant seedlings were phenotypically similar to wild type seedlings when grown at 16–18 °C but died within several days after transfer to 28 °C. This phenotype was observed throughout both vegetative and reproductive development. Protein extracts derived from pmm-12 plants had lower PMM protein and enzyme activity levels. In vitro biochemical analysis of recombinant proteins showed that the mutant PMM protein was compromised in its catalytic efficiency (Kcat/Km). Despite significantly decreased AsA levels in pmm-12 plants, AsA deficiency could not account for the observed phenotype. Since, at restrictive temperature, total glycoprotein patterns were altered and glycosylation of protein-disulfide isomerase was perturbed, we propose that a deficiency in protein glycosylation is responsible for the observed cell death phenotype.
Received for publication, June 18, 2007 , and in revised form, December 13, 2007. * This work was supported by Interuniversity Poles of Attraction Programme-Belgian Science Policy Grant P5/13, Research Fund of Ghent University Geconcerteerde Onderzoeksacties Grant 12051403, and Research Foundation-Flanders Grant G.0350.04. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 These authors contributed equally to this work. 2 Recipient of a predoctoral fellowship of the Institute for the Promotion of Innovation by Science and Technology in Flanders. Present address: Ablynx N.V., 9052 Gent, Belgium. 3 Present address: Lumora Ltd., Ely CB7 4ET, United Kingdom. 4 Present address: Dept. of Psychiatry and Behavioral Sciences, Stanford University, Palo Alto, CA 94304-5742. 5 Present address: Dept. of Pathology, University Hospital of Liège, 4000 Liège, Belgium. 6 Present address: School of Agriculture, Food and Rural Development, University of Newcastle upon Tyne, Newcastle upon Tyne NE1 7RU, United Kingdom. 7 To whom correspondence should be addressed: Dept. of Plant Systems Biology, VIB, Ghent University, Technologiepark 927, 9052 Gent, Belgium. Tel.: 32-9-3313800; Fax: 32-9-3313809; E-mail: frank.vanbreusegem{at}psb.ugent.be.
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