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BIOCHEMICAL PROCESSES AND MACROMOLECULAR STRUCTURES

Analysis of Metabolic Flux Phenotypes for Two Arabidopsis Mutants with Severe Impairment in Seed Storage Lipid Synthesis^1,[W],[OA]

Biology Department, Brookhaven National Laboratory, Upton, New York 11973

Major storage reserves of Arabidopsis (Arabidopsis thaliana) seeds are triacylglycerols (seed oils) and proteins. Seed oil content is severely reduced for the regulatory mutant wrinkled1 (wri1-1; At3g54320) and for a double mutant in two isoforms of plastidic pyruvate kinase (pkpβ₁pkp; At5g52920 and At3g22960). Both already biochemically well-characterized mutants were now studied by ¹³C metabolic flux analysis of cultured developing embryos based on comparison with their respective genetic wild-type backgrounds. For both mutations, in seeds as well as in cultured embryos, the oil fraction was strongly reduced while the fractions of proteins and free metabolites increased. Flux analysis in cultured embryos revealed changes in nutrient uptakes and fluxes into biomass as well as an increase in tricarboxylic acid cycle activity for both mutations. While in both wild types plastidic pyruvate kinase (PK_p) provides most of the pyruvate for plastidic fatty acid synthesis, the flux through PK_p is reduced in pkpβ₁pkp by 43% of the wild-type value. In wri1-1, PK_p flux is even more reduced (by 82%), although the genes PKpβ₁ and PKp are still expressed. Along a common paradigm of metabolic control theory, it is hypothesized that a large reduction in PK_p enzyme activity in pkpβ₁pkp has less effect on PK_p flux than multiple smaller reductions in glycolytic enzymes in wri1-1. In addition, only in the wri1-1 mutant is the large reduction in PK_p flux compensated in part by an increased import of cytosolic pyruvate and by plastidic malic enzyme. No such limited compensatory bypass could be observed in pkpβ₁pkp.

The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) is: Jörg Schwender (schwend{at}bnl.gov).

^[W] The online version of this article contains Web-only data.

^[OA] Open Access articles can be viewed online without a subscription.

www.plantphysiol.org/cgi/doi/10.1104/pp.109.144121

^* Corresponding author; e-mail schwend{at}bnl.gov.

Received July 1, 2009; accepted September 7, 2009; published September 15, 2009.