Function of plastidial pyruvate kinases in seeds of Arabidopsis thaliana^†

Abstract

Summary Pyruvate kinase (PK) catalyses the irreversible synthesis of pyruvate and ATP, which are both used in multiple biochemical pathways. These compounds are essential for sustained fatty acid production in the plastids of maturing Arabidopsis embryos. Using a real‐time quantitative reverse transcriptase (RT)‐PCR approach, the three genes encoding putative plastidial PKs (PKps) in Arabidopsis, namely PKp1 (At3g22960), PKp2 (At5g52920) and PKp3 (At1g32440), were shown to be ubiquitously expressed. However, only PKp1 and PKp2 exhibited significant expression in maturing seeds. The activity of PKp1 and PKp2 promoters was consistent with this pattern, and the study of the PKp1:GFP and PKp2:GFP fusion proteins confirmed the plastidial localization of these enzymes. To further investigate the function of these two PKp isoforms in seeds comprehensive functional analyses were carried out, including the cytological, biochemical and molecular characterization of two pkp1 and two pkp2 alleles, together with a pkp1pkp2 double mutant. The results obtained outlined the importance of these PKps for fatty acid synthesis and embryo development. Mutant seeds were depleted of oil, their fatty acid content was drastically modified, embryo elongation was retarded and, finally, seed germination was also affected. Together, these results provide interesting insights concerning the carbon fluxes leading to oil synthesis in maturing Arabidopsis seeds. The regulation of this metabolic network by the WRINKLED1 transcription factor is discussed, and emphasizes the role of plastidial metabolism and the importance of its tight regulation.

A comparative transcriptomic approach to understanding the formation of cork

“…Some genes related to these categories, such as the putative orthologs of Arabidopsis sucrose synthase SUS4 and the plastidial pyruvate kinase PKp1, are clearly upregulated in the cork oak outer bark (Table S6B). In fact the plastidial pyruvate kinase has been involved in fatty acid synthesis in Arabidopsis seeds (Andre et al 2007;Baud et al 2007) and sucrose synthase is involved in oil accumulation, suggesting a coordination between glycolytic and fatty acid biosynthetic pathways (Troncoso-Ponce et al 2011). However, the upregulation of sucrose synthases in cork could also be a consequence of a higher demand of cell wall synthesis (Andersson-Gunnerås et al 2006;Wei et al 2015).…”

Microalgal triacylglycerols as feedstocks for biofuel production: perspectives and advances

“…In the chloroplast, photosynthesis provides an endogenous source of acetyl CoA, and more than one pathway may contribute to maintaining the acetyl CoA pool. In oil seed plants, a major route of carbon flux to fatty acid synthesis may involve cytosolic glycolysis to phosphoenolpyruvate (PEP), which is then preferentially transported from the cytosol to the plastid, where it is converted to pyruvate and consequently to acetyl CoA (Baud et al, 2007;Ruuska et al, 2002;Schwender and Ohlrogge, 2002). In green algae, as glycolysis and pyruvate kinase (PK), which catalyzes the irreversible synthesis of pyruvate from PEP, occur in the chloroplast in addition to the cytosol (Andre et al, 2007), it is possible that glycolysis-derived pyruvate is the major photosynthate to be converted to acetyl CoA for de novo fatty acid synthesis.…”

Oil and Protein Accumulation in Developing Seeds Is Influenced by the Expression of a Cytosolic Pyrophosphatase in Arabidopsis

“…Characterization of a recessive high-oil mutant has led to the identification of maternally controlled deposition polysaccharides of the seed coat mucilage as a carbon sink that diverts carbon from seed storage lipid biosynthesis (Shen et al, 2006;Shi et al, 2012). Transfer (T)-DNA-tagged alleles of plastidic subunits of pyruvate kinase are associated with low seed oil content and have demonstrated the importance of plastidic pyruvate supply in seed storage lipid biosynthesis (Andre et al, 2007;Baud et al, 2007b). Defects in plastidic starch biosynthesis, due to recessive loss-of-function mutations of plastidic phosphoglucomutase, show low seed oil content, thus emphasizing the importance of transient starch as a carbon source supporting seed oil biosynthesis in Arabidopsis (Periappuram et al, 2000).…”