Highly Divergent Methyltransferases Catalyze a Conserved Reaction in Tocopherol and Plastoquinone Synthesis in Cyanobacteria and Photosynthetic Eukaryotes

Abstract

Tocopherols are lipid-soluble compounds synthesized only by photosynthetic eukaryotes and oxygenic cyanobacteria. The pathway and enzymes for tocopherol synthesis are homologous in cyanobacteria and plants except for 2-methyl-6-phytyl-1,4-benzoquinone/2-methyl-6-solanyl-1,4-benzoquinone methyltransferase (MPBQ/MSBQ MT), which catalyzes a key methylation step in both tocopherol and plastoquinone (PQ) synthesis. Using a combined genomic, genetic, and biochemical approach, we isolated and characterized the VTE3 (vitamin E defective) locus, which encodes MPBQ/MSBQ MT in Arabidopsis. The phenotypes of vte3 mutants are consistent with the disruption of MPBQ/MSBQ MT activity to varying extents. The ethyl methanesulfonate-derived vte3-1 allele alters tocopherol composition but has little impact on PQ levels, whereas the null vte3-2 allele is deficient in PQ and ␣ -and ␥ -tocopherols. In vitro enzyme assays confirmed that VTE3 is the plant functional equivalent of the previously characterized MPBQ/MSBQ MT (Sll0418) from Synechocystis sp PCC6803, although the two proteins are highly divergent in primary sequence. Sll0418 orthologs are present in all fully sequenced cyanobacterial genomes, Chlamydomonas reinhardtii , and the diatom Thalassiosira pseudonana but absent from vascular and nonvascular plant databases. VTE3 orthologs are present in all vascular and nonvascular plant databases and in C. reinhardtii but absent from cyanobacterial genomes. Intriguingly, the only prokaryotic genomes that contain VTE3-like sequences are those of two species of archea, suggesting that, in contrast to all other enzymes of the plant tocopherol pathway, the evolutionary origin of VTE3 may have been archeal rather than cyanobacterial. In vivo analyses of vte3 mutants and the corresponding homozygous Synechocystis sp PCC6803 sll0418 :: aphII mutant revealed important differences in enzyme redundancy, the regulation of tocopherol synthesis, and the integration of tocopherol and PQ biosynthesis in cyanobacteria and plants.

RNAi-Mediated Tocopherol Deficiency Impairs Photoassimilate Export in Transgenic Potato Plants

“…However, most of our understanding of tocopherol functions has been derived from nonplant systems, since tocopherols (vitamin E) are essential dietary components for humans and other mammals. Recent efforts to identify the enzymatic steps involved in the tocopherol biosynthetic pathway and to improve vitamin E contents in crop plants by metabolic engineering have provided a number of different mutant and transgenic plants for the analysis of tocopherol function in planta (Shintani and DellaPenna, 1998;DellaPenna, 2001, 2003;Porfirova et al, 2002;Savidge et al, 2002;Bergmü ller et al, 2003;Cheng et al, 2003;Van Eenennaam et al, 2003). In this respect, it was discovered that the maize sxd1 mutant, originally isolated based on a block in Suc transport from leaves (Russin et al, 1996;Provencher et al, 2001), and the Arabidopsis vte1 mutant were both defective in the TC gene, resulting in a complete lack of tocopherols (Porfirova et al, 2002;Sattler et al, 2003).…”

Protein Profiling of Plastoglobules in Chloroplasts and Chromoplasts. A Surprising Site for Differential Accumulation of Metabolic Enzymes

“…A possible explanation is that accumulation of phytol diphosphate leads to feedback regulation of the nonmevalonate pathway. It appears that the biosynthetic pathway of plastoquinone is rather simple and seems resolved (Cheng et al, 2003;Sattler et al, 2004). In contrast, several enzymes in the phylloquinone pathway have not yet been identified.…”

Tocopherols Play a Crucial Role in Low-Temperature Adaptation and Phloem Loading in Arabidopsis

“…Although plausible, such hypotheses are based primarily on correlations and circumstantial evidence and have yet to be rigorously tested in planta. The isolation of Arabidopsis thaliana mutants disrupting steps of the tocopherol biosynthetic pathway provides powerful tools with which to directly investigate tocopherol functions in plants (Figure 1) (Shintani and DellaPenna, 1998;Collakova and DellaPenna, 2001;Porfirova et al, 2002;Cheng et al, 2003;Sattler et al, 1 To whom correspondence should be addressed. E-mail dellapen@ msu.edu; fax 517-353-9334.…”