Antisense inhibition of the plastidial glucose‐6‐phosphate/phosphate translocator inViciaseeds shifts cellular differentiation and promotes protein storage

Abstract

SummaryThe glucose-6-phosphate/phosphate translocator (GPT) acts as an importer of carbon into the plastid. Despite the potential importance of GPT for storage in crop seeds, its regulatory role in biosynthetic pathways that are active during seed development is poorly understood. We have isolated GPT1 from Vicia narbonensis and studied its role in seed development using a transgenic approach based on the seed-specific legumin promoter LeB4. GPT1 is highly expressed in vegetative sink tissues, flowers and young seeds. In the embryo, localized upregulation of GPT1 at the onset of storage coincides with the onset of starch accumulation. Embryos of transgenic plants expressing antisense GPT1 showed a significant reduction (up to 55%) in the specific transport rate of glucose-6-phosphate as determined using proteoliposomes prepared from embryos. Furthermore, amyloplasts developed later and were smaller in size, while the expression of genes encoding plastid-specific translocators and proteins involved in starch biosynthesis was decreased. Metabolite analysis and stable isotope labelling demonstrated that starch biosynthesis was also reduced, although storage protein biosynthesis increased. This metabolic shift was characterized by upregulation of genes related to nitrogen uptake and protein storage, morphological variation of the protein-storing vacuoles, and a crude protein content of mature seeds of transgenics that was up to 30% higher than in wild-type. These findings provide evidence that (1) the prevailing level of GPT1 abundance/activity is rate-limiting for the synthesis of starch in developing seeds, (2) GPT1 exerts a controlling function on assimilate partitioning into storage protein, and (3) GPT1 is essential for the differentiation of embryonic plastids and seed maturation.

Comparative transcriptome and metabolite analysis of oil palm and date palm mesocarp that differ dramatically in carbon partitioning

“…6 B and C), and represented one of the highest EST counts of all carbohydrate-related genes of oil palm mesocarp. Interestingly, antisense inhibition of a GPT transporter in Vicia seeds altered carbon partitioning and promoted protein storage at the expense of carbohydrates (25). ESTs for the phosphoenolpyruvate (PEP) transporter PPT1 were 3.7 times higher than in date palm and increased fourfold during oil palm ripening (Figs.…”

The Import and Export Business in Plastids: Transport Processes across the Inner Envelope Membrane

“…A detailed analysis of heterozygous GPT1/gpt1 plants revealed that the development of both the female and the male gpt1 gametophyte are impaired. Two more recent investigations showed that GPT1 is also involved in embryo development and seed maturation (Rolletschek et al, 2007;Andriotis et al, 2010). A reduction of GPT1 expression of about 50% in seeds of Vicia narbonensis by an antisense approach with a seed-specific promoter led to a shift in assimilate partitioning from starch/ lipids into storage proteins, indicating that GPT1 exerts a significant control on seed filling and maturation (Rolletschek et al, 2007).…”

“…Two more recent investigations showed that GPT1 is also involved in embryo development and seed maturation (Rolletschek et al, 2007;Andriotis et al, 2010). A reduction of GPT1 expression of about 50% in seeds of Vicia narbonensis by an antisense approach with a seed-specific promoter led to a shift in assimilate partitioning from starch/ lipids into storage proteins, indicating that GPT1 exerts a significant control on seed filling and maturation (Rolletschek et al, 2007). A reduction of more than 80% of GPT1 expression in Arabidopsis seeds is lethal because embryo development stops at the globular stage (Andriotis et al, 2010).…”

Starch Turnover and Metabolism during Flower and Early Embryo Development

“…Accumulating evidence in all dicotyledonous plant species studied so far (e.g. Arabidopsis, broad bean [Vicia faba], and potato [Solanum tuberosum]) points to Glc-6-P as the preferred translocated hexose phosphate and the Glc-6-P/phosphate antiporter GPT1 as the active translocator (Kammerer et al, 1998;Geigenberger et al, 2004;Niewiadomski et al, 2005;Rolletschek et al, 2007;Zhang et al, 2008). This is further supported by the fact that the loss of plastidial PGI1 activity impacts starch accumulation only in autotrophic Arabidopsis cells (Yu et al, 2000;Tsai et al, 2009), while mutations affecting plastidial PGM1 result in a low-starch phenotype in both autotrophic and heterotrophic tissues (Caspar et al, 1985).…”