Complex Assembly and Metabolic Profiling of Arabidopsis thaliana Plants Overexpressing Vitamin B6 Biosynthesis Proteins

Abstract

In plants, vitamin B₆ biosynthesis requires the activity of PDX1 and PDX2 proteins. Arabidopsis thaliana encodes for three PDX1 proteins, named PDX1.1, 1.2, and 1.3, but only one PDX2. Here, we show in planta complex assembly of PDX proteins, based on split-YFP and FPLC assays, and can demonstrate their presence in higher complexes of around 750 kDa. Metabolic profiling of plants ectopically expressing the different PDX proteins indicates a negative influence of PDX1.2 on vitamin B₆ biosynthesis and a correlation between aberrant vitamin B6 content, PDX1 gene expression, and light sensitivity specifically for PDX1.3. These findings provide first insights into in planta vitamin B₆ synthase complex assembly and new information on how the different PDX proteins affect plant metabolism.

Balancing of B6 Vitamers Is Essential for Plant Development and Metabolism in Arabidopsis

“…Intriguingly, most of the metabolite compounds detected were increased rather than decreased. This is similar to what has been observed in other vitamin B 6 biosynthesis mutants with the exception of the amino acid glycine, which has been observed to decrease in other mutants of this pathway (Wagner et al, 2006;Raschke et al, 2011), although glycine accumulation has been reported upon overexpression of the de novo biosynthesis gene PDX1.3 (Leuendorf et al, 2010). Interestingly, we did not observe substantial changes in the expression of the majority of the other genes involved in the biosynthesis of PLP, such as PDX1.1/ PDX1.3 and PDX2 involved in biosynthesis de novo (TambascoStudart et al, 2005) or the salvage pathway kinase SOS4 Sang et al, 2007).…”

The Pseudoenzyme PDX1.2 Boosts Vitamin B6 Biosynthesis under Heat and Oxidative Stress in Arabidopsis

“…Previous observations based on yeast two-hybrid and split YFP data indicated that the Arabidopsis PDX1 proteins can form heteromers, but the nature of these complexes and a physiological reason for their existence were not established (26,27). Therefore, we were prompted to investigate the PDX1 proteins in more detail and our own hypothesis concerning the possible function of PDX1.2 as a regulator of vitamin B 6 biosynthesis.…”

“…In this context, it is intriguing that such a seemingly inactive homolog of PDX1 (annotated as PDX1.2 in Arabidopsis) is present among its catalytic counterparts in certain plant species. Moreover, this paralog is known to interact and form complexes with its catalytic paralogs (26,27). However, the precise nature of these complexes and whether PDX1.2 has any function have remained elusive to date.…”

Vitamin Deficiencies in Humans: Can Plant Science Help?

“…In plants, while activity of the de novo pathway and the kinase appear to be cytosolic (although additional localizations have been visualized) (Shi et al, 2002;Chen and Xiong, 2005;Tambasco-Studart et al, 2005), the oxidase has been shown to be restricted to plastids (Sang et al, 2011). A few recent studies in Arabidopsis have independently demonstrated that overexpressing the biosynthetic enzymes (Chen and Xiong, 2009;Leuendorf et al, 2010;Raschke et al, 2011) leads to increased accumulation of vitamin B 6 . The most recent of these studies, which demonstrated the highest boost in B 6 content, showed that such an increase can lead to larger organ size and, thus, potentially higher yield rates (Raschke et al, 2011).…”