Characterising the functional importance of folate polyglutamation in plants (PhD)

Folate cannot be synthesized by humans and plant food is the main source of this essential vitamin. Folate deficiency can have severe health consequences such as birth defects e.g. spina bifida, and marginal deficiency may impact on cardiovascular vascular diseases and certain cancers. Research suggests that folate intake needs to increase to improve human health. In the USA, cereal-based products are fortified with folic acid and there has been a recommendation for its introduction into the UK. There is some concern over fortification with folic acid since this is not the “natural form” of folate. This project aimed to establish the potential to harness natural variation to enhance folate content in wheat.

The potential for natural variation was demonstrated by a three fold range of folate levels within a population of the model plant Arabidopsis. The total folate levels were then determined in the seed from a range of elite wheat lines. In this case there was minimal natural variation with a range of folate of between 40 and 55μg.100g-1. The term folate encompasses a wide range of related compounds. The project developed a novel technique to profile these folates. The method was able to differentiate and quantify around 20 individual folates in Arabidopsis leaf samples. When applied to folates from wheat seed, it proved possible to separate and quantify several mono-glutamated folates but there was no evidence for any folates with higher levels of glutamation as found in the leaf tissue. The effect of partial germination on the folate content of the wheat seed was also determined. Germination resulted in a significant increase in total folate, and in particular in the levels of 5-methyl tetrahydrofolate, but again only mono-glutamated forms were detected. An investigation into the physiological role of polyglutamation was thus carried out using Arabidopsis. Three isoforms of folylpolyglutamate synthase (FPGS) were identified - cytoplasmic (cFPGS), mitochondrial (mFPGS) or plastid (pFPGS). The expression of these within the plant and the effect of knocking out their activity on growth and folate metabolism was examined. Spatial expression, particularly for cFPGS and pFPGS was observed. There was minimal phenotypic response to knocking these enzymes out individually, although silencing of pFPGS appears to have the most significant impact on the accumulation of polyglutamated forms of folate.