Thiamin Metabolism in Plants: Elucidation of the 4-Methyl-5-(b-hydroxyethyl)Thiazole Phosphate (HET) Biosynthetic Pathway in Arabidopsis

Board Of Regents, Nshe, Obo University Of Nevada, Reno, Reno NV

Investigators

Abstract

Thiamin pyrophosphate (TPP), which is formed through the condensation of two major intermediates, 4-methyl-5-(b-hydroxyethyl)thiazole phosphate (HET) and 2-methyl-4-amino-5-hydroxymethylpyrimidine pyrophosphate (HMP), is an essential enzyme co-factor required for the viability of all organisms. While TPP is only synthesized by bacteria, fungi and photosynthetic organisms, animals and humans must derive TPP from dietary sources of thiamin. Although plants represent a major source of dietary thiamin, surprisingly little is understood about most aspects of its synthesis in plants. This is particularly true for the HET intermediate of TPP. Most of what is known about HET biosynthesis has been derived from genetic and biochemical studies in microorganisms where this compound is derived from different synthetic precursors using different enzymatic machinery. Although a previous study suggested that HET is synthesized as described for E. coli, where deoxy-xylulose-5-phosphate (DXP) is a precursor, the Shintani lab has performed studies showing that plants can synthesize HET in the absence of DXP. This result suggests that plants may possess an alternative pathway to HET that exists in place of or in addition to the previously described "E. coli-like" pathway. The overall goal of this research is to use a combination of molecular and biochemical approaches to determine the biosynthetic pathway(s) leading to formation of HET in plants. To achieve this goal the following research objectives will be performed: 1) Determine the biosynthetic precursors leading to thiamin biosynthesis in plants, 2) Determine the role of the plant orthologue of the yeast Thi4 protein in HET biosynthesis, 3) Identify and functionally analyze additional Arabidopsis HET biosynthetic genes. Thiamin (Vitamin B1) deficiencies in humans can lead to a condition known as Beriberi that is manifested by severe neurological disorders and a general wasting phenomenon. This disease is primarily associated with poverty-stricken populations of developing countries whose diets subsist primarily of polished grain products such as polished rice or bleached wheat flour. A sustainable solution to thiamin deficiencies in humans would be to increase the nutritional content of staple food crops that endogenous populations of the world commonly consume. By genetic engineering such crops for increased thiamin, it should be possible to positively impact the nutritional needs of the global population. Unfortunately, the major impediment to such an effort is a current lack of knowledge pertaining to the biosynthesis of thiamin in plants. The increased biosynthetic knowledge obtained through these studies will be important for the rational design of crops engineered for elevated thiamin levels for improved human and animal nutrition.