Thiamin diphosphate (TDP, vitamin B1) is an essential coenzyme present in all organisms. Animals obtain TDP from their diets, but plants synthesize TDP de novo. The vitamin B1 biosynthesis pathway was thought to be interconnected between chloroplasts and the cytosol in plants. A research team led by Hsieh Ming-Hsiun from the Institute of Plant and Microbial Biology at Academia Sinica has isolated a thiamin deficient mutant pale green1 (pale1) in the model plant Arabidopsis thaliana. The PALE1 gene encodes a thiamin monophosphate phosphatase that localizes in the mitochondrion. Therefore, a complete vitamin B1 biosynthesis pathway may involve the chloroplasts, mitochondria and cytosol.
TDP is an essential cofactor for many important enzymes involved in the catabolism of sugars and amino acids. In addition to its role in primary metabolism, TDP is also involved in tolerance to DNA damage, biotic and abiotic stresses. It is well established that thiamin monophosphate (TMP) is synthesized in the chloroplast. Instead of direct phosphorylation of TMP to form TDP, TMP is first dephosphorylated to thiamin. Thiamin is then pyrophosphorylated to TDP in the cytosol. Thus, it was expected that the TMP phosphatase involved in the dephosphorylation of TMP is most likely located in the chloroplast or in the cytosol.
The Arabidopsis pale1 mutant contains higher concentrations of TMP and less thiamin and TDP than the wild type. Supplementation with thiamin fully rescued the mutant phenotype, indicating that the pale1 mutant is a thiamin-deficient mutant. By map-based cloning and whole-genome sequencing, Dr. Hsieh’s group identified that the pale1 mutant has a mutation in a gene encoding TMP phosphatase of the vitamin B1 biosynthesis pathway. Dr. Hsieh’s group further demonstrated that the TMP phosphatase (PALE1) is localized to the mitochondrion. Thus, the biosynthesis of vitamin B1 in plant cells involves at least three different subcellular components, the chloroplasts, mitochondria, and cytosol.
With the complete vitamin B1 biosynthesis pathway genes in hand, it is now feasible to genetically engineer these genes in a crop plant. The production of high vitamin B1 crops promises to have practical applications in agriculture and medicine.
About the Author
Ming-Hsiun Hsieh, Ph.D.
Associate Research Fellow
Institute of Plant and Microbial Biology, Academia Sinica
Dr Hsieh’s research focus is on the regulation of chloroplast and mitochondrial gene expression. He and his team isolated a collection of slow growth (slo) mutants in Arabidopsis to study genes that are important for plant growth. They previously showed that the slo1 mutant is defective in a nuclear gene encoding a pentatricopeptide repeat (PPR) protein. In addition to slo mutants, studies on an Arabidopsis pale green mutant have led to the identification of a novel protein involved in organellar tRNA modification. This novel protein may play an important role in translational regulation in chloroplast and/or mitochondria. Dr Hsieh’s research also involves the amino acid sensing and signaling in plants. Their team has identified a novel type of ACT domain-containing protein family whose members contain ACT domain repeats (ACR), and hypothesizes that these novel ACT domain-containing proteins may serve as amino acid sensors in plants. They are using reverse genetic approaches to study the functions of these ACR proteins in Arabidopsis and the forward genetic approaches to isolate Arabidopsis mutants that could not grow well on medium containing amino acid as the sole nitrogen source. These mutants may be defective in amino acid uptake, metabolism, transport, and possibly signalling.