Research
Analysis of Carbohydrate Metabolism Mutants of Arabidopsis
Carbohydrate synthesis and subsequent utilization are among the most fundamental processes in plants. In spite of the importance of these processes, little is known about the regulation of starch synthesis and degradation, especially in vegetative tissues. We isolated Arabidopsis mutants with altered starch content in leaves and applied the reverse genetics to study genes involved in starch metabolism. Mutants are characterized by genetic, biochemical, and molecular methods to understand the function of mutated genes. These approaches resulted in the discovery of previously unknown proteins (e.g., SEX1, DPE2 and MEX1) essential for leaf starch degradation.
The pathway of starch metabolism derived from the analysis of Arabidopsis mutants is substantially different from the pathway in germinating cereal seeds. £\- Amylase was thought to be the enzyme initiating the degradation of starch granules. However, we found that no difference in transitory starch metabolism in the knockout mutants of chloroplast and cytosolic £\-amylase genes, compared to the wild type, indicating that £\- amylase is not essential for starch breakdown in leaves. Analysis of Arabidopsis starch metabolism mutants: mex1, dpe1 and dpe2, showed that starch and maltose or malto-oligosaccharides are accumulated in leaves of these mutants. These results suggest that maltose is a major product derived from starch degradation and £]-amylase would be one of the primary enzymes for transitory starch degradation. Indeed, our analysis of £]-amylase mutants showed that these mutants have impaired starch degradation.
Currently, we are analyzing starch-excess mutants, sex4 and ke103. We found that sex4 and ke103 mutants are deficient in plastidial £\- and £]-amylase, respectively. However, neither sex4 nor ke103 encodes the deficient enzyme. Thus, analysis of these mutants would provide insight for the regulation in leaf starch metabolism.
We also found temporal and spatial coordinated expression of several starch metabolism genes in plants. Transcription regulation of starch metabolism genes is the primary mechanism of this coordinated expression.It would be interesting to isolate mutants with defective transcription factors and to identify the cis regulatory elements for starch metabolism genes.
