Poster: Metabolic engineering

Abs # 297: Genetic modification of ADP-glucose pyrophosphorylase in Arabidopsis: Effects on enzyme activity, starch biosynthesis, and physiology.

Presenter:	Nagai, Yasuko S, ynagai@wsu.edu
Authors	Nagai, Yasuko S (A)   Hwang, Seon-Kap  (A)   Edwards, Gerald E (B)   Okita, Thomas W (A)   Kato, Chikako  (C)   Ito, Hiroyuki  (C)   Matsui, Hirokazu  (C)
Affiliations:	(A): Institute of Biological Chemistry, Washington State University (B): School of Biological Sciences, Washington State University (C): Graduate School of Agriculture, Hokkaido University

We are studying the significance of the capacity for starch biosynthesis in leaves, through manipulations of ADP-glucose pyrophosphorylase (AGPase), to source-sink relationships and growth using Arabidopsis as a model system. AGPase, a key enzyme in plant starch synthesis, consists of a pair of regulatory large subunits and catalytic small subunits. We previously demonstrated that the starch deficient mutant of Arabidopsis TL46 which contains only a functional small subunit gene, has lower starch synthesis, lower rates of CO₂ assimilation and growth than wildtype (Arabidopsis thaliana (L.) Heynh cv. Columbia) (Sun et al. 1999 Plant Physiol. 119:267, Sun et al. 2002 Plant Physiol. 130:1573). The large subunit gene, ApL1, was isolated using a combination of PCR amplification and cloning techniques. This cloned gene was engineered by site-directed mutagenesis to select for mutant forms having modified responses to allosteric effectors and catalysis. Arabidopsis TL46 plants lacking a functional largel subunit of AGPase were transformed using the wildtype and mutant forms of the large subunit gene, ApL1. These transformants, along with TL46 and wildtype, were grown under controlled environments under normal atmospheric conditions versus CO₂ enrichment. Transformed Arabidopsis lines having wildtype and modified large subunits were identified which have higher AGPase activity and higher starch levels than wildtype, while TL46 has low AGPase activity and low starch. Preliminary results indicate that elevation of starch metabolism in TL46 results in more normal growth and increases in photosynthetic capacity. The specific effects of the modification of the ApL1 gene on AGPase activity, starch accumulation and turnover, photosynthesis, growth and plant biomass will be presented.