Poster: Photosynthesis

Abs # 347: PEPC expression and regulation in a same-cell C₄-system

Presenter:	Bowes, George , gbowes@botany.ufl.edu
Authors	Bowes, George  (A)   Rao, Srinath K (A)   Reiskind, Julia B (A)
Affiliations:	(A): University of Florida

Hydrilla leaves exposed to low [CO₂] change from C₃ to C₄ photosynthesis. The C₄ and Calvin cycles function in the same cell to concentrate CO₂ in the chloroplasts. This is a model for producing a "minimalist" C₄ system within a C₃ plant, such as rice, which lacks Kranz anatomy. We have reported sequence and expression data for two leaf PEPC isoforms, Hvpepc3 and 4. Northern analyses indicated that diel regulation of Hvpepc4 expression was induced during the C₄ shift. Western analyses with a maize anti-PEPC showed two bands in both C₃ and C₄ extracts, but the lower molecular mass band, potentially corresponding to Hvpepc4, dominated in the C₄. Antibodies against a phosphorylated Ser/Thr Akt substrate gave positive signals only in the lower band of light and dark C₄ extracts, suggesting that Hvpepc4 was phosphorylated in the light and dark, possibly more in the former. Hvpepc3 appeared largely non-phosphorylated. PEPC kinetics, using C₃ and C₄ leaf extracts assayed at cytosolic pH values, showed differences. The C₃ activity was lower with little malate inhibition, unlike the C₄ leaf activity. The effector glucose-6-P increased activities and reduced malate inhibition, but in C₄ extracts it also reduced the K_0.5PEP values. C₄ and C₃ extracts exhibited sigmoidal and Michaelis-Menten [PEP] responses, respectively. Light-harvested C₄ extracts had higher PEPC activities at cytosolic pH than dark-harvested, and in vitro dephosphorylation of the light C₄ PEPC increased malate sensitivity. It is clear from these studies that, to optimize a transgenic C₄ system, careful consideration should be made of the expression, kinetic and regulatory characteristics of the isoform used to introduce into a C₃ background. Supported by USDA NRICGP 98-35306-6449 and 2002-35318-12540.