Poster: Redox regulation

Abs # 238: AtPOS5-2 is a calmodulin-independent NAD⁺ kinase found in Arabidopsis stroma

Presenter:	Guenther, James F, jguenth@utk.edu
Authors	Guenther, James F (A)   Ashley, David W (A)   Choi, Won-Gyu  (A)   Kabachinski, Gregory L (A)   Roberts, Daniel M (A)
Affiliations:	(A): The University of Tennessee

Regulation of the cell pools of nicotinamide adenine nucleotides is mediated by the enzyme NAD⁺ kinase (NADK) which converts NAD⁺ to NADP⁺ using ATP as a phosphodonor. Conversion of NAD⁺ to NADP⁺ is associated with physiological responses such as light regulated shifts in redox co-enzymes during photosynthesis, and also possibly in plant defense and regulation of reactive oxygen levels. Biochemical analysis shows two activities of NADK in plants: 1) A Ca²⁺ and calmodulin-dependent activity, and 2) a calmodulin-independent activity. Examination of the Arabidopsis genome reveals three genes that encode putative NADKs with similarity to the yeast POS5p protein, an NAD⁺(H) kinase involved in oxidative stress-adaptation in yeast. All three AtPOS5 proteins possess a conserved carboxyl-terminal NADK domain fused to an extended amino-terminal region which is not conserved in yeast and bacterial NADKs. However, orthologs with high similarity are found in Oryza sativa (OsPOS5). Functional analysis of AtPOS5 and OsPOS5 proteins produced by invitro transcription/translation or by expression of the recombinant proteins in E. coli shows that these enzymes are NADKs with calmodulin-independent activity. Antibodies against AtPOS5-2 detect a protein with an identical molecular weight in the chloroplast stroma of both spinach and Arabidopsis, coinciding with a calmodulin-independent NADK activity found in both fractions. The data suggest that AtPOS5-2 is a calmodulin-independent chloroplastic NADK which may be responsible for the light-regulated increase in NADP⁺ associated with photosynthesis. (Supported by National Science Foundation grant MCB-0237219 to DMR)