Location: Children's Nutrition Research CenterTitle: Characterization of "Arabidopsis" Ca(2+)/H(+) exchanger CAX3) Author
Submitted to: Biochemistry
Publication Type: Peer reviewed journal
Publication Acceptance Date: 6/8/2011
Publication Date: 6/9/2011
Citation: Manohar, M., Shigaki, T., Mei, H., Park, S., Marshall, J., Aguilar, J., Hirschi, K.D. 2011. Characterization of "Arabidopsis" Ca(2+)/H(+) exchanger CAX3. Biochemistry. 50(28):6189-6195. Interpretive Summary: Plant transporters can move nutrients and toxin across membranes. We seek to alter nutrient transport function in order to improve nutrient content within edible plant tissues. Here we demonstrate that a calcium transporter is highly active in plants when we "tailor" the transporter by removing some of the protein. This "extra" portion of the transporter may negatively regulate function, and we can utilize our findings to generate designer transporters to enhance a host of nutrients within edible tissues.
Technical Abstract: Plant calcium (Ca2+) gradients, millimolar levels in the vacuole and micromolar levels in the cytoplasm, are regulated in part by high-capacity vacuolar cation/H(+) exchangers (CAXs). Several CAX transporters, including CAX1, appear to contain an approximately 40-amino acid N-terminal regulatory region (NRR) that modulates transport through N-terminal autoinhibition. Deletion of the NRR from several CAXs (sCAX) enhances function in plant and yeast expression assays; however, to date, there are no functional assays for CAX3 (or sCAX3), which is 77% identical and 91% similar in sequence to CAX1. In this report, we create a series of truncations in the CAX3 NRR and demonstrate activation of CAX3 in both yeast and plants by truncating a large portion (up to 90 amino acids) of the NRR. Experiments with endomembrane-enriched vesicles isolated from yeast expressing activated CAX3 demonstrate that the gene encodes Ca(2+)/H(+) exchange with properties distinct from those of CAX1. The phenotypes produced by activated CAX3-expressing in transgenic tobacco lines are also distinct from those produced by sCAX1-expressing plants. These studies demonstrate shared and unique aspects of CAX1 and CAX3 transport and regulation.