ARS | Publication request: Characterization of CAX9 from Arabidopsis, a homolog of mammalian K(+) dependant Na(+), Ca(2+)/H(+) (NCKX) antiporters

Submitted to: American Society of Plant Biologists Annual Meeting
Publication Type: Proceedings
Publication Acceptance Date: June 1, 2005
Publication Date: July 16, 2005
Citation: Morris, J., Shigaki, T., Hirschi, K. 2005. Characterization of CAX9 from Arabidopsis, a homolog of mammalian K(+) dependant Na(+), Ca(2+)/H(+)(NCKX) antiporters [abstract]. The 2005 Annual Meeting of the American Society of Plant Biologists Meeting, July 16-20, 2005, Seattle, Washington. Minisymposium 18: Membrane Transport, Presentation 29001, p. 69.

Technical Abstract: The Arabidopsis genome contains 11 members of the CAX (cation exchanger) family of transporters that have the ability to transport cations across the vacuolar membrane. To date only a small portion of the genes have been characterized. Here we present novel functions of CAX9, which differ from previously characterized CAX genes. Yeast cells expressing a truncated version of CAX1(sCAX1) suppress a Ca(2+) sensitive phenotype in mutant yeast strains deficient in vacuolar Ca(2+) transport. Several of the CAXs are also regulated by an N-terminal auto-inhibitory domain. However, yeast cells expressing full-length CAX9 (CAX9) and sCAX9 are unable to suppress the Ca(2+) sensitive phenotype, but CAX9 can suppress Na(+) sensitive phenotypes in several strains defective in Na(+) transport. CAX9 is different than other CAX genes in that removal of the N-terminal domain does not deregulate the protein. These biological functions are different than other characterized CAX transporters. The CAX9 ORF contains a single exon, which encodes a 643 amino acid protein (70.1 kDa) with 12 predicted trans-membrane domains with 2 highly conserved Ca(2+)/Na(+) exchanger domains in CAX9, which more so resembles characteristics found in the mammalin NCKX family of transporters. Ectopic expression of full-length "CAX9" in tobacco dramatically alters leaf morphology and plant development. These plants have drastic regions of leaf chlorosis as well as a significant decrease in Na(+) and Mn(2+) concentrations. Histochemical GUS analysis shows low levels of "CAX9" expression in developing seeds and pollen in 30-day-old Arabidopsis flowers. Localization of a CAX9 fused to eGFP protein in yeast showed the fusion protein localized to the vacuole. Furthermore this c-terminal tag did not alter the function of CAX9, since the fusion was also able to suppress the Na(+) sensitive phenotype of mutant yeast strains. Alternatively, T-DNA knock out lines of CAX9 in Arabidopsis have no discernable phenotype and have proper floral development. Our preliminary data suggests full-length CAX9 can transport Na(+) and possibly Mn(2+), and truncation of an N-terminal region can abolish this function. Furthermore ectopic expression in tobacco causes drastic phenotypic changes, suggesting a unique role of CAX9 in ion homeostasis.