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A cation/[H.sup.+] exchangers (CAXs) from yeast are analyzed along with zebrafish CAX (CAX1). The studies have provided insights into Vnx1 function and have supported an unexpected role of CAX transporters in animal growth and development.

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 Ca2+/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. [ABSTRACT FROM AUTHOR]

CAtiOn/H+ exchangers (CAxs) are integral membrane proteins that transport Ca2+ or other cations by exchange with protons. While several yeast and plant CAX proteins have been characterized, no functional analysis of a vertebrate CAX homologue has yet been reported. In this study, we further characterize a CAX from yeast, VNXl and initiate characterization ofa zebrafish CAX (Cax I). Localization studies indicated that both Vnxl and CaxI proteins are found in endomembrane compartments. Biochemical characterization of endomembrane fractions from vnx I mutant cells and zebrafish Cax I-expressing yeast cells suggested that both yeast and fish CAXs have Ca2+/H+ antiport activities. Additionally, the vnxl mutation was associated with heightened pH-sensitivity. In zebrafish embryos, cax I was specifically expressed in neural crest cells. Morpholino knockdown ofcaxl caused defects in neural crest development, including alterations in pigmentation, defects in jaw development, and reduction in expression of the neural crest marker, Pax7. Collectively, our findings provide insights into Vnxl function and support an unexpected role of CAX transporters in animal growth and development. [ABSTRACT FROM AUTHOR]