Grapevine and Arabidopsis Cation-Chloride Cotransporters Localize to the Golgi and Trans-Golgi Network and Indirectly Influence Long-Distance Ion Transport and Plant Salt Tolerance

Plant cation-chloride cotransporters (CCCs) have been implicated in conferring salt tolerance. They are predicted to improve shoot salt exclusion by directly catalysing the retrieval of sodium (Na+) and chloride ions (Cl-) from the root xylem. We investigated whether VvCCC from grapevine (Vitis vinifera L.) has a role in salt tolerance by cloning and functionally characterising the gene from Cabernet Sauvignon. Amino acid sequence analysis revealed that VvCCC shares a high degree of similarity with other plant CCCs. A VvCCC-YFP translational fusion protein localised to the Golgi and trans-Golgi network (TGN), not the plasma membrane, when expressed transiently in tobacco (Nicotiana benthamiana) leaves and Arabidopsis thaliana mesophyll protoplasts. AtCCC-GFP from Arabidopsis also localised to Golgi and TGN. In Xenopus laevis oocytes, VvCCC targeted to the plasma membrane where it catalysed bumetanide sensitive 36Cl-, 22Na+ and 86Rb+ uptake, suggesting that VvCCC (like AtCCC) belongs to the NKCC class of CCCs. Expression of VvCCC in an Arabidopsis ccc knockout mutant abolished the mutant9s stunted growth phenotypes, and reduced shoot Cl- and Na+ content to wild-type levels after growing plants in 50 mM NaCl. In Vitis vinifera roots, VvCCC transcript abundance was not regulated by Cl- treatment and was present at similar levels in both the root stele and cortex of three Vitis genotypes that exhibit differential shoot salt exclusion. Our findings indicate that CCC function is conserved between grapevine and Arabidopsis, but neither protein is likely to directly mediate ion transfer with the xylem or have a direct role in salt tolerance.