水稻Ca^(2+)/H^+反向转运体OsCAX3的功能分析和亚细胞定位研究

Functional Analysis of Rice Ca^(2+)/H^+ Antiporter OsCAX3 in Yeast and Its Subcellular Localization in Plant

Ca2+/H+反向转运体作为一类Ca2+外向转运器,在植物的营养和信号转导中起着非常重要的作用.克隆了水稻Ca2+/H+反向转运体基因OsCAX3,序列分析表明OsCAX3具有11个跨膜区,其中在第6和第7个跨膜区之间有一个17个氨基酸组成的酸性基序(acidmotif),功能互补实验证明OsCAX3具有转运Ca2+的功能,并且其N端26个氨基酸序列对转运Ca2+具有一定的抑制作用.RT-PCR分析表明OsCAX3的表达受到外源Ca2+的诱导.利用PSORTprediction进行亚细胞定位分析,和利用OsCAX3-GFP融合蛋白瞬时表达分析证明,OsCAX3定位于细胞质膜.以上结果表明,OsCAX3是一种定位于细胞质膜上的Ca2+/H+反向转运体.

Ca^2＋/H^＋ antiporters play important roles in plant nutrition and signal transduction. A novel Ca^2＋/H^＋ antiporters gene OsCAX3 was identified from rice. Sequence analysis revealed that OsCAX3 has 11 transmembrane domains, and an acid motif consisting of 17 amino acids between sixth and seventh transmembrane domains. When expressed in yeast, OsCAX3 complemented the mutant growing deficiency at high Ca^2＋ concentrations, and its N-terminal（1～26 amino acid） partially suppressed its ability to transport Ca^2＋. RT-PCR analysis showed that OsCAX3 mRNA was induced by exogenous calcium. OsCAX3 was predicted to localize in plant plasma membrane by PSORT prediction, and this was further confirmed by transiently expressing OsCAX3-GFP fusion protein in mesophyll cell protoplast of Arabidopsis. All these findings strongly suggested that OsCAX3 is a Ca^2＋/H^＋ antiporter located in plasma membrane.