Study On Function, Expression Property And Regulation Sequence Of Rice Ca～(2+)/H～+ Antiporter OsCAX3

Ca²⁺ /H⁺ antiporters play important roles in plant nutrition and signal transduction. To keep the Ca²⁺ balance, plant must transport Ca²⁺ through calcium efflux system. Being one kind of calcium efflux transporters, Ca²⁺/H⁺ antiporters are proposed to provide three critical housekeeping functions. First, they restore [Ca²⁺]_cyt to resting levels, thereby terminating Ca²⁺ signals. Second, they load Ca²⁺ into compartments such as the ER and vacuole to be used as sources for regulated Ca²⁺ release. Third, they supply Ca²⁺ to various organelles to support specific biochemical functions. Rice is one of the model material in molecular biology, further more, one of the most important food crops in the world. To study the Ca²⁺/H⁺ antiporter of rice has theoretical and practical sense at the same time.In this paper, we focued our researches on the Ca²⁺/H⁺ antiporter gene OsCAX3 from rice. We identifed OsCAX3 from rice. Sequence analysis revealed that OsCAX3 had 11 transmembrane domains (TMs) and high similarity with other Ca²⁺/H⁺ antiporters. RT-PCR analysis showed that OsCAX3 mRNA was strongly induced by exogenous calcium, and slightly induced by exogenous K⁺, Na⁺, Mg²⁺and 4℃, and slightly down induced by Mn²⁺and PEG. It was predicted to be localized in plasma membrane by PSORT Prediction and this result was confirmed by transiently expressing OsCAX3-GFP fusion protein in mesophyll cell protoplast of arabidopsis and by stably expressed OsCAX3-GFP in tobacco. When expressed in yeast mutant, it suppressed the deficiency at high Ca²⁺ concentrations of the mutant, but could not suppress Mn²⁺, Cu²⁺, Ni²⁺, Na⁺, Zn²⁺ deficiency. We expressed OsCAX3 under 35S promoter in tobacco to study its function in plant. The transgenic tobaccohad no difference in pH 5.7 1/2 MS medium and in pH 5.7 1/2 MS supplemented with 100mM CaCl₂, and there also no difference in content between two type of plant. However, when grown in pH 7.2 1/2MS medium supplemented with 100mM CaCl₂, the transgenic tobacco growed better than wild type tobacco, furthermore, the calcium content in the transgenic tobacco is lower than in wild type tobacco.We also studied the regulation mechanism of OsCAX3, and the result showed that the 1-26 N-terminal amino acid could partially suppress its ability to transport Ca²⁺.