Fuction Analysis Of AtSCAMP Family Genes In Arabidopsis Thaliana

Salt stress is one of the most important factors that threaten the world food production. Soil salinization is a more outstanding problem in China. The study of the function of salt resistance genes can help to understanding salt-tolerant mechanism and improving salt-tolerance in plant. In recent years, with the deepening of the research, many salt-resistant genes in wheat constantly have been found. But because of the huge genome and the long growth cycles, it is more difficult to do research with wheat. Through analyzing the homologous gene in arabidopsis, it can help to knowing function of the homologous gene in wheat effectively.Previous work in our lab has cloned three genes, TaSCAMP1, TaSCAMP2 and TaSCAMP3, whose expression were increased treated by salt stress. We also verified the interaction between TaSCAMP1 and the Na+/H+ transport protein which located on the tonoplast membrance.This paper is based on the results of the previous experiments, and the purpose is to verify whether the SCAMP family protein can directly interact with Na+/H+ antiporter protein in Arabidopsis.There are five SCAMP family proteins in Arabidopsis. Because there is no exact name, they were named by their full length nucleotides respectively, that is ATSCAMP849 (AT2G20840)、ATSCAMP870 (AT1G61250)、ATSCAMP876 (AT1G11180) ATSCAMP852 (AT1G03550) and ATSCAMP795 (AT1G32050). There are 7 Na+/H+ antiporter protein in Arabidopsis, AtNHX1, AtNHX2, AtNHX3, AtNHX4, AtNHX5, AtNHX6 and AtNHX7, which mainly located on the tonoplast membrane except AtNHX7 located on the plasma membrane.4 Na+/H+ antiporter protein genes and 4 AtSCAMP genes in Arabidopsis have been cloned, and they are all constructed into the expression vectors for BiFC analysis. Then the interaction between the two protein families has been detected respectively. Our results show that AtNHX2 can directly inteact with AtSCAMP849 through BiFC analysis. Besides no interactions are observed between AtNHXl and AtSCAMP849、AtNHX1 and AtSCAMP876、AtNHX2 and AtSCAMP876. Because of the auto-fluorescence of AtNHX3、AtNHX6, we cannot identify whether the direct interaction exists between AtSCAMP795, AtSCAMP870 and the Na+/H+ antiporter protein. We will exchange the vector or use other strategies to eliminate auto-fluorescence of empty contrasts.In order to ensure the authenticity of the interaction between AtS CAMP 849 and AtNHX2, we use the traditional yeast hybrid (Y2H) to verify the interaction. The result showed that there was no interaction between AtNHX2 and AtSCAMP849. We think there are two reasons. One is that the result of BiFC is false positive. The other is that the traditional Y2H cannot be applied to the two proteins. Because AtNHX2 and AtSCAMP849 are membrane proteins which can not enter the nuclear, the report genes in the yeast cannot be activated.Another technology, that is mating-based split ubiquitin system (mbSUS), has been used to eliminate the second possibility and to verify the interaction between AtNHX2 and AtSCAMP849. The result showed that there has interaction between AtNHX2 and AtSCAMP849.