Identification Of Bdf1p-dependent Salt Tolerance And Screening The Suppressors Of The Bdf1△ Salt Sensitive Phenotype In Saccharomyces Cerevisiae

Microorganisms continuously experience rapid and drastic changes of the surrounding environment. Consequently, cell need to sense and to respond to such alterations in a highly controlled fashion in order to survive and adapt to the ever-changing environment. Currently, the molecular mechanism research of cell response to high salt stress is not only convenient and effective functional genomics research model to the interpretation of gene function, Meanwhile, it has important significance to the transformation of the industrial microbial and improvement of the resistance of plant crops against saline conditions.Using the pools of the yeast genomic library with mini-transposon to transform the haplaid Saccharomyces cerevisiae strain W303-1A, we got many mutants sensitive to high salt stress totally. By the rescue plasmid, the insertion sites of the transposon in mutants were sought out, and those insertions affected 61 genes. The mutant named 258-B10 was studied more. The insertion site of mTn3 transposon in 258-B10 was in the intergenic region 1002bp of the BDF1 gene. 258-B10 showed hypersensitivity under high salt stress condition. The S. cerevisiae BDF1 gene, was first isolated in a large-scale screen for salt-sensitivity mutants following transposon mutugenesis.This paper based on the original work, and further confirm the relationship between high salt stress sensitivity and BDF1 gene, We constructed BDF1 deletion strain and BDF1 gene expression vector pYX- PB1. Our data indicated that bdf1Δstrain show the same high-salt (0.65mol/ L) stress-sensitive phenotype with transposition insertion mutants 258-B10 labels. And the expression vector pYX- PB1 transformed into bdf1Δstrains can fill the stress-sensitive phenotype. These results show BDF1 gene play a certain role in the respond to the high-salt stress condition.We focus on two aspects, on the one hand, high salt stress on the yeast cells includes the ion toxic effects and hyperosmotic stress. Therefore, we confirmed the Bdf1p respond to ion toxicity. In addition, ENA1 gene is important Na~+ efflux pump in S. cerevisiae. It is controlled by several important high-salt stress signal pathway, such as high osmolarity glycerol mitogen activated protein kinase signaling transduction pathway and Calcineurin pathway. We studied Genetic interaction between BDF1 and ENA1 gene and proved Bdf1p is not responsed to high salt stress through ENA1 gene.On the other hand, we used the pHR81 yeast chromosome library to transform 258-B10, we got two plasmid that can recover the hypersensitive phenotype of 258-B10. By sequencing, the yeast chromosome fragment was obtained. It respectively contains the entire BDF1 and the BDF2 fragment. Then we constructed high- and low-copy BDF2 gene expression vector into bdf1Δstrain. The result showed up this phenotype is not affected by changes of BDF2 expression dose while BDF2 deletions did not show salt sensitive phenotype. It indicated there is some functional overlap between Bdf1p and Bdf2p in yeast salt resistance.In short, we studied the mechanism of response to high salt stress, we confirmed Bdf1p participated in ion toxicity stress, furthermore we found its homologous gene BDF2 can complement the sensitive phenotype. These results provide good foundation for further study. More study is going on.