Functional Analysis Of WNK Family Gene AtWNK9in Arabidopsis

Arabidopsis AtWNK9gene belongs to WNK gene family. In this research, wefirst analyzed AtWNK9in protein structure, phosphorylation site, promotercis-element, gene expression pattern by using bioinformatic methods. The resultsshowed AtWNK9belong to the serine-threonine protein kinase family, there aremultiple phosphorylation sites, located in the nucleus, the highest expression organwas in the root of Arabidopsis, the promoter region contained multiple hormone andabiotic stress response elements.Constructed the over-expression vector of AtWNK9and transformed intoArabidopsis to obtain the AtWNK9over-expression strains;Bought and identifiedT-DNA insertion mutants to obtain wnk9deletion mutant.Protoplast transformedexperiment proved that AtWNK9located in the nucleus.Real-time quantitative PCRand GUS stain results confirmed AtWNK9gene had a high expression level in the rootof Arabidopsis.Treatment14-day-old wild-type Arabidopsis with ABA (100μM), salt (300mM),glucose (2%), heat (37℃), cold (4℃),quantitative analysis theexpression level ofAtWNK9, found AtWNK9gene expression was induced by ABA、NaCl、glucose、hotand cold, and ABA had the most obvious induce effect.Treated with exogenous ABA,the over-expression lines of AtWNK9(AtWNK9ox3, AtWNK9ox4, AtWNK9ox7) andT-DNA insertion mutant wnk9showed a sharp difference in taproot elongate、stomatalopening、 water loss and proline accumulation. AtWNK9over-expression linesappeared hypersensitive to ABA relative to the wild type Col-4, while AtWNK9T-DNA insertion mutant wnk9suggested insensitive to ABA relative to the wild typeCol-0.Drought treatment to the over-expression mutants of AtWNK9(AtWNK9ox3,AtWNK9ox4,AtWNK9ox7) and the deletion mutant wnk9for10to12days, found theAtWNK9over-expression mutants showed a better drought resistance than wild type,while the function deletion mutant was weaker in drought-resistant ability than wildtype.Quantitative PCR analysis and found the expression level of RAB18, RD22, ABI3and ABF3were higher expressed in over-expression mutant AtWNK9ox3than the wildtype, but lower expressed in deletion function mutant wnk9; ABI1, ERA1were lowerexpressed in over-expression mutant AtWNK9ox3than the wild type, but higherexpressed in deletion function mutant wnk9. In conclusion, the Arabidopsis AtWNK9gene influenced ABA related genesexpression and acted as a positive regulator involved in ABA signal transduction,enhanced the drought resistance of Arabidopsis.