The Role Of Chromatin Remodeling Complex ATPase CHR16 In Response To Drought Stress In Arabidopsis Thaliana

Chromatin remodeling affects the binding of various transcription factors to chromatin through the precise control of chromatin modification by a series of complexes,thus making cells feel the stimulation from various signals and environments.SNF2 chromatin remodeling complexes play an important role in regulating plant growth,development and response to stresses.As a member of the SNF2 family,the biological function of CHR16 is rarely reported.In the previous work,our group found that CHR16 gene of Arabidopsis thaliana plays an important role in SAM and RAM formation and stem cell identity maintenance by regulating the expression of CLV3,WUS,STM,PLT2,SCR,CYCB,QC46 and QC184 genes.To explore other biological function of CHR16,Col-0,chr16-1,chr16-2,chr16-3,and chr16-4 mutants were used as experimental materials.Through the phenotypic analysis,histochemical staining,yeast two hybrid,qPCR,physiological,and transcriptome,the biological functions of CHR16 was further investigated,the main results are as follows:1.Through phenotypic analysis,identification of pure heterozygosity by three-primer method and RT-PCR,we found that chr16-1,chr16-2,chr16-3 and chr16-4 mutants were single-gene recessive mutations.Furthermore,the homozygous mutants were completely deletion mutations.Compared with the wild type(WT),the homozygous mutant showed the phenotype of short root and smaller cotyledon,which could complete the life story,but the time of mossy was later,and could blossom but be sterile.2.By construction of pCHR16-GUS transgenic lines,we studied the tissue localization of CHR16 in Arabidopsis thaliana.GUS staining results showed that there were strong GUS signals in seed coat,cotyledon,true leaf,hypocotyl,long sill,rosette leaves,flower organs and anther of Arabidopsis thaliana,indicating that CHR16 was widely expressed in plants.3.The yeast library of Arabidopsis thaliana was screened by yeast two-hybrid method,14 candidate substrates were preliminarily obtained.Through comparison,it was found that RD2,DI19 and PP2C62 were all involved in drought stress response.Therefore,we speculated that CHR16 of Arabidopsis might be related to drought stress response.4.We treated pCHR16-GUS transgenic plants with ABA,the staining results showed that GUS activity was inhibited.Subsequent qPCR results also showed that ABA treatment could reduce the transcription level of CHR16 in Arabidopsis.The results above indicate that CHR16 plays a negative regulatory role in ABA response.5.ABA can not only promote dormancy of seeds,but also cause growth stagnation when the embryo after seed germination is subjected to water stress.ABA response after germination in Arabidopsis thaliana is mediated to a large extent by the ABA-induced transcription factor ABA INSENSITIVE5(ABI5).We found that the absence of CHR16 function of ATPase CHR16 in SWI2/SNF2 chromatin remodeling caused hypersensitivity of Arabidopsis to ABA after germination.In the absence of ABA,both ABI5 and ABI3 genes were upregaged in chr16-2 mutant,and the upregaged multiple of ABI5 was lower in chr16-2 mutant than WT under ABA stress,but the upregaged multiple of ABI3 in both was not as significant as that in ABI5.Therefore,the loss of CHR16 function will lead to the selective ABA response gene expression changes.Most obviously,the inhibition of ABI5 and ABI3 expression will be removed in the absence of ABA.6.Analysis of the drought sensitivity shown that the tolerance of chr16-2 mutant response to drought stress increased compared with WT,and have a reduced water loss rate in isolated plants,suggesting that CHR16 may play a negative regulatory role in Arabidopsis response to drought stress.7.ABA induction of stomatal closure showed that stomatal closure was faster in chr16-2 mutant under ABA treatment,which may be the direct cause of increased drought tolerance.8.Transcriptome analysis results showed that under drought stress,a total of 1816 differentially expressed genes were associated with CHR16,and most of them were concentrated in catabolic pathway and plant hormone signal transduction pathway.Most of the members of ABA signaling pathway were differentially expressed,suggesting that CHR16 gene may participate in the response to drought stress by regulating the transcription level of ABA signaling pathway members.In summary,our results suggest that CHR16 can participate in the response to drought stress by regulating stomatal closure,as a regulatory factor of drought stress,interact with members of ABA pathway and become a downstream target of drought stress/ABA signaling pathway.In addition,CHR16 may play a role in the balance between plant growth and stress response.These results are helpful for us to further understand the response of epigenetic chromatin remodeling in plant growth,development and drought stress,and provide more ideas for the genetic breeding of crops.