Proteomic And Khib Modification Analysis Of Arabidopsis Thaliana Siliques Under Salt Stress And Function Study On ATATPase-? Subunit

In the natural environment,many abiotic and biotic stresses affect the normal growth and development of plants.Under salt stress,which is one of the abiotic stress factors,plants ensure survival through physiological,biochemical,molecular and protein regulatory mechanisms.Proteomics and posttranslational modification proteomics are research methods to explore the impact of protein function on life activities.From the aspect of protein functions,proteomics and modification studies will be more reliable than the transcriptomics.Khib is one of the relatively new post-translational modifications,which is involved in transcription and cell metabolism.In this study,the differentially expressed proteins and differentially Khib modified proteins of Arabidopsis thaliana siliques under salt stress and control were analyzed,and the At ATPase-? gene was studied.The results are as follows:1.Under salt stress,the Arabidopsis silique was found to be chili-like shape with an enlarged base.The subcellular structure of the silique base was observed,and it was found that the outer layer cells were neatly arranged,while the endocarp cells were small and numerous in control.Under salt stress,size and shape of the cells were irregularly pitted,and the arrangement of outer layer in the base of siliques was more irregular.2.A total of 6603 differentially expressed proteins were identified under salt stress,293 of which were up-regulated and 179 were down-regulated(>1.3 fold).The results of GO,protein domains and KEGG showed that the differentially expressed proteins were mainly distributed in defense response,response to water diprivation and response to ABA during the biological processes.In terms of cellular composition,the differentially expressed proteins were mainly concentrated in vacuoles and membrane protein complexes.As for molecular function,the differentially expressed proteins were mainly associated with nutrient reservoir activity and xyloglucan: xyglucosyl transferase activity.The subcellular localization analysis revealed that the differentially expressed proteins were mainly concentrated in the Bifunctional inhibitor/plant lipid transfer protein/seed storage helix region,RNA recognition motif domain.Differentially expressed proteins are mainly enriched in spliceosomes,oxidative phosphorylation and photosynthesis.3.Under salt stress,a total of 3810 normalized Khib sites on 1254 proteins were identified in siliques,with Khib up-regulated at 96 sites on 78 proteins and down-regulated at 282 sites on 205 proteins(>1.3 fold).There were 19 Motif types of differentially Khib modified proteins,of which G*Khib was the most adundant.The results of GO classification,protein domains,and KEGG annotation showed that the differentially Khib modified proteins were mainly distributed in responses to cadmium ions,reproductive system development and reproductive structure development in during the biological processes.Among the cell component,differentially Khib modified proteins were mainly distributed in vacuole and vacuole membrane.In molecular function,differently Khib modified proteins were related to transmembrane and active transmembrane transporter activities as well,copper ion binding.Subcellular localization analysis revealed that the differential Khib modified proteins were mainly enriched in the C-terminal and peptide binding domain of HSP 70 k D,apical domain and equatorial domain of Groel-like.Khibmodified proteins were enriched in glycolysis/gluconeogenesis,pyruvate metabolism.4.The correlation analysis of proteomics and Khib modification revealed that 1451 proteins had changeds in both protein expression level and Khib modification level in siliques under salt stress,and 257 proteins differed only in Khib.5.Under salt stress,the protein expression level of At ATPase-? was unchanged,and the Khib modification expression was down-regulated with,9 modified sites,among which552 sites underwent significant Khib modifications with a 2-fold differences in expression.The PRM verification showed that,the protein K amino acid hads Khib modification.At ATPase-? gene was more sensitive to salt stress in silique and root than in leaf.At ATPase-? gene editing lines were successfully obtained by constructing vectors.The results showed that the At ATPase-? gene was responsive to salt stress.