Multi-omics Analysis Of Nicotiana Bethamiana In Response To Dark Stress

Nicotiana benthamiana was an important model plant,as well a heliophile,the growth and quality of N.benthamiana was closely related to light intensity.In this study,multiple omics analysis(including proteome,phosphoroteome,transcriptome and metabolome)was employed to explore the changes of proteins,genes and metabolites during the process of plant response to dark stress.The main conclusions are as follows:1.Quantitative proteomics and phosphoproteomics research by liquid chromatography-mass spectrometryA total of 5,763 proteins and 1405 phosphorylated proteins were identified from dark-stressed N.benthamiana leaves.Weighted gene co-expression analysis(WGCNA)was applied to integrate and analysis the protein data and four co-expression modules were obtained.We found out some candidate proteins which respond to dark stress and autophagy in the module proteins,such as ribosomal protein L24/L5A/S5A,heat shock protein 70(HSP 70),isopropylmalate isomerase large/small subunit 1(IIL1/AT2G43090),eukaryotic elongation factor 5A(ELF5A),sucrose non-fermentation related protein kinase 1(SnRK1),type 2A phosphatase associated protein of 46 kDa(Tap46)and glutamate dehydrogenase(GDH).The results indicated that autophagy played a positive role in the defence against dark stress.The results of phosphoproteins showed that N.benthamiana adopted to environmental changes by differentially expressing autophagy-related protein 13(ATG13)and thylakoid lumen 18.3 kDa protein(TLP18.3).2.Transcriptomics study based on RNA-seq technologyA total of 12871 genes with a significantly different expression were identified from the N.benthamiana leaves during dark-treatment.It was found that tobacco plants can eliminate excess reactive oxygen species(ROS)through autophagy pathway and enzymatic antioxidant mechanisms under dark stress.WGCNA was performed on the differentially expressed genes and six specific modules were identified.Some hub genes were filtered out form the six modules,such as histone H2A.4/H4,mitochondrial carrier family protein(MCF),SWIB/MDM2 domain superfamily protein,his-asn-his(HNH)endonuclease,high chlorophyll fluorescence 101(HCF 101),late elongated hypocotyl/circadian clock associated-l-like protein 1(LHY1/CCA1 like 1),general control non-repressible 5(GCN5),phosphate-responsive 1 family protein(EXO),3-Ketoacyl-CoA synthase 11(KCS11)and plant invertase/pectin methylesterase inhibitor superfamily proteins(PMEI).It was speculated that these genes were involved in the process of photosynthesis,circadian rhythm regulation,growth and development of plants in response to dark stress.In addition,the differentially expressed genes ATP synthase and ribulose-1,5-bisphosphate carboxylase/oxygenase(Rubisco)also played an important role in the dark stress resistance of N.benthamiana,supposedly.3.Metabonomics research based on chromatography mass spectrometry technology.183 differential metabolites were identified in metabolomics analysis.Pathway enrichment showed that the metabolic pathways of amino acids and carbohydrates changed significantly.Plant hormones and flavonoids can not only play an important role as plant growth regulators in response to dark stress,but also participate in the process of autophagy regulation.