The ITRQA Analysis Reveals Mechanisms Of Heavy Metal Enrichment In Eichhornia Crassipes Due To Excess Cd And Zinc

Eichhornia crassipes can enrich many kinds of heavy metals to conduct phytoremediation for the pollutional water.However,it still remains further discussion about various functions played by relevant proteins in the process as well as involved mechanism.Here we studied the change of physiological and biochemical indexes such as plant biomass,enrichment level of heavy metals when Eichhornia crassipes under stress treatment of Cd and Zn.Meanwhile,qPCR and SDS-PAGE were used to determine the expression of Eichhornia crassipes metallothionein EiMT2.The expression and puritication conditions of EiMT2 in Escherichia coli were preliminarily explored.By means of isobaric tags for relative and absolute quantitation(iTRAQ)technology,various parts of proteins in the plant were studied to make clear the obvious difference protein and analyze their functions.The molecular mechanism of heavy metal enrichment in Eichhornia crassipes were further explored,providing more clues to further improve the remediation efficiency of heavy metals.The main research results are as follows:1.Under the Cd and Zn stress,the total biomass of Eichhornia crassipes has been obviously reduced.Under the complex treatment of Cd and Zn,the addition of Zn stress makes the roots fix more Cd;while the addition of Cd reduces the enrichment ability to Zn,but improves the transport capacity of Zn in the plant.Cd and Zn stress is conducive for the Eichhornia crassipes to absorb the nutrient element of Fe.2.The EiMT2 in the roots,stems and leaves of Eichhornia crassipes has a strong response to Cd and Zn stress.Zn induces EiMT2 expression twice relative to Cd induction.Under Cd and Zn complex treatment,EiMT2 relative expression is over ten times higher than that of Cd and Zn single stress.It is stable and reproducible for EiMT2 expression in the Coli BL21(DE3)pLys S and favorable for the further exploration of its structure and function.3.The i TRAQ marking method has detected 149 differential proteins.Under the Cd stress,we identify 40 up-regulated proteins and 30 down-regulated proteins,including the metabolic-related proteins which play a role in the metablism of nitrogen,and carbohydrate,photosynthesis,photorespiration,the synthesis of proteins and so on.Under the Zn stress,we identify 28 up-regulated proteins and 51 down-regulated proteins,which take part in carbohydrate metablism,photosynthesis,photorespiration,the composition and decomposition of intracellular compound and so on.4.The analysis of proteomics shows that in the process of both Cd and Zn stress,the Rubisco in Eichhornia crassipes is remarkable up-regulated,while the otherphotosynthesis-related proteins are more for down-regulated expression,it reveals that Rubisco catalyze the photorespiration of plant instead of photosynthesis.In addition,Rubisco may be related to the metabolic of nitrogen-containing compounds in the plant.By the means of QPCR,the result of relative expression of Rubisco is contraty iTRAQ,it turns out that the stress of Cd or Zn may lead to a certain delay or differences between the gene regulation and protein expression.