Functional Analysis Of The XCD1 Gene Involved In The Regulation Of Zinc Stress In Arabidopsis

Heavy metal contamination is an environmental problem that is increasing worldwide as a result of mining operations, industrial production and domestic and agricultural use of metal and metal containing compound have resulted in the release of toxic metals into the environment. As one of the world’s recognized heavy metal elements, zinc will be a serious threat to human health and the environment through the food chain on the zinc contaminated soil. The use of phytoremediation for heavy metal pollution has been identified as an economical and practical approach. And the main content of this research is how to combine phytoremediation technology with genetic engineering, and then to apply for repairing the zinc polluted soil, which depending on our understanding on the mechanisms of plant heavy metal tolerance.In our previous work, we found that XCD1 (At3g10890) gene regulated cadmium tolerance through the glutathione-dependent pathway in Arabidopsis thaliana. In this study, the biological function of XCD1was analyzed in regulation the zinc tolerance of plants. The results were as follows:(1)No significant differences in growth were observed between the WT,xcd1mutants and overexpressing transgenic plants grown on plain 1/2 MS medium. However, when ZnS04was applied, the mutants were more sensitive to Zn stress than were WT, and all overexpressing seedlingsdisplayed greater Zn tolerance than WT seedlings. Quantitative analyses confirmed that FW and root length of theoverexpressing seedlings were significantly (P< 0.05) higher thanthose of WT seedlings grown on Zn-containing medium, and the FW and root length of themutantswere lower thanthose of WT seedlings. Together, these results suggest that XCD1 plays an important role in the regulation of Zn tolerance inArabidopsis.(2) We next determined the spatial expression pattern of theXCD1gene in Arabidopsis by qRT-PCR analysis. XCD1 wasexpressed in most of the examined tissues, including root, inflorescences, siliques, rosette leaves, cauline leaves and stems, with thehighest expression in the inflorescence organs.(3) We found that the Zn content was significantlyincreased in overexpressing lines but was decreased inxcdl mutant plants subjected to Zn treatment.(4) The GSH content was significantly decreased byZnSO4, but this effect was alleviated in overexpressingplants which was correlated with increased activation of theGSHl gene that plays an important role in the controlof GSH biosynthesis. Moreover, under Zn stress, PC concentrations increased significantly and a larger increase in PC content was determined in the overexpressing line, whichwas associated with greatly increased PCS1 and PCS2 expression. But the contents of GSH and PC in xcdl mutant plants were decreased compared with those of WT seedlings under Zn stress. These results collectively suggest that XCD1 gene mediates Zn tolerance via the GSH-dependent PC synthesis pathway bycoordinated activation of expression of genes involved in PC biosynthesis.(5) TheZn-triggered growth inhibition was abolished in both WT and mutant seedlings grown on 1/2 MS mediacontaining mannose. The decrease of GSH content was alleviated by mannose. PC content and the Zn content were greatly increased under mannose added. These collectively suggest that mannose plays a general rolein the response to Zn stress and functions downstream of XCD1 in the signaling pathway.(6) Mannose was also added in hydroponics and it still increased the Zn content of Arabidopsis.