Mechanisms Which By The XCD1Gene Regulates Cadmium Resistance In Arabidopsis

Heavy metal cadmium (Cd) pollution in soil is one of the global environmentproblems. Phytoremediation of genetic engineering is an important way to solve theCd pollution in soil. The key of this work is to understand their underlyingmolecular mechanisms of the tolerance to Cd toxicity and excessive accumulationof Cd and to clone the key genes involved in the abiotic stress responses in plants.In preliminary studies, we screened a T-DNA insert mutant xcd1-D, and cloneda Cd-resistant gene XCD1in Arabidopsis, and gain-of-function of this gene led toCd tolerance in Arabidopsis. To further determine the role of XCD1gene, itsfunction and action mechanisms were been studied by using loss-of-functionmethods. The results are as follows:1. Under normal growth conditions, there was no significant difference in thegrowth between the wild type and XCD1knockout mutants xcd1-1xcd1-2, xcd1-3;however, xcd1-1, xcd1-2and xcd1-3mutants were more sensitive to Cd stress thanthe wild type, as indicated by increased inhibition of their root length and freshweight.2. Analysis of the subcellular localization of XCD1protein showed that thesubcellular localization of XCD1-GFP fusion protein were mainly located in thecell wall. Moreover, analysis of ProXCD1:GUS expression showed that theexpression of XCD1gene was in roots of plant.3. The biochemical function of the protein encoded by XCD1was analyzedusing prokaryotic expression technology, and it was shown that the protein has theβ-mannanase activity.4. Analysis of Cd content showed that inducible expression of XCD1gene led toincreased accumulation of Cd in the mutant xcd1-D, which was increased1.39-foldcompared to wild-type. Under Cd stress,35S::XCD1transgenic plants had increasedcontent of Cd, which was increased an average of1.72-fold to the wild type. Moreover, Cdcontent of the knockout mutants was slightly lower than that of the wild type. Under theCd and β-estradiol treatment, the Cd content was measured in the cell wallextracted from wild-type and xcd1-D mutant. The results showed the Cdaccumulation of xcd1-D mutant was significantly increased compared with the wild type and improved1.25times.5. To further understand the mechanisms of XCD1-mediated Cd accumulationin the plant roots, the xcd1-D mutant and wild-type were treated with Cd andβ-estradiol. It was found that the β-mannanase activity of cell wall in the xcd1-Dmutant was significantly increased, and that the content of hemicellulose in xcd1-Dmutant was decreased, but the Cd content in the hemicellulose was increased,suggesting that their substances might bind to Cd molecule.6. Under normal growth conditions, there was no significant difference in thewild-type and XCD1knockout mutant plants subjected to exogenous mannosetreatment. However, under Cd stress, exogenous mannose treatment could improveCd resistance of wild type and xcd1-2mutant, but more increased Cd resistance wasobserved in xcd1-2mutant. Moreover, Cd contents of the roots and the cell wallwere also increased in wild type and xcd1-2mutant, indicating that it is possiblethat mannose-Cd conjugates were adsorbed on the cell wall of the root.