Mechanism Research On Cadmium-stress Response Of Brassica Based On Transcriptomic

Heavy metal pollution seriously affects the yield and quality of crops, and poses a threat to human health due to the migration of heavy metals in the food chain. According to the investigation report of Ministry of Environmental Protection and Ministry of Land and Resources of the People’s Republic of China, Cadmium contamination is the most serious and largest heavy metal pollution of China. Plant is one of the main ways for humans to absorb heavy metals. However, it is also the critical carrier used for the phytoremediation of heavy metal contamination. There are a lot of Brassica germplasm in China, and some species of Brassica napus and Brassica juncea have a certain tolerance and accumulation to Cd, which means that Brassica has the potential to remediate the Cd contamination and, at the same time, maintain its original economic value. Thus, revealing the Cd-stress response mechanisms of B. napus and B. juncea and identifying the key transporters or genes involved in Cd uptake and metabolism in Brassica are of great significance for the cultivation of plant which can be used for the phytoremediation of contaminated soils or have high resistance and low accumulation to Cd when planted in the Cd contaminated soils.In this study, some physiological and biochemical parameters of B. napus and B. juncea, including height, number of effective branch, content of Cd and mineral elements, grain quality and relative expression levels of transporter genes involved in Cd transport, under various Cd treatments have been fully assayed. Meanwhile, to identify the candidate genes and pathways involved in Cd-stress response in leaves of B. napus thereby elucidating the molecular mechanism of Cd-stress tolerance and accumulation of it and providing some theoretical basis for the construction of applied Cd hyperaccumulators, the transcriptomes of B. napus leaves under two different Cd-stress conditions were compared using RNA-Sequencing and bioinformatics. The main results are as following:(1)The soil culture experiment results showed that Cd stress scarcely affected the growth of Brassica at the bolting stage. Cd disturbed the net CO2 assimilation of Brassica leaves to a certain extent, but no significant phenomenon was observed in plant height, number of effective branch and biomass under Cd stress conditions. The Cd content in Brassica was increased along with the rise of stress concentration of Cd and changed along with Brassica species. Compared with B. napus, B. juncea had a stronger alibilty to translocate Cd from roots to shoots. Meanwhile, the orders of the total Cd accumulation in 4 Brassica species were as follows: Bn#2>Bj#2>Bn#1>Bj#1. The Cd absorbed by plant was mainly accumulated in aboveground parts and proportional to the concentration of Cd treatment and biomass, meaning that the biomass of tissue is the main factor of total Cd accumulation.(2)The experiment results showed that the Cd content of the mature rapeseeds harvested after 70-days Cd treatment was more than the limit value of Cd in crops and the quality of rapeseed was also decreased. The rapeseed Cd concentrations in all 4 Brassica species were more than 2.02 μg/g and proportional to the Cd concentration in the soil. The fatty acid compositions of rapeseeds detected by Gas Chromatography were affected by Cd, obviously. The results showed that the content of the fatty acid beneficial to human, such as Oleic acid and linoleic acid, was decreased but the harmful erucic acid content was increased.(3)Cd stress disturbed the absorption, transport and redistribution of mineral nutrient elements in Brassica. The accumulation of metal metals, including calcium, magnesium, iron, manganese, zinc and copper, in plant tissues was obviously changed under Cd treatment, and the influences, promotion or inhibition, were different among Brassica species or tissues. Significantly, the content of Fe and Mn in plants was obviously enhanced under 50 ppm Cd treatment.(4)Cd influenced the expression of the transporter genes involved in Cd uptake and transport in Brassica. The qRT-PCR results of Lower Leaf, Upper Leaf and Silique in Bn#1 and Bj#1 showed that the relative expression levels of many transporter genes, including ABCC family, CET family, HMA family and Nramp family, were related to Cd. Especially, the expression levels of ABCC1/2 and HMA2/3 were sinificantly affected by Cd.(5)The total RNA of upper leaves in Bn#1 under 0 ppm and 200 ppm Cd treatment was used for RNA-Sequencing and bioinformatics. The RNA-Seq results showed that the numbers of differentially expressed genes(DEGs), mainly enriched GO term and KEGG pathway were 2012, 112 and 10, respectively. Of the 2102 DEGs, 680 and 1422 genes were detected to be up-regulated and down-regulated, respectively. GO and KEGG enrichment analysis of these DEGs indicated that oxidation-reduction process and many biosynthesis of secondary metabolites including sulfur metabolism were up-regulated under Cd stress. Meanwhile, 82 transporter genes, including 16 metal ion transporter genes, were found according to the results of functional annotation of DEGs. It is noteworthy that AtHMA2 and OsHMA2, homologous proteins of BnHMA2, have been shown to have the ablitiy to absorb and transport Cd in Arabidopsis thaliana and Oryza sativa, respectively. In addition, the results of qRT-PCR analysis of DEGs were consistent with those of RNA-Seq data, which confirmed the reliability of RNA-Seq results.