Phenotypic and Proteomic Insights into Differential Cadmium Accumulation in Maize Kernels.

The contamination of agricultural soil with cadmium (Cd), a heavy metal, poses a significant environmental challenge, affecting crop growth, development, and human health. Previous studies have established the pivotal role of the ZmHMA3 gene, a P-type ATPase heavy metal transporter, in determining variable Cd accumulation in maize grains among 513 inbred lines. To decipher the molecular mechanism underlying mutation-induced phenotypic differences mediated by ZmHMA3, we conducted a quantitative tandem mass tag (TMT)-based proteomic analysis of immature maize kernels. This analysis aimed to identify differentially expressed proteins (DEPs) in wild-type B73 and ZmHMA3 null mutant under Cd stress. The findings demonstrated that ZmHMA3 accumulated higher levels of Cd compared to B73 when exposed to varying Cd concentrations in the soil. In comparison to soil with a low Cd concentration, B73 and ZmHMA3 exhibited 75 and 142 DEPs, respectively, with 24 common DEPs shared between them. ZmHMA3 showed a higher induction of upregulated genes related to Cd stress than B73. Amino sugar and nucleotide sugar metabolism was specifically enriched in B73, while phenylpropanoid biosynthesis, nitrogen metabolism, and glyoxylate and dicarboxylate metabolism appeared to play a more significant role in ZmHMA3. This study provides proteomics insights into unraveling the molecular mechanism underlying the differences in Cd accumulation in maize kernels.

Dynamic changes of rhizosphere soil bacterial community and nutrients in cadmium polluted soils with soybean-corn intercropping.

BACKGROUND: Soybean-corn intercropping is widely practised by farmers in Southwest China. Although rhizosphere microorganisms are important in nutrient cycling processes, the differences in rhizosphere microbial communities between intercropped soybean and corn and their monoculture are poorly known. Additionally, the effects of cadmium (Cd) pollution on these differences have not been examined. Therefore, a field experiment was conducted in Cd-polluted soil to determine the effects of monocultures and soybean-corn intercropping systems on Cd concentrations in plants, on rhizosphere bacterial communities, soil nutrients and Cd availability. Plants and soils were examined five times in the growing season, and Illumina sequencing of 16S rRNA genes was used to analyze the rhizosphere bacterial communities. RESULTS: Intercropping did not alter Cd concentrations in corn and soybean, but changed soil available Cd (ACd) concentrations and caused different effects in the rhizosphere soils of the two crop species. However, there was little difference in bacterial community diversity for the same crop species under the two planting modes. Proteobacteria, Chloroflexi, Acidobacteria, Actinobacteria and Firmicutes were the dominant phyla in the soybean and corn rhizospheres. In ecological networks of bacterial communities, intercropping soybean (IS) had more module hubs and connectors, whereas intercropped corn (IC) had fewer module hubs and connectors than those of corresponding monoculture crops. Soil organic matter (SOM) was the key factor affecting soybean rhizosphere bacterial communities, whereas available nutrients (N, P, K) were the key factors affecting those in corn rhizosphere. During the cropping season, the concentration of soil available phosphorus (AP) in the intercropped soybean-corn was significantly higher than that in corresponding monocultures. In addition, the soil available potassium (AK) concentration was higher in intercropped soybean than that in monocropped soybean. CONCLUSIONS: Intercropped soybean-corn lead to an increase in the AP concentration during the growing season, and although crop absorption of Cd was not affected in the Cd-contaminated soil, soil ACd concentration was affected. Intercropped soybean-corn also affected the soil physicochemical properties and rhizosphere bacterial community structure. Thus, intercropped soybean-corn was a key factor in determining changes in microbial community composition and networks. These results provide a basic ecological framework for soil microbial function in Cd-contaminated soil.

Natural variations in the P-type ATPase heavy metal transporter gene ZmHMA3 control cadmium accumulation in maize grains.

Cadmium (Cd) accumulation in maize grains is detrimental to human health. Developing maize varieties with low Cd content is important for safe consumption of maize grains. However, the key genes controlling maize grain Cd accumulation have not been cloned. Here, we identified one major locus for maize grain Cd accumulation (qCd1) using a genome-wide association study (GWAS) and bulked segregant RNA-seq analysis with a biparental segregating population of Jing724 (low-Cd line) and Mo17 (high-Cd line). The candidate gene ZmHMA3 was identified by fine mapping and encodes a tonoplast-localized heavy metal P-type ATPase transporter. An ethyl methane sulfonate mutant analysis and an allelism test confirmed that ZmHMA3 influences maize grain Cd accumulation. A transposon in intron 1 of ZmHMA3 is responsible for the abnormal amino acid sequence in Mo17. Based on the natural sequence variations in the ZmHMA3 gene of diverse maize lines, four PCR-based molecular markers were developed, and these were successfully used to distinguish five haplotypes with different grain Cd contents in the GWAS panel and to predict grain Cd contents of widely used maize inbred lines and hybrids. These molecular markers can be used to breed elite maize varieties with low grain Cd contents.

[Cashmere goat bacterial artificial chromosome recombination and cell transfection system].

The Cashmere goat is mainly used to produce cashmere, which is very popular for its delicate fiber, luscious softness and natural excellent warm property. Keratin associated protein (KAP) and bone morphogenetic protein (BMP) of the Cashmere goat play an important role in the proliferation and development of cashmere fiber follicle cells. Bacterial artificial chromosome containing kap6.3, kap8.1 and bmp4 genes were used to increase the production and quality of Cashmere. First, we constructed bacterial artificial chromosomes by homology recombination. Then Tol2 transposon was inserted into bacterial artificial chromosomes that were then transfected into Cashmere goat fibroblasts by Amaxa Nucleofector technology according to the manufacture's instructions. We successfully constructed the BAC-Tol2 vectors containing target genes. Each vector contained egfp report gene with UBC promoter, Neomycin resistant gene for cell screening and two loxp elements for resistance removing after transfected into cells. The bacterial artificial chromosome-Tol2 vectors showed a high efficiency of transfection that can reach 1% to 6% with a highest efficiency of 10%. We also obtained Cashmere goat fibroblasts integrated exogenous genes (kap6.3, kap8.1 and bmp4) preparing for the clone of Cashmere goat in the future. Our research demonstrates that the insertion of Tol2 transposons into bacterial artificial chromosomes improves the transfection efficiency and accuracy of bacterial artificial chromosome error-free recombination.

Higher methylation in genomic DNA indicates incomplete reprogramming in induced pluripotent stem cells.

Pluripotent stem cells can be created successfully through the inner cell mass (ICM), nuclear transfer, and defined-factor induction. Unfortunately, the epigenetic characteristics of the cells produced are poorly understood. In this article, we compared expression levels of enzymes involved in epigenetic modifications across six pluripotent stem cell lines. Six of the 11 genes evaluated here (Dnmt3a, Dnmt3b, Tet1, Ezh2, Mll1, and Lsd1) showed abnormally low levels of expression in the two germ-line chimeric induced pluripotent stem cell (iPSC) lines. We also conducted locus-specific analysis of DNA methylation at 9 loci. Although iPSCs did express Oct4, the Oct4 promoter region was shown to have a higher level of DNA methylation. The Xist and Line-1 repeating sequences differed relatively little in methylation level across the cell lines, but Peg3, Peg10, and H19 exhibited high degrees of variation in the pattern of DNA methylation. Meg3 in the Dlk1-Dio3 imprinting cluster was incompletely methylated in embryonic stem cells (ESCs) and nuclear transfer (nt) ESCs. However, in germ-line chimeric iPSCs, Meg3 was almost entirely methylated. ESC and ntESC lines showed twice as much Meg3 expression than in the iPSC lines. The genomic 5mC contents detected by reverse-phase high-performance liquid chromatography (HPLC) indicated that, despite their germ-line chimeric abilities, iPSCs remained incompletely reprogrammed, even though no direct evidence is shown here.