A phased small interfering RNA-derived pathway mediates lead stress tolerance in maize.

Phased small interfering RNAs (phasiRNAs) are a distinct class of endogenous small interfering RNAs, which regulate plant growth, development and environmental stress response. To determine the effect of phasiRNAs on maize (Zea mays L.) tolerance to lead (Pb) stress, the roots of 305 maize lines under Pb treatment were subjected to generation of individual databases of small RNAs. We identified 55 high-confidence phasiRNAs derived from 13 PHAS genes (genes producing phasiRNAs) in this maize panel, of which 41 derived from nine PHAS loci were negatively correlated with Pb content in the roots. The potential targets of the 41 phasiRNAs were enriched in ion transport and import. Only the expression of PHAS_1 (ZmTAS3j, Trans-Acting Short Interference RNA3) was regulated by its cis-expression quantitative trait locus and thus affected the Pb content in the roots. Using the Nicotiana benthamiana (N. benthamiana) transient expression system, 5'-rapid amplification of cDNA ends, and Arabidopsis heterologously expressed, we verified that ZmTAS3j was cleaved by zma-miR390 and thus generated tasiRNA targeting ARF genes (tasiARFs), and that the 5' and 3' zma-miR390 target sites of ZmTAS3j were both necessary for efficient biosynthesis and functional integrity of tasiARFs. We validated the involvement of the zma-miR390-ZmTAS3j-tasiARF-ZmARF3-ZmHMA3 pathway in Pb accumulation in maize seedlings using genetic, molecular, and cytological methods. Moreover, the increased Pb tolerance in ZmTAS3j-overexpressed lines was likely attributed to the zma-miR390-ZmTAS3j-tasiARF-ZmARF3-SAURs pathway, which elevated indole acetic acid levels and thus reactive oxygen species scavenging capacity in maize roots. Our study reveals the importance of the TAS3-derived tasiRNA pathway in plant adaptation to Pb stress.

Genetic variations in ZmSAUR15 contribute to the formation of immature embryo-derived embryonic calluses in maize.

The ability of immature maize (Zea mays) embryos to form embryonic calluses (ECs) is highly genotype dependent, which limits transgenic breeding development in maize. Here, we report the association map-based cloning of ZmSAUR15 using an association panel (AP) consisting of 309 inbred lines with diverse formation abilities for ECs. We demonstrated that ZmSAUR15, which encodes a small auxin-upregulated RNA, acts as a negative effector in maize EC induction. Polymorphisms in the ZmSAUR15 promoter that influence the expression of ZmSAUR15 transcripts modulate the EC induction capacity in maize. ZmSAUR15 is involved in indole-3-acetic acid biosynthesis and cell division in immature embryo-derived callus. The ability of immature embryos to induce EC formation can be improved by the knockout of ZmSAUR15, which consequently increases the callus regeneration efficiency. Our study provides new insights into overcoming the genotypic limitations associated with EC formation and improving genetic transformation in maize.

A genome-wide co-expression network analysis revealed ZmNRAMP6-mediated regulatory pathway involved in maize tolerance to lead stress.

KEY MESSAGE: A metal transporter ZmNRAMP6 was identified by using a trait-associated co-expression network analysis at a genome-wide level. ZmNRAMP6 confers maize sensitivity to Pb by accumulating it to maize shoots. ZmNRAMP6 knockout detains Pb in roots, activates antioxidant enzymes, and improves Pb tolerance. Lead (Pb) is one of the most toxic heavy metal pollutants, which can penetrate plant cells via root absorption and thus cause irreversible damages to the human body through the food chain. To identify the key gene responsible for Pb tolerance in maize, we performed a trait-associated co-expression network analysis at a genome-wide level, using two maize lines with contrasting Pb tolerances. Finally, ZmNRAMP6 that encodes a metal transporter was identified as the key gene among the Pb tolerance-associated co-expression module. Heterologous expression of ZmNRAMP6 in yeast verified its role in Pb transport. Combined Arabidopsis overexpression and maize mutant analysis suggested that ZmNRAMP6 conferred plant sensitivity to Pb stress by mediating Pb distribution across the roots and shoots. Knockout of ZmNRAMP6 caused Pb retention in the roots and activation of the antioxidant enzyme system, resulting in an increased Pb tolerance in maize. ZmNRAMP6 was likely to transport Pb from the roots to shoots and environment. An integration of yeast one-hybrid and dual-luciferase reporter assay uncovered that ZmNRAMP6 was negatively regulated by a known Pb tolerance-related transcript factor ZmbZIP54. Collectively, knockout of ZmNRAMP6 will aid in the bioremediation of contaminated soil and food safety guarantee of forage and grain corn.

Genetic dissection of maize seedling traits in an IBM Syn10 DH population under the combined stress of lead and cadmium.

The heavy metals lead and cadmium have become important pollutants in the environment, which exert negative effects on plant morphology, growth and photosynthesis. It is particularly significant to uncover the genetic loci and the causal genes for lead and cadmium tolerance in plants. This study used an IBM Syn10 DH population to identify the quantitative trait loci (QTL) controlling maize seedling tolerance to lead and cadmium by linkage mapping. The broad-sense heritability of these seedling traits ranged from 65.8-97.3% and 32.0-98.8% under control (CK) and treatment (T) conditions, respectively. A total of 53 and 64 QTL were detected under CK and T conditions, respectively. Moreover, 42 QTL were identified using lead and cadmium tolerance coefficient (LCTC). Among these QTL, five and two major QTL that explained > 10% of phenotypic variation were identified under T condition and using LCTC, respectively. Furthermore, eight QTL were simultaneously identified by T and LCTC, explaining 5.23% to 9.21% of the phenotypic variations. Within these major and common QTL responsible for the combined heavy metal tolerance, four candidate genes (Zm00001d048759, Zm00001d004689, Zm00001d004843, Zm00001d033527) were previously reported to correlate with heavy metal transport and tolerance. These findings will contribute to functional gene identification and molecular marker-assisted breeding for improving heavy metal tolerance in maize.

ZmbZIP54 and ZmFDX5 cooperatively regulate maize seedling tolerance to lead by mediating ZmPRP1 transcription.

Extensive lead (Pb) accumulation in plants exerts toxic effects on plant growth and development and enters the human food chain. Combining linkage mapping, transcriptome analysis, and association studies, we cloned the ZmbZIP54 transcription factor, which confers maize tolerance to Pb. Combined overexpression and knockdown confirmed that ZmbZIP54 mitigates Pb toxicity in maize by alleviating Pb absorption into the roots. Yeast one-hybrid and dual-luciferase assays revealed that ZmbZIP54 binds to the ZmPRP1 promoter and promotes its transcription. Yeast two-hybrid and bimolecular fluorescence complementation assays indicated that ZmFdx5 interacts with ZmbZIP54 in the nucleus. ZmFdx5 acts as a switch that controls the regulation of ZmPRP1 expression by ZmbZIP54 when maize encounters Pb stress. Furthermore, we revealed that variation in the 5'-UTR of ZmbZIP54 affects its expression level under Pb stress and contributes to the difference in Pb tolerance among maize lines. Finally, we proposed a model to summarize the role of ZmbZIP54 in Pb tolerance, which involves the cooperative effect of ZmbZIP54 and ZmFdx5 on the ZmPRP1 transcription in maize response to Pb. This study provides novel insights into the development of Pb-tolerant maize varieties and bioremediation of Pb-contaminated soils.

Association mapping uncovers maize ZmbZIP107 regulating root system architecture and lead absorption under lead stress.

Lead (Pb) is a highly toxic contaminant to living organisms and the environment. Excessive Pb in soils affects crop yield and quality, thus threatening human health via the food chain. Herein, we investigated Pb tolerance among a maize association panel using root bushiness (BSH) under Pb treatment as an indicator. Through a genome-wide association study of relative BSH, we identified four single nucleotide polymorphisms (SNPs) and 30 candidate genes associated with Pb tolerance in maize seedlings. Transcriptome analysis showed that four of the 30 genes were differentially responsive to Pb treatment between two maize lines with contrasting Pb tolerance. Among these, the ZmbZIP107 transcription factor was confirmed as the key gene controlling maize tolerance to Pb by using gene-based association studies. Two 5' UTR_variants in ZmbZIP107 affected its expression level and Pb tolerance among different maize lines. ZmbZIP107 protein was specifically targeted to the nucleus and ZmbZIP107 mRNA showed the highest expression in maize seedling roots among different tissues. Heterologous expression of ZmbZIP107 enhanced rice tolerance to Pb stress and decreased Pb absorption in the roots. Our study provided the basis for revelation of the molecular mechanism underlying Pb tolerance and contributed to cultivation of Pb-tolerant varieties in maize.

Transcription Factors Responding to Pb Stress in Maize.

Pb can damage the physiological function of human organs by entering the human body via food-chain enrichment. Revealing the mechanisms of maize tolerance to Pb is critical for preventing this. In this study, a Pb-tolerant maize inbred line, 178, was used to analyse transcription factors (TFs) expressed under Pb stress based on RNA sequencing data. A total of 464 genes expressed in control check (CK) or Pb treatment samples were annotated as TFs. Among them, 262 differentially expressed transcription factors (DETs) were identified that responded to Pb treatment. Furthermore, the DETs were classified into 4 classes according to their expression patterns, and 17, 12 and 2 DETs were significantly annotated to plant hormone signal transduction, basal transcription factors and base excision repair, respectively. Seventeen DETs were found to participate in the plant hormone signal transduction pathway, where basic leucine zippers (bZIPs) were the most significantly enriched TFs, with 12 members involved. We further obtained 5 Arabidopsis transfer DNA (T-DNA) mutants for 6 of the maize bZIPs, among which the mutants atbzip20 and atbzip47, representing ZmbZIP54 and ZmbZIP107, showed obviously inhibited growth of roots and above-ground parts, compared with wild type. Five highly Pb-tolerant and 5 highly Pb-sensitive in maize lines were subjected to DNA polymorphism and expression level analysis of ZmbZIP54 and ZmbZIP107. The results suggested that differences in bZIPs expression partially accounted for the differences in Pb-tolerance among the maize lines. Our results contribute to the understanding of the molecular regulation mechanisms of TFs in maize under Pb stress.

Short-term rosuvastatin therapy for prevention of contrast-induced acute kidney injury in patients with diabetes and chronic kidney disease.

OBJECTIVES: This study sought to evaluate the safety and efficacy of rosuvastatin in preventing contrast-induced acute kidney injury (CI-AKI) in patients with diabetes mellitus (DM) and chronic kidney disease (CKD). BACKGROUND: CI-AKI is an important complication after contrast medium injection. While small studies have shown positive results with statin therapy, the role of statin therapy in prevention of CI-AKI remains unknown. METHODS: We randomized 2,998 patients with type 2 DM and concomitant CKD who were undergoing coronary/peripheral arterial angiography with or without percutaneous intervention to receive rosuvastatin, 10 mg/day (n = 1,498), for 5 days (2 days before, and 3 days after procedure) or standard-of-care (n = 1,500). Patients' renal function was assessed at baseline, 48 h, and 72 h after exposure to contrast medium. The primary endpoint of the study was the development of CI-AKI, which was defined as an increase in serum creatinine concentration ≥0.5 mg/dl (44.2 µmol/l) or 0.25% above baseline at 72 h after exposure to contrast medium. RESULTS: Patients randomized to the rosuvastatin group had a significantly lower incidence of CI-AKI than controls (2.3% vs. 3.9%, respectively; p = 0.01). During 30 days' follow-up, the rate of worsening heart failure was significantly lower in the patients treated with rosuvastatin than that in the control group (2.6% vs. 4.3%, respectively; p = 0.02). CONCLUSIONS: Rosuvastatin significantly reduced the risk of CI-AKI in patients with DM and CKD undergoing arterial contrast medium injection. (Rosuvastatin Prevent Contrast Induced Acute Kidney Injury in Patients With Diabetes [TRACK-D]; NCT00786136).