Integrated multi-omics analysis reveals candidate genes for cuticular wax biosynthesis and molecular characteristics of a glossy mutant in rapeseed under natural drought stress
Results suggest that the drought response in the glossy mutant involves not only a defective cuticular barrier but also extensive metabolic and signaling reprogramming, which offers a promising approach for developing more sustainable farming methods amid environmental challenges.
Abstract
Introduction Cuticular wax plays a crucial role in drought tolerance. However, the regulatory mechanisms controlling cuticular wax biosynthesis and the drought response in rapeseed are not well understood. Methods and Results In this study, we identified a glossy mutant, hy7201, in rapeseed (Brassica napus L.). Compared to the wild-type ‘HY7201’, hy7201 exhibited a significant reduction in total cuticular wax content, altered composition, and decreased crystal density, along with a significant increase in cuticle permeability. Genetic analysis revealed that the glossy phenotype of the hy7201 mutant is controlled by a single dominant gene. Through bulked segregant analysis coupled with next-generation sequencing (BSA-seq) of waxy and glossy pools derived from F1 individuals of the self-crossed hy7201 population, BnaA09G0721400ZS, the homolog gene of AtCER1, was identified as a key candidate gene. This gene encodes a very-long-chain aldehyde decarbonylase, which contributes to the differences in leaf cuticular wax accumulation between the wild-type ‘HY7201’ and the glossy mutant ‘hy7201’. To validate the key genes involved in cuticular wax biosynthesis and characterize the molecular features under natural drought conditions in glossy plants, we performed an integrated analysis of the leaf transcriptome, proteome, and metabolome using KEGG enrichment, Pearson correlation, and two-way orthogonal partial least squares (O2PLS) methods with three biological replicates. The genes CER1 (BnaA09G0721400ZS) and its paralog CER1-2 (BnaA09G0698500ZS) were significantly downregulated at both the transcriptional and protein levels. Additionally, they exhibited significant negative correlations with differentially expressed metabolites (DEMs) in glossy plants under drought stress. The multi-omics approach uncovered that pathways related to cutin, suberin, and wax biosynthesis, as well as ABC transporters, glucosinolate biosynthesis, glutathione metabolism, linoleic acid metabolism, sphingolipid metabolism, and arginine and proline metabolism, significantly contribute to the glossy plant’s response to drought. Discussion These results suggest that the drought response in the glossy mutant involves not only a defective cuticular barrier but also extensive metabolic and signaling reprogramming. This insight could facilitate the identification of genes related to cuticular wax biosynthesis and drought stress response, which could be utilized in molecular breeding programs to enhance drought tolerance in rapeseed. Consequently, this offers a promising approach for developing more sustainable farming methods amid environmental challenges.
Introduction Plant cuticular wax has gained increasing attention owing to its critical role in biotic and abiotic stress tolerance. Identifying key genes regulating cuticular wax biosynthesis and understanding their mechanisms are important for rapeseed resistance breeding. However, the candidate genes and mechanisms involved in cuticular wax biosynthesis in rapeseed remain poorly understood. Methods and results In this study, a dominant glossy mutant 39J7H was identified in rapeseed through genetic analysis. Two stable and pure sister lines ‘4074’ (glaucous) and ‘4075’ (glossy) were developed from the glossy mutant 39J7H via successive self-crosses. The leaves of ‘4075’ exhibited significantly increased cuticle permeability as indicated by water loss rate and toluidine blue (TB) staining. Conversely, scanning electron microscopy (SEM) and GC-MS analysis demonstrated a dramatic reduction in wax crystals density, total wax content, and chemical components in ‘4075’ leaves compared to ‘4074’. Gene mapping analysis using F2 populations from the ‘4074’ × ‘4075’ cross identified the qCWA9.1 locus within a 2.3 Mb region on chromosome A09 of rapeseed through BSA-seq. Subsequent linkage analysis in the F2 population, along with phenotypic data from the F2:3 generation, refined the qCWA9.1 region to a 49.0-kb interval between the SSR markers 9AS337 and 9AS339. This interval contains nine genes, among which BnaA09G0695600ZS, an orthologue of the Arabidopsis gene AT1G01600 (AtCYP86A4), encoding a fatty acid omega-hydroxylase CYP86A4, was identified as a potential candidate gene for qCWA9.1 associated with the glossy phenotype. Sequence analysis uncovered SNP variations in the first exon and promoter region of BnaA09G0695600ZS in the glossy line ‘4075’. Additionally, qRT-PCR analysis indicated that a significant upregulated in BnaA09G0695600ZS expression in this line. Discussion Based on these results, we hypothesize that BnaA09G0695600ZS emerged as one of candidate genes for cuticular wax synthesis. The absence of two key cis-regulatory elements, I-box and TATC-box, in the promoter region of BnaA09G0695600ZS in the glossy line ‘4075’ may potentially alter cuticular wax metabolism in rapeseed leaves. These findings enhance our understanding of cuticular wax biosynthesis in rapeseed and provide valuable genetic resources for the development of high-stress-resistant rapeseed germplasms.
Lei Lei, Xirong Zhou, Weirong Wang et al.· Frontiers in Plant Science· 0 citations
Drought is a major abiotic stress that reduces crop productivity due to global climate change. The plant cuticular wax layer plays a crucial role in reducing non-stomatal water loss and enhancing drought tolerance. KCS6, encoding 3-ketoacyl-CoA synthase 6, is a key enzyme in the biosynthesis of very-long-chain fatty acids (VLCFAs), the main components of cuticular wax. However, its specific function in wax accumulation and drought resistance remains incompletely elucidated. This study aimed to characterize the role of KCS6 in wax biosynthesis and drought tolerance in Arabidopsis thaliana using CRISPR/Cas9-mediated gene knock-out. kcs6 mutants were generated by targeting the first exon of KCS6 to create a loss-of-function allele. Phenotypic analysis, cuticular wax composition, and drought tolerance assays were performed in the mutants compared to wild-type plants. The results showed that kcs6 mutants exhibited a significant reduction in VLCFA content and cuticular wax load. The mutants also displayed a higher rate of water loss and increased sensitivity to drought stress compared to the wild type. These findings provide direct genetic evidence that KCS6 is essential for cuticular wax biosynthesis and for maintaining plant water status under drought conditions. This study is the first to demonstrate, using CRISPR/Cas9-mediated targeted mutagenesis, that disruption of KCS6 directly links altered VLCFA-derived cuticular wax biosynthesis to impaired plant water retention and drought adaptation, providing mechanistic evidence for KCS6 as a promising genetic target for improving crop drought resilience.
H. Sultan, Muhammad Zubair, A. Ashraf· Jurnal Mangifera Edu· 0 citations
Simple Summary Cotton is one of the world’s most important fiber and oil crops, but drought and salty soils strongly reduce its growth and yield. Enzymes of the BXL family help remodel plant cell walls and are thought to contribute to a plant’s ability to cope with environmental stress. In this study, we searched the cotton genome and identified 25 BXL genes, and we examined their evolutionary relationships, gene structures, and the regulatory regions that control their activity. We then tested how these genes respond when cotton seedlings are exposed to drought-like and salt-like conditions. Several BXL genes were strongly activated under both treatments, and three in particular were identified as central “hub” genes within networks of stress-responsive genes. These results indicate that specific BXL genes may help cotton survive dry or salty conditions, and they provide candidate targets for future efforts to improve cotton’s tolerance to drought and salinity. Such improvements could help stabilize cotton production under increasingly challenging environmental conditions.
Zhenzhen Wei, An-Xing Zhu, Yang Liu et al.· Biology· 0 citations
Functional characterization demonstrated that transgenic tobacco plants overexpressing AsGRF17 displayed significantly enhanced drought tolerance compared with wild-type plants, and this study provides new insights into the molecular mechanisms underlying AsGRF17 -mediated drought tolerance in oat.
Jingyan Li, Shi Tong, Kailian Li et al.· 0 citations
A chromosome-level genome assembly for ZY821, an elite high-GSL variety, is generated using long-read sequencing and Hi-C scaffolding and uncovered several novel candidate genes implicated in GSL metabolism.
Yizhou He, Zetao Bai, Zengfeng Wang et al.· Molecular Horticulture· 0 citations
The TIFY gene family comprises plant-specific transcriptional regulators central to jasmonic acid (JA) signaling and responses to biotic and abiotic stresses. Despite the economic importance of the banana (Musa spp.), the TIFY family remains largely uncharacterized in this crop. Here, we conducted a genome-wide identification and comprehensive analysis of the MaTIFY gene family in Musa acuminata. A total of 47 MaTIFY genes were identified, distributed across all 11 chromosomes. Phylogenetic analysis classified these into four subfamilies (TIFY, ZIZ/ZML, PPD, and JAZ), and conserved motif and domain analyses revealed a core TIFY domain architecture with subfamily-specific structural features. Gene Ontology (GO) enrichment and cis-acting regulatory element analyses suggested potential involvement in JA-mediated signaling, defense response, and hormone cross-talk. Expression profiling under drought, Fusarium oxysporum f. sp. cubense race 4 (Foc 4), and cold stress revealed distinct transcriptional responses, with MaTIFY5, MaTIFY16, MaTIFY20, MaTIFY26, and MaTIFY30 exhibiting enhanced induction in resistant cultivars compared to their susceptible counterparts. Functional characterization of MaTIFY20 confirmed its significant upregulation under drought stress and its ability to confer enhanced osmotic tolerance when heterologously expressed in yeast. These findings provide novel insights into the evolutionary dynamics and stress-responsive functions of banana TIFY genes and identify candidate targets for molecular breeding to improve abiotic and biotic stress resilience in banana.
Sheraz Ahmad, Huimin Song, Hangbo Cao et al.· International Journal of Mol...· 0 citations