Elsevier

Plant Science

Volume 292, March 2020, 110380
Plant Science

Transcriptome and GWAS analyses reveal candidate gene for seminal root length of maize seedlings under drought stress

https://doi.org/10.1016/j.plantsci.2019.110380Get rights and content

Highlights

Abstract

Water deficits are a major constraint on maize growth and yield, and deep roots are one of the major mechanisms of drought tolerance. In this study, four root and shoot traits were evaluated within an association panel consisting of 209 diverse maize accessions under well-watered (WW) and water-stressed (WS) conditions. A significant positive correlation was observed between seminal root length (SRL) under WS treatment and the drought tolerance index (DI) of maize seedlings. The transcriptome profiles of maize seminal roots were compared between four drought-tolerant lines and four drought-sensitive lines under both water conditions to identify genes associated with the drought stress response. After drought stress, 343 and 177 common differentially expressed genes (DEGs) were identified in the drought-tolerant group and drought-sensitive group, respectively. In parallel, a coexpression network underlying SRL was constructed on the basis of transcriptome data, and 10 hub genes involved in two significant associated modules were identified. Additionally, a genome-wide association study (GWAS) of the SRL revealed 62 loci for the two water treatments. By integrating the results of the GWAS, the common DEGs and the coexpression network analysis, 7 promising candidate genes were prioritized for further research. Together, our results provide a foundation for the enhanced understanding of seminal root changes in response to drought stress in maize.

Introduction

Drought affects crop growth and greatly reduces crop yields globally and is considered as one of the most severe environmental stressors [1,2]. Maize (Zea mays L.) is the most widely grown staple food crop, but maize production is frequently compromised as a result of the increasing frequency and intensity of drought [3]. Thus, improvement of drought tolerance has become a priority during modern maize breeding.

Plant roots play a crucial role in the absorption of water and nutrients from the soil and are also an important component of the drought stress response [[4], [5], [6]]. Optimal root architecture is beneficial for improving plant drought tolerance and maintaining plant productivity under drought stress [5,7,8]. The ‘steep, cheap and deep’ (SCD) ideotype root architecture, involving reduced lateral branching density and reduced number of axial roots, has been proposed to enhance the rapid exploitation of deep soil resources in maize [9]. Reduced lateral branching density and reduced number of axial roots can improve drought tolerance in maize by reducing intra-plant root competition, permitting greater axial root elongation, thereby improving deep water capture [8,10]. During the development of drought, soil moisture is lost from the surface, and deep roots have been demonstrated to serve as a strategy to access water stored deep in the soil profile, for example, maize and wheat under moderate drought [6,11,12]. The angle at which roots penetrate the soil may also relate to root depth. Under moderate drought or severe drought treatments, steeper root growth angles increase rooting depth and drought tolerance in rice [13]. In maize, seminal root angles are strongly positively correlated with grain yield under severe drought [14].

For drought adaptation, seminal roots can improve access to residual moisture within deep soil layers [15]. For instance, previous studies in wheat and barley have reported that relatively deep seminal roots are beneficial for the absorption of water under drought stress [16,17]. For the first 2–3 weeks of maize seedling growth, seminal roots are crucial for seedling survival, prompt establishment and early development after germination [9,18,19]. Seminal roots are produced in maize [20], wheat [15] and barley [16], whereas rice [21] and sorghum [22] lack this type of root. Lynch et al. [9] hypothesized that maize seminal roots have a relatively larger diameter to improve deep soil exploration and contribute to foraging in deeper soil horizons. In addition, previous studies have shown that seminal root length (SRL) is positively correlated with shoot biomass under low phosphorus and water stress conditions [20,23]. To increase maize drought tolerance and yield under drought stress, it is important to explore the genes underlying SRL.

Several genes controlling the development of seminal roots have been cloned in maize. For example, both rtcs, which encodes a LOB-domain protein [24], and rum1, which encodes an Aux/ indoleacetic acid (IAA) protein [25] affect the initiation of seminal roots. In addition, mutation of the gene bige1, which encodes a multidrug and toxin extrusion (MATE) transporter, results in increased number of seminal roots [26]. By the evaluation of both biparental and natural populations, quantitative trait locus (QTL) mapping and genome-wide association studies (GWAS) have been used to elucidate the genetic basis underlying SRL in maize. Many QTLs regulating maize SRL have been reported under different conditions, including various phosphorus deficiencies [20], nitrogen deficiencies [27], water stress [23], and normal conditions [28,29]. With the advent of increasing types of economically viable sequencing technologies, GWAS has become a popular method for identifying natural genetic variation associated with target traits. Although previous GWAS have examined the wide natural variation associated with SRL under normal conditions in maize [[30], [31], [32]], the genetic basis of SRL under water stress, especially under soil drought stress conditions, remains to be elucidated by GWAS.

RNA sequencing (RNA-seq) has become the favored technique for detecting genome-wide gene expression patterns. Tai et al. [33] compared transcriptomic profiling differences between primary, seminal and crown roots of maize, and found diverse patterns of gene activity across all root types and highlighted the unique transcriptome of the seminal roots. Moreover, many important differentially expressed genes (DEGs) in primary root tissue have been identified by RNA-seq in maize under water stress [34,35]. To identify the complex mechanism governing transcriptional regulations, Hwang et al. [36] utilized weighted gene coexpression network analysis (WGCNA) to identify functional connections between genes and confirmed the presence of reliable hub genes involved in a potential subnetwork related to initial root development at different developmental stages in maize seedlings. These studies provided general insight into root development and abiotic stress responses, but the identification of potential key candidate genes underlying target traits is difficult because of the large amounts of DEGs obtained by RNA-seq. Recently, examination of candidate genes identified from RNA-seq via significant SNPs from GWAS or via candidate gene association analysis has been employed to interpret RNA-seq results. For example, Sekhon et al. [37] showed that an integrated genomic and transcriptomic analysis could be used to identify novel genes and networks involved in the genetic control of senescence in maize.

In this study, four drought-tolerant lines and four drought-sensitive lines selected from 209 diverse inbred lines were used for RNA-seq analysis of SRL under WW and WS conditions. The expression profiles of genes were analyzed to identify DEGs. Using WGCNA, we detected important modules of coexpressed genes and hub genes involved in SRL under water stress. Using the 209 inbred lines with SRL phenotypes under WW and WS conditions, we also conducted a GWAS to determine genomic loci associated with SRL under drought stress. Lastly, we integrated the transcriptomic analysis and GWAS results to identify candidate genes that respond to drought stress. The goal of this study was to identify genomic loci and candidate genes affecting SRL under drought conditions for improving drought tolerance breeding in maize.

Section snippets

Plant materials and growth conditions

A natural population including 209 diverse accessions of maize inbred lines was used for measuring the shoot dry weight (SDW), primary root length (PRL), seminal root length (SRL), and seminal root number (SRN) under well-watered (WW) and water-stressed (WS) conditions. Pot experiments were arranged in a greenhouse located on the campus of Chinese Academy of Agricultural Sciences under constant conditions (14 h of day at 28 °C/10 h of night at 24 °C), during 2017-2018. The pots consisting of

Evaluation of root traits and drought tolerance in diverse maize germplasm

To systematically evaluate the relationship between root traits and drought tolerance of maize grown in soil, a natural population comprising 209 diverse accessions was grown under WW and WS conditions, and four traits, SDW, SRL, SRN, and PRL, were phenotyped at the seedling stage (35 days after emergence). The lines within each treatment were significantly (P < 0.001) different for all traits (Table S2). The repeatability of the root-related traits was moderate, ranging from 0.62 (PRL) to 0.71

Precise root phenotyping of plants growing in soil

Drought stress is a major limiting factor for crop production worldwide, and plant root system is of great significance in drought tolerance [5]. Understanding the relationship between root architecture and drought tolerance could help in the optimization of plant root system to improve drought tolerance. Phenotypic characterization and quantification of the root architecture of the plants growing in the soil are major challenges. PVC tubes and plastic pots filled with soil are often used to

Conclusions

In the present study, we accurately phenotyed the root architecture and drought tolerance index of a diverse maize association panel at seedling stage, and found that the seminal root length was more conducive to improving drought tolerance than the primary root length. Comparative transcriptome analyses of seminal roots provided a comprehensive overview of the transcriptome variations between drought-tolerant lines and drought-sensitive lines, and revealed the gene expression patterns of

Author contributions

CL, YuL, DY and TW conceived and designed the experiment and revised the manuscript; JG, XZ, YuS, and YaS performed the field experiment and phenotype evaluation; JG, DZ and YoL collected and analyzed the data; JG wrote the first draft of the manuscript; CL, YuL, DY and TW modified the manuscript. All authors have read and approved the publication of the manuscript.

Declaration of Competing Interest

The authors declare that they have no competing interests.

Acknowledgements

This research was supported by the Programs of MOST and MOA of China (2016YFD0101201), the National Natural Science Foundation of China (31971891), Young Elite Scientists Sponsorship Program by CAST (2016QNRC001), and the CAAS Innovation Program.

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