Genome-Wide Differential DNA Methylation and miRNA Expression Profiling Reveals Epigenetic Regulatory Mechanisms Underlying Nitrogen-Limitation-Triggered Adaptation and Use Efficiency Enhancement in Allotetraploid Rapeseed.

Ying-Peng Hua,Zhen-Hua Zhang,Chun-Yun Guan,Ting Zhou,Jin-Yong Huang,Cai-Peng Yue,Hai-Xing Song

doi:10.3390/ijms21228453

Ying-Peng Hua, Zhen-Hua Zhang + Show 5 more

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https://doi.org/10.3390/ijms21228453

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Abstract

Improving crop nitrogen (N) limitation adaptation (NLA) is a core approach to enhance N use efficiency (NUE) and reduce N fertilizer application. Rapeseed has a high demand for N nutrients for optimal plant growth and seed production, but it exhibits low NUE. Epigenetic modification, such as DNA methylation and modification from small RNAs, is key to plant adaptive responses to various stresses. However, epigenetic regulatory mechanisms underlying NLA and NUE remain elusive in allotetraploid B. napus. In this study, we identified overaccumulated carbohydrate, and improved primary and lateral roots in rapeseed plants under N limitation, which resulted in decreased plant nitrate concentrations, enhanced root-to-shoot N translocation, and increased NUE. Transcriptomics and RT-qPCR assays revealed that N limitation induced the expression of NRT1.1, NRT1.5, NRT1.7, NRT2.1/NAR2.1, and Gln1;1, and repressed the transcriptional levels of CLCa, NRT1.8, and NIA1. High-resolution whole genome bisulfite sequencing characterized 5094 differentially methylated genes involving ubiquitin-mediated proteolysis, N recycling, and phytohormone metabolism under N limitation. Hypermethylation/hypomethylation in promoter regions or gene bodies of some key N-metabolism genes might be involved in their transcriptional regulation by N limitation. Genome-wide miRNA sequencing identified 224 N limitation-responsive differentially expressed miRNAs regulating leaf development, amino acid metabolism, and plant hormone signal transduction. Furthermore, degradome sequencing and RT-qPCR assays revealed the miR827-NLA pathway regulating limited N-induced leaf senescence as well as the miR171-SCL6 and miR160-ARF17 pathways regulating root growth under N deficiency. Our study provides a comprehensive insight into the epigenetic regulatory mechanisms underlying rapeseed NLA, and it will be helpful for genetic engineering of NUE in crop species through epigenetic modification of some N metabolism-associated genes.

Highlights

Nitrogen (N) is a key macronutrient for plant growth, development, and yield capacity [1]
In terms of N transporter genes that are responsible for N absorption, we focused on the dual-affinity nitrate transporter (NRT) gene, NRT1.1/NPF6.3 [23], and the two-component high-affinity nitrate transport system genes, NRT2.1 and NAR2.1/NRT3.1 [24]
We investigated the expression of NRTs that are involved in long-distance translocation of N nutrients, such as NRT1.5/NPF7.3 and NRT1.8/NPF7.2, which are responsible for root nitrate xylem loading and unloading [25,26], respectively

Summary

Introduction

Nitrogen (N) is a key macronutrient for plant growth, development, and yield capacity [1]. Low nitrate availability is often observed in most agricultural soils; a huge amount of energy is consumed to annually apply large numbers of chemical N fertilizers to ensure crop production [3]. Increasing plant adaptability to N limitation is key to reducing N fertilizer application in modern agriculture production, which contributes to development of environmentally friendly agriculture and a sustainable ecosystem. Epigenetic DNA modification, not changing genomic sequences, plays a pivotal role in maintaining genome stability, orchestrating gene expression across plant development, and regulating plant responses to environmental stimulations [5]. Nutrient stresses in plants, such as N limitation [8], phosphate starvation [9,10], zinc deficiency [11], and sulfur deficiency [12], have been shown to change global DNA methylation at the whole genome level

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