基于uORF编辑的作物精准育种研究进展与展望

doi:10.3864/j.issn.0578-1752.2026.21.001

摘要/Abstract

摘要：

上游开放阅读框（upstream open reading frame，uORF）是位于真核生物mRNA 5′非翻译区（5′UTR）的关键顺式作用元件，在基因翻译和性状形成调控中发挥重要作用。uORF主要通过漏扫描、翻译重起始、核糖体停滞或触发无义介导的衰变等机制拦截扫描中的核糖体，进而抑制下游主阅读框（mORF）的翻译。这种抑制作用的强度高度依赖于uORF起始密码子两侧序列及RNA二级结构等特定的基因组结构特征。该机制赋予了植物在不改变转录的前提下，敏锐响应外部环境信号的能力，在维持正常生长发育和逆境适应的动态平衡中发挥着独特作用。进化分析表明，具有适应性优势的uORF在正向自然选择驱动下往往被保留并呈现高度保守性，同时，植物群体中自然发生的变异uORF序列能够显著微调目标基因的翻译效率，是驱动作物群体抗逆响应和农艺表型多样性分化的重要遗传基础。随着高通量组学技术的发展，大量潜在功能性uORF的翻译图谱被精准解析，为后续的靶向编辑奠定了坚实的数据基础。相较于启动子或3′UTR等其他区域的编辑，靶向uORF具有多效性低、预期性强且无需引入外源序列的显著优势。可通过CRISPR/Cas9、引导编辑（prime editing）等基因编辑技术实现uORF长度的改变、Kozak序列修饰，以及偶联环境响应型uORF等多种工程化策略。这些微调基因表达的策略能够突破传统强启动子过表达引发的负面多效性瓶颈，实现目标基因表达量的适当上调。通过编辑uORF精准调控内源目标蛋白丰度在改良作物广谱抗病性、耐逆性，优化生育期等发育性状，以及提升维生素含量等营养品质方面存在巨大的潜力。本文以建立基于uORF编辑的高效作物精准育种技术体系为目标，阐述了uORF的主要特征特性及其分子调控机制，梳理了uORF编辑的策略及其在性状改良应用中取得的进展，探讨了uORF编辑在作物精准育种中的应用前景和面临的挑战，提出了结合深度学习模型提升uORF编辑效果及其可预见性的策略和途径。

关键词: 上游开放阅读框, 翻译调控, 基因编辑, 作物育种, 5′非翻译区

Abstract:

Upstream open reading frames (uORFs) are key cis-acting elements in the 5′ untranslated region (5′UTR) of eukaryotic mRNAs, which play important roles in regulation of gene translation and ultimately phenotypes. uORFs mainly block scanning ribosomes through mechanisms like leaky scanning, translation reinitiation, ribosome stalling, or nonsense-mediated mRNA decay (NMD), which consequently inhibits the translation of the downstream main open reading frame (mORF). The strength of such inhibition depends heavily on specific uORF structural features, such as the sequence around its start codon and RNA secondary structures. The uORF-based translation regulation allows plants to quickly respond to environmental signals without changing transcription levels, playing a unique role in balancing normal growth with stress adaptation. Evolutionary analysis shows that beneficial uORFs are often kept by natural selection and highly conserved. Moreover, natural uORF sequence variations in plant populations can finely adjust the translation efficiency of target genes. This provides an important genetic basis for the diversity of stress responses and agronomic traits in crops. With the development of high-throughput omics technologies, the translation profiles of many functional uORFs have been accurately mapped, providing a solid data foundation for targeted editing. Compared to editing other regions like promoters or 3′UTRs, targeting uORFs has major advantages: lower pleiotropy, higher predictability, and no need to introduce foreign sequences. Various engineering strategies—such as changing uORF length, modifying Kozak sequences, and using environment-responsive uORFs—can be achieved using genome editing technologies like CRISPR/Cas9 and prime editing (PE). These fine-tuning strategies avoid the negative side effects often caused by traditional overexpression with strong promoters, allowing for a moderate increase in target gene expression. Precisely adjusting endogenous protein levels through uORF editing has great potential for crop improvement. This includes improving broad-spectrum disease resistance and stress tolerance, optimizing traits like the growth period, and increasing nutritional quality, such as vitamin content. To help establish an efficient precision crop breeding system based on uORF editing, this review explains the main characteristics and molecular mechanisms of uORFs, summarized current uORF editing strategies and progress in trait improvement. Also discussed are t future applications and challenges of uORF editing in precision crop breeding, and the prospect of using deep learning models to improve the efficiency and predictability of uORF editing.

Key words: uORF, translational regulation, gene editing, precision breeding, 5′UTR

虞布卡, 谭瑗瑗, 舒庆尧. 基于uORF编辑的作物精准育种研究进展与展望[J]. 中国农业科学, 2026, 59(21): 4697-4709.

YU BuKa, TAN YuanYuan, SHU QingYao. Progress and Prospects of Crop Precision Breeding Based on uORF Editing[J]. Scientia Agricultura Sinica, 2026, 59(21): 4697-4709.

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调控策略 Regulatory strategy	作用区域 Target region	调控水平 Regulatory level	作用机制 Mechanism of action	参考文献 References
uORF编辑 uORF editing	5′UTR	翻译水平 Translational	消除uORF对下游mORF翻译抑制，增加核糖体对mORF的翻译 Eliminating uORF-mediated translational inhibition of the downstream mORF, thereby increasing ribosome recruitment and translation of the mORF	[11]
启动子编辑 Promoter editing	启动子 Promoter	转录水平 Transcriptional	破坏转录抑制因子结合位点，提高转录效率 Disrupting transcriptional repressor binding sites to enhance transcription efficiency	[40]
3′UTR编辑 3′UTR editing	3′UTR	转录后水平 Post-transcriptional	敲除3′UTR负调控区域（通常包含 miRNA特异性识别的顺式元件或形成比较稳定RNA二级结构的区域） Knocking out negative regulatory regions in the 3′UTR (typically containing cis-elements for specific miRNA recognition or regions forming stable RNA secondary structures)	[41]
染色体结构重排 Chromosomal rearrangement	基因组大片段 Large genomic segments	转录水平 Transcriptional	通过染色体的倒置或重复，让同一条染色体上的强启动子去驱动弱表达的基因 Leveraging chromosomal inversions or duplications to allow a strong promoter on the same chromosome to drive the expression of a weakly expressed target gene	[42]

类别 Category	数据库名称 Name of the database	适用物种、输入数据 Applicable species or input data	主要功能与特色 Main functions and features	参考文献 References
鉴定工具 Identification tool	ORFfinder	序列 Sequence	NCBI的经典工具。常用于对5′UTR序列进行初步的ORF识别 A classic tool of NCBI. It is often used for the initial ORF scanning of 5′UTR sequences	[49]
	uORF-seqr	Ribo-seq数据、RNA-seq数据、基因组注释文件 Ribo-seq data, RNA-seq data, genomic annotation files	高效鉴定非AUG起始的近同密码子uORF，并给出每个uORF的显著性评估值 Efficiently identify near-similar codon uORFs that do not start with AUG, and provide a significance evaluation value for each uORF	[50]
	uORF4u	序列 Sequence	支持原核生物预测其核糖体结合位点 Support the prediction of ribosome binding sites for prokaryotes	[51]
数据库 Database	uORFlight	拟南芥、水稻等 Arabidopsis thaliana, Oryza sativa, etc.	可在种群尺度评估uORF出现频率，通过数据库挖掘，找出单核苷酸多态性或插入/缺失 By mining the database, it enables the evaluation of uORF occurrence frequencies at the population level and the identification of SNPs or InDels	[52]
	PsORF	拟南芥、棉花、水稻、玉米等 Arabidopsis thaliana, Gossypium spp., Oryza sativa, Zea mays, etc.	植物小ORF库，包含uORF、dORF等共5类小ORF Plant small ORF library. It includes five types of small ORFs such as uORF and dORF	[53]
	uORFdb	人、拟南芥等 Homo sapiens, Arabidopsis thaliana, etc.	计算预测、试验文献与群体变异数据深度融合 The deep integration of computational prediction, experimental literature and population variation data	[54]

方法 Method	物种 Species	基因 Gene	突变体性状 Mutant trait	参考文献 References
缺失uORF Delete uORF	水稻O. sativa	Hd2	延迟开花Delay flowering	[46]
	水稻O. sativa	GLW7	改善粒型Improve grain shape	[45]
	水稻 O. sativa	GW7	增加粒长和长宽比 Increased grain length and length/width ratio	[45]
	大麦H. vulgare	Mla13	加快免疫应答Accelerate immune response	[57]
	生菜L. sativa	GGP2	增加维生素C含量Increase vitamin C content	[11]
	拟南芥 A. thaliana	bZIP11	消除蔗糖的抑制效应 Eliminate the inhibitory effect of sucrose	[58]
生成uORF Generate uORF	水稻O. sativa	Waxy	降低直链淀粉含量Reduce amylose content	[59]
生成uORF Generate uORF	水稻O. sativa	DLT	降低株高和分蘖数Reduce plant height and tiller numbers	[60]
改变uORF长度 Change uORF length	水稻O. sativa	Hd2	延迟开花Delay flowering	[46]
改变uORF长度 Change uORF length	生菜L. sativa	GGP1	增加维生素C含量Increase vitamin C content	[11]
转换uORF种类 Switch uORF types	水稻O. sativa	GS2	增强抗病性Enhance disease resistance	[61]
转换uORF种类 Switch uORF types	番茄Solanum lycopersicum	RabGAP22	增强耐盐性Enhance salt tolerance	[62]
改变uORF起始密码子上下文序列 Mutant uORF initiation codon context	水稻O. sativa	Waxy	降低直链淀粉含量Reduce amylose content	[63]
改变uORF起始密码子上下文序列 Mutant uORF initiation codon context	番茄S. lycopersicum	GGP1	增加维生素C含量Increase vitamin C content	[64]
偶联环境响应型 uORF Coupled environment-responsive uORF	水稻O. sativa	NPR1	抗病但正常生长Resistance but minimal fitness costs	[55]
偶联环境响应型 uORF Coupled environment-responsive uORF	拟南芥A. thaliana	snc1	抗病但正常生长Resistance but minimal fitness costs	[55]