Identification of MYB gene family and functional analysis of ElMYB114 in lipid synthesis of Euphorbia lathyris L.
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摘要:
MYB转录因子广泛参与植物的代谢和种子发育等生命过程。为探究其在续随子（Euphorbia lathyris L.）油脂合成及积累过程中的调控作用，本研究对续随子MYB转录因子进行了全基因组鉴定，分析了其基因结构、编码蛋白理化性质、启动子顺式元件以及时空表达谱，并克隆了ElMYB114基因的编码序列，将其转入烟草（Nicotiana tabacum L.），进一步鉴定该基因的生物学功能。结果显示：（1）全基因组鉴定获得199个续随子MYB基因，命名为ElMYB1~ElMYB199，编码蛋白均具有SANT结构域；（2）ElMYB114编码的蛋白定位于细胞核；（3）荧光定量qRT-PCR分析结果表明，ElMYB114基因在种子发育后期的表达量最高；（4）与转空载对照相比，异源表达ElMYB114基因的烟草株系，其叶片总油脂含量分别提高了3.7%和4.2%，种子总脂含量分别提高了11.0%和10.7%，表明过表达ElMYB114基因可提高植物组织的总油脂积累。研究为深入解析MYB介导的续随子油脂生物合成调控网络奠定了基础。
Abstract:MYB transcription factors play pivotal roles in plant metabolism and seed development. This study conducted a comprehensive investigation into the regulatory functions of these transcription factors in lipid synthesis and accumulation in Euphorbia lathyris L.. The analysis included genome-wide identification of MYB transcription factors, characterization of gene structures, physicochemical properties of encoded proteins, distribution of promoter cis-acting elements, and spatiotemporal expression patterns. The ElMYB114 coding sequence was cloned and genetically transformed into tobacco (Nicotiana tabacum L.) to characterize its biological function. Genome-wide identification revealed 199 MYB genes in E. lathyris, designated ElMYB1 to ElMYB199, all encoding proteins with SANT domains. The protein encoded by ElMYB114 was localized to the nucleus. Quantitative fluorescence qRT-PCR analysis showed that ElMYB114 expression peaked during the late stage of seed development. Overexpression of ElMYB114 in transgenic tobacco lines led to a 3.7% and 4.2% increase in total oil content in leaves and an 11.0% and 10.7% increase in total lipid content in seeds compared to the empty vector control. These findings suggest that ElMYB114 enhances carbon flux within the heterologous host toward lipid synthesis pathways, thereby promoting lipid accumulation in plant tissues. This study provides foundational insights into the MYB-mediated regulatory networks governing oil biosynthesis in E. lathyris.
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图 1 续随子ElMYB基因的染色体定位
Figure 1. Chromosomal localization of ElMYB genes in Euphorbia lathyris
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图 2 ElMYB基因在续随子不同组织中的相对表达量
不同小写字母表示在P<0.05水平差异显著，下同。
Figure 2. Relative expression level of ElMYB genes in different tissues of Euphorbia lathyris
Different lowercase letters in the same column indicate significant differences at 0.05 level. Same below.
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图 3 转基因株系烟草叶片中ElMYB114基因的表达量
Figure 3. Expression of ElMYB114 gene in tobacco leaves of transgenic plants
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图 4 ElMYB114的亚细胞定位
A：GFP，B：DAPI暗场；C：明场；D：叠加场。
Figure 4. Subcellular localization of ElMYB114
A: GFP, B: DAPI dark field for green fluorescence; C: Bright field; D: Observed in combination.
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图 5 不同基因型烟草叶片总油脂含量
WT：野生型；EV：pCAMBIA1303空载转烟草；OE1、OE2：转基因株系。下同。
Figure 5. Total lipid content in tobacco leaves
WT: Wild type; EV: pCAMBIA1303 no-load; OE1, OE2: Transgenic strain. Same below.
表 1 引物信息
Table 1 Primer information
引物名称
Primer name引物序列（5'→3'）
Primer sequence (5'→3')用途
ApplicationElAction-F ATGTATGTCGCCATTCAAGCCGTTC 内参 ElAction-R TCATCAGAGCATCAGTAAGGTCACG ElMYB33-F TGCATGCTAGATGGGGGAAC 实时荧光定量PCR ElMYB33-R TGTCGCAGTTTTCGGGTACT ElMYB63-F ATTCGAGTTCCAGCGCTACG ElMYB63-R GATCCATCCGAGTTCGCACA ElMYB100-F CACCGCAAGGACCGATAACT ElMYB100-R ATATGCCGATACCAGGGGGT ElMYB111-F GGACGAGATGTCGCAATGGA ElMYB111-R TAACACGTGGCTTTGGACGA ElMYB114-F TCGAGGAATGTGGTGGTGAC ElMYB114-R AGGGTTGCTCTCCACACTTG ElMYB188-F AACGTTACTCCTCAAGCCCG ElMYB188-R CTGAGGAGGCAATTGAGCCA ElMYB114-1303-F ATGACCGCTTGCTGGACTCG 基因克隆 ElMYB114-1303-R TCACATTGGGGGGAAGTTATAGTT ElMYB114-1300-F ATGACCGCTTGCTGGACTCG ElMYB114-1300-R CATTGGGGGGAAGTTATAGTTTTG 
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表 2 续随子ElMYB家族蛋白的基本理化性质
Table 2 Basic information of ElMYB family proteins
蛋白类型
Protein type数量
Quantity分子量
Molecular
weight / kD氨基酸数
Number of amino acids等电点
Theoretical pI不稳定指数
Instability index / (II)亲水性系数
GRAVY亚细胞定位
Subcellular
localizationMYB-related 78 8.23~219.86 72~2 026 4.25~9.93 32.06~89.20 −1.316~−0.137 细胞核 R2R3-MYB 115 12.34~177.75 105~1 619 4.62~9.98 35.43~81.68 −1.220~−0.319 细胞核 3R-MYB 5 39.08~119.47 330~1 081 5.26~9.00 43.07~60.47 −0.642~−1.220 细胞核 4R-MYB 1 118.07 1 043 6.94 55.65 −0.828 细胞核 合计 199 8.23~219.86 72~2 026 4.25~9.98 32.06~89.20 −1.316~−0.137 细胞核
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表 3 续随子ElMYB基因上游启动子顺式作用元件
Table 3 Cis-acting elements in the upstream promoter of ElMYB genes of Euphorbia lathyris
顺式作用元件
Cis-acting element核心序列
Core sequence功能
FunctionWRE3 CCACCT 损伤诱导元件 LTR CCGAAA 低温响应元件 MYB CAACAG MYB结合位点 Myc TCTCTTA MYC结合位点 MRE AACCTAA 与光反应有关的MYB结合位点 WUN-motif TTATTACAT 创伤反应元件 STRE AGGGG 热休克、营养饥饿和低pH诱导响应元件 GATA-motif GATAGGA 光响应元件 CAAT-box CAAT 启动子和增强子区域的共同顺式作用元件 TC-rich repeats GTTTTCTTAC 参与防御和应激反应的顺式作用元件 GT1-motif GGTTAA 光响应元件 Box 4 ATTAAT 参与光反应的保守DNA模块的一部分 MYB-like sequence TAACCA MYB结合位点 ABRE ACGTG 脱落酸响应元件 ABRE3a TACGTG 脱落酸响应元件 AAGAA-motif GAAAGAA 脱落酸响应元件 ABRE4 CACGTA 脱落酸响应元件 TATA-box TATAAATA 启动子基本元件 P-box CCTTTTG 赤霉素响应元件 Myb-binding site CAACAG MYB结合位点 ARE AAACCA 厌氧响应元件 G-box TACGTG 光响应相关的顺式作用元件 TCA-element CCATCTTTTT 水杨酸反应涉及的顺式作用元件 3-AF3 bind site CACTATCTAAC 光应答 A-box CCGTCC α-淀粉酶启动子保守序列 ABRE2 CCACGTGG 脱落酸响应元件 AT1-motif AATTATTTTTTATT 光应答 AE-box AGAAACTT 光响应模块 ATCT-motif AATCTAATCC 部分光反应元件 as-1 TGACG 压力响应元件 CAT-box GCCACT 与分生组织表达相关的顺式作用元件 TCA TCATCTTCAT 水杨酸响应元件 TGA-box TGACGTAA 部分生长素响应元件 MBS CAACTG 与干旱诱导有关的MYB结合位点
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