College of Horticulture, Shenyang Agricultural University, Key Laboratory of Protected Horticulture of Education Ministry and Liaoning Province, National & Local Joint Engineering Research Center of Northern Horticultural Facilities Design & Application Technology (Liaoning), Liaoning Shenyang, 110866, People's Republic of China.
BMC Plant Biol. 2019 Feb 15;19(1):75. doi: 10.1186/s12870-019-1678-1.
Lipoxygenases (LOXs) play significant roles in abiotic stress responses, and identification of LOX gene promoter function can make an important contribution to elucidating resistance mechanisms. Here, we cloned the CmLOX08 promoter of melon (Cucumis melo) and identified the main promoter regions regulating transcription in response to signalling molecules and abiotic stresses.
The 2054-bp promoter region of CmLOX08 from melon leaves was cloned, and bioinformatic analysis revealed that it harbours numerous cis-regulatory elements associated with signalling molecules and abiotic stress. Five 5'-deletion fragments obtained from the CmLOX08 promoter-2054 (LP1), 1639 (LP2), 1284 (LP3), 1047 (LP4), and 418 bp (LP5)-were fused with a GUS reporter gene and used for tobacco transient assays. Deletion analysis revealed that in response to abscisic acid, salicylic acid, and hydrogen peroxide, the GUS activity of LP1 was significantly higher than that of the mock-treated control and LP2, indicating that the - 2054- to - 1639-bp region positively regulates expression induced by these signalling molecules. However, no deletion fragment GUS activity was induced by methyl jasmonate. In response to salt, drought, and wounding treatments, LP1, LP2, and LP4 promoted significantly higher GUS expression compared with the control. Among all deletion fragments, LP4 showed the highest GUS expression, indicating that - 1047 to - 1 bp is the major region regulating promoter activity and that the - 1047 to - 418-bp region positively regulates expression induced by salt, drought, and wounding, whereas the - 1284 to - 1047-bp region is a negative regulatory segment. Interestingly, although the GUS activity of LP1 and LP2 was not affected by temperature changes, that of LP3 was significantly induced by heat, indicating that the - 1284- to - 1-bp region is a core sequence responding to heat and the - 2054- to - 1284-bp region negatively regulates expression induced by heat. Similarly, the - 1047- to - 1-bp region is the main sequence responding to cold, whereas the - 2054- to - 1047-bp region negatively regulates expression induced by cold.
We cloned the CmLOX08 promoter and demonstrated that it is a signalling molecule/stress-inducible promoter. Furthermore, we identified core and positive/negative regulatory regions responding to three signalling molecules and five abiotic stresses.
脂氧合酶（LOXs）在非生物胁迫响应中发挥重要作用，鉴定 LOX 基因启动子功能可以为阐明抗性机制做出重要贡献。在这里，我们克隆了甜瓜（Cucumis melo）的 CmLOX08 启动子，并鉴定了主要的启动子区域，以调节对信号分子和非生物胁迫的转录反应。
从甜瓜叶片中克隆了 2054bp 的 CmLOX08 启动子区域，生物信息学分析表明它包含与信号分子和非生物胁迫相关的许多顺式调控元件。从 CmLOX08 启动子-2054（LP1）、1639（LP2）、1284（LP3）、1047（LP4）和 418bp（LP5）获得的五个 5'缺失片段与 GUS 报告基因融合，并用于烟草瞬时测定。缺失分析表明，在响应脱落酸、水杨酸和过氧化氢时，LP1 的 GUS 活性明显高于对照和 LP2，表明-2054 至-1639bp 区域正向调节这些信号分子诱导的表达。然而，茉莉酸甲酯处理没有诱导任何缺失片段的 GUS 活性。在盐、干旱和创伤处理下，LP1、LP2 和 LP4 促进了比对照更高的 GUS 表达。在所有缺失片段中，LP4 表现出最高的 GUS 表达，表明-1047 至-1bp 是主要的调节启动子活性的区域，-1047 至-418bp 区域正向调节盐、干旱和创伤诱导的表达，而-1284 至-1047bp 区域是一个负调控片段。有趣的是，尽管 LP1 和 LP2 的 GUS 活性不受温度变化的影响，但 LP3 的 GUS 活性在受热时显著增加，表明-1284 至-1bp 区域是响应热的核心序列，-2054 至-1284bp 区域负调节热诱导的表达。同样，-1047 至-1bp 区域是响应冷的主要序列，而-2054 至-1047bp 区域负调节冷诱导的表达。
我们克隆了 CmLOX08 启动子，并证明它是一个信号分子/胁迫诱导启动子。此外，我们鉴定了响应三个信号分子和五种非生物胁迫的核心和正/负调控区域。
