Wahyudi Anung, Ariyani Dinni, Ma Gang, Inaba Ryosuke, Fukasawa Chikako, Nakano Ryohei, Motohashi Reiko
Graduate School of Science and Technology, Shizuoka University, 3-5-1 Johoku, Naka-ku, Hamamatsu, Shizuoka 432-8561, Japan.
Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, Shizuoka 422-8529, Japan.
Plant Biotechnol (Tokyo). 2018 Dec 25;35(4):303-312. doi: 10.5511/plantbiotechnology.18.0620a.
In this study, two temperature-induced lipocalin genes and , and a chloroplastic lipocalin gene were isolated from 'Micro-Tom' tomato. The coding sequences of , and were 558, 558, and 1002 bp, respectively. By TargetP analysis, no characteristic transit peptides were predicted in the proteins of SlTIL1 and SlTIL2, while a chloroplastic transit peptide was predicted in the protein of SlCHL. The subcellular localization results indicated that SlTIL1 and SlTIL2 proteins were major localized in the plasma membrane, while SlCHL was localized in chloroplast. To understand the function of lipocalins, transgenic tomato over-expressed , and and their virus-induced gene silencing (VIGS) plants were generated. The phenotypes were significantly affected when the , and were over-expressed or silenced by VIGS, which suggested that the three lipocalins played important roles in regulating the growth and development of tomato. In addition, the level of ROS (O and HO) was low in , and over-expressed plants, while it was high in their silenced plants. The changes in the expression of s were consistent with the accumulations of ROS, which indicated that lipocalins might have an important role in abiotic oxidative stress tolerance in tomato plants. Especially SlTIL1 and SlTIL2 are localized around their membranes and protect them from ROS. The results will contribute to elucidating the functions of lipocalin in plants, and provide new strategies to improve the tolerance to abiotic stress in tomato plants.
在本研究中，从“Micro-Tom”番茄中分离出两个温度诱导的脂质运载蛋白基因SlTIL1和SlTIL2，以及一个叶绿体脂质运载蛋白基因SlCHL。SlTIL1、SlTIL2和SlCHL的编码序列分别为558、558和1002 bp。通过TargetP分析，在SlTIL1和SlTIL2的蛋白质中未预测到特征性转运肽，而在SlCHL的蛋白质中预测到一个叶绿体转运肽。亚细胞定位结果表明，SlTIL1和SlTIL2蛋白主要定位于质膜，而SlCHL定位于叶绿体。为了解脂质运载蛋白的功能，构建了过表达SlTIL1、SlTIL2和SlCHL的转基因番茄及其病毒诱导基因沉默（VIGS）植株。当SlTIL1、SlTIL2和SlCHL过表达或通过VIGS沉默时，番茄表型受到显著影响，这表明这三种脂质运载蛋白在调控番茄生长发育中发挥重要作用。此外，在SlTIL1、SlTIL2和SlCHL过表达植株中活性氧（O₂和H₂O₂）水平较低，而在其沉默植株中活性氧水平较高。SlTIL1、SlTIL2和SlCHL表达的变化与活性氧的积累一致，这表明脂质运载蛋白可能在番茄植株非生物氧化胁迫耐受性中起重要作用。特别是SlTIL1和SlTIL2定位于其膜周围并保护它们免受活性氧的侵害。这些结果将有助于阐明植物中脂质运载蛋白的功能，并为提高番茄植株对非生物胁迫的耐受性提供新策略。
