LI Yuanyuan,YU Qiuyue,SHANG Qinwang,et al.Preliminary Analysis of Subcellular Localization and Pathogenicity of Little cherry virus 1 Protein P3.5[J].Northern Horticulture,2023,(16):48-52.[doi:10.11937/bfyy.20224507]
樱桃小果病毒1编码的P3.5蛋白生物学功能初探
- Title:
- Preliminary Analysis of Subcellular Localization and Pathogenicity of Little cherry virus 1 Protein P3.5
- 文献标志码:
- A
- 摘要:
- 以樱桃小果病毒-1（Little cherry virus 1,LChV-1）甜樱桃分离物为试材，采用分子克隆及浓杆菌侵染等方法，初步研究了其基因组编码的P3.5蛋白的亚细胞定位及致病性特征，以期为研究植物病毒编码的小分子量蛋白的生物学功能提供参考依据。结果表明：1）P3.5基因全长为96 nt，编码31个氨基酸，大小约3.5 kDa的跨膜蛋白。2）通过构建了带有GFP标签的荧光表达载体35S::P35-GFP，浸润本氏烟叶片后利用激光共聚焦显微镜观察，发现P35-GFP在细胞膜和细胞核膜均有分布。3）将P3.5构建到马铃薯X病毒（Potato virus X，PVX），以此获得异源表达载体的PVX-P3.5。将PVX-P35接种本氏烟，14 d后，发现接种PVX-P3.5的本氏烟新叶并没有出现典型的症状，同时接种PVX空载体的植株也仅有轻微的花叶、黄化，表明在PVX异源表达系统 中，P35并没有明显增强PVX的致病性而诱导寄主植物出现严重的症状，暗示P35可能不是作为症状形成的致病因子在病毒的侵染 中发挥作用。
- Abstract:
- Little cherry virus 1 (LChV-1) was as research subject in this study.The subcellular localization and pathogenicity of P3.5 protein were studied by using molecular biology methods,in order to provide reference for the studying of the very small proteins encoded by plant viruses.The results showed that,1) P3.5 was encoded by ORF2 of LChV-1,a small hydrophobic protein of unknown function.Primarily,P3.5 was cloned from an isolate of LChV-1 from Tai′an,Shandong province using specific primers by RT-PCR,which was 96 bp in length and encoded a protein of approximately 3.5 kD with a transmembrane domain.2) Subcellular localization analysis by tobacco leaves showed that P3.5 localized in cytomembrane and nuclear membrane.3) To further clarify the pathogenicity of P3.5,a potato virus X (PVX)-based vector was introduced and PVX-P3.5 was constructed.The PVX-P3.5 was transformed into Agrobacterium tumefaciens GV3101 and infiltrated into the N.benthamiana.Slight mosaic symptom was observed in the plants inoculated by PVX 14 days later.Meanwhile,when the plants inoculated by PVX-P3.5,the plants didn′t have visible symptoms.It demonstrated that heterologous expression of P3.5 in N.benthamiana didn′t enhanced the pathogenicity of PVX,and not be related to the symptom-formation.
参考文献/References:
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备注/Memo
第一作者简介：李媛媛（1989-），女，博士，讲师，现主要从事作物病毒基因表达调控等研究工作。E-mail：lyy 329715173@126.com.责任作者：王德亚（1989-），男，博士，副教授，现主要从事植物病毒学等研究工作。E-mail：wangdeyasdny@163.com.基金项目：国家自然科学基金资助项目（32102143）；山东省自然科学基金资助项目（ZR2019PC011，ZR2020QC122）；枣庄学院博士启动基金资助项目（2018BS040）；枣庄市科技计划资助项目（NS03）。收稿日期：2022-11-05