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| 东方蜜蜂突触囊泡糖蛋白2B基因的生物信息学和表达模式 |
| Bioinformatics and expression pattern of synaptic vesicle glycoprotein 2B gene in Apis cerana |
| 投稿时间：2024-06-28 修订日期：2024-10-01 |
| DOI： |
| 中文关键词: 东方蜜蜂 突触囊泡糖蛋白 分子特征 系统进化 表达模式 |
| 英文关键词:Apis cerana synaptic vesicle glycoprotein 2B molecular characteristics phylogenetic tree conserved sequence motifs |
| 基金项目:国家自然科学基金面上项目(32372943,32172792); 国家现代农业产业技术体系建设专项资金(CARS-44-KXJ7); 福建农林大学硕士生导师团队项目(郭睿); 福建省自然科学基金面上项目(2022J01131334, 2023J01133656); 福建农林大学科技创新专项基金(KFb22060XA) |
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| 中文摘要: |
| 【目的】解析东方蜜蜂(Apis cerana)突触囊泡糖蛋白2B(synaptic vesicle glycoprotein 2B, AcSV2B)的理化性质和分子特征,检测AcSV2B基因在东方蜜蜂工蜂不同组织和发育阶段的表达模式,丰富AcSV2B信息并为进一步的功能研究提供基础。【方法】通过Stem-loop RT-PCR和Sanger测序验证AcSV2B在东方蜜蜂工蜂的表达和序列真实性。通过相关生物信息学软件预测AcSV2B的理化性质、信号肽、磷酸化位点、二级结构及三级结构,鉴定和比较东方蜜蜂和其他物种SV2B的结构域及保守基序,并进行系统进化树的构建和分析。采用实时荧光定量PCR (RT-qPCR)检测AcSV2B在工蜂的触角、中肠、脑、毒腺、表皮、咽下腺和脂肪体等不同组织,卵、幼虫、预蛹、蛹和成虫,以及不同日龄工蜂成虫中的相对表达量。【结果】AcSV2B在东方蜜蜂工蜂体内真实存在,符合预测结果。AcSV2B含3 814个核苷酸,编码671个氨基酸；AcSV2B的分子量约为72.87 ku,分子式为C3317H5162N842O940S31,脂溶系数为102.32,等电点为6.02,包含24个丝氨酸磷酸化位点和11个跨膜结构域,不含典型的信号肽,主要分布于内质网。AcSV2B含6个结构域和5个保守基。东方蜜蜂、高加索蜂(Apis mellifera caucasica)、小蜜蜂(Apis florea)和西方蜜蜂(Apis mellifera)的SV2B在进化树上聚为一支。AcSV2B的表达量在脑中最高且显著高于触角、中肠、毒腺、表皮、咽下腺和脂肪体中的表达量(P＜0.05)；AcSV2B在8日龄预蛹中的表达量最高且显著高于卵、3日龄幼虫和7日龄预蛹中的表达量(P＜0.05)；AcSV2B在18日龄成虫中的表达量最高且显著高于1、2、6和12日龄成虫中的表达量(P＜0.05)。【结论】AcSV2B可能是一种亲水性蛋白、跨膜蛋白和胞内蛋白,东方蜜蜂、高加索蜂、小蜜蜂和西方蜜蜂的SV2B保守性和同源性较高,AcSV2B在脑、8日龄预蛹和18日龄成虫中特异性高表达。 |
| 英文摘要: |
| 【Objective】The objective of this work is to enrich the information about AcSV2B and provide a basis for further functional dissection, by analyzing the physicochemical property and molecular characteristics of the Apis cerana synaptic vesicle glycoprotein 2B protein (AcSV2B)and detecting the expression profile of AcSV2B gene in different tissues and developmental stages of worker.【Method】The expression and sequence authenticity of AcSV2B in Apis cerana worker bees were verified by Stem-loop RT-PCR and Sanger sequencing. Relevant bioinformatic software were employed to predict the physicochemical property, signal peptide, phosphorylation site, secondary structure and tertiary structure of AcSV2B, to identify and compare structural domains and conserved motifs in SV2B among Apis cerana and other species followed by construction and investigation of the phylogenetic tree. RT-qPCR was utilized to detect the relative expression level of AcSV2B in various tissues of worker including antenna, midgut, brain, venom gland, epidermis, hypopharyngeal gland and fat body, in egg, larva, prepupa, pupa and adult of worker, and worker adults at different day-old.【Result】AcSV2B is really present in the worker bees of Apis cerana, which is consistent with the predicted results. AcSV2B contained 3 814 nucleotides, encoding 671 amino acids. The molecular weight of AcSV2B was approximately 72.87 ku, the molecular formula was C3317H5162N842O940S31, the aliphatic index was 102.32, and the theoretical pI was 6.02. AcSV2B included 24 serine phosphorylation sites and 11 transmembrane domains, but there was no typical signal peptides. AcSV2B was mainly distributed in the endoplasmic reticulum. Additionally, AcSV2B included 6 structural domains and 5 conserved motifs. SV2B from Apis cerana, Apis mellifera caucasica, Apis florea and Apis mellifera were clustered into a clade on the phylogenetic tree. Furthermore, the relative expression level of AcSV2B was the highest in brain and significantly higher (P＜0.05) than that in antenna, midgut, venomous gland, epididymis, hypopharyngeal gland and fat body; the relative expression level of AcSV2B was the highest in 8-day-old prepupa and significantly higher (P＜0.05) than that in egg, the 3-day-old larva, and the 7-day-old prepupa; the relative expression level of AcSV2B was the highest in 18-day-old adult and significantly higher (P＜0.05) than that in 1-, 2-, 6- and 12-day-old adults.【Conclusion】AcSV2B may be a hydrophilic protein, transmembrane protein and intracellular protein, The SV2B of Apis cerana, Apis mellifera caucasica, Apis florea and Apis mellifera were highly conserved and homologous,and AcSV2B was highly expressed in brain, 8-day-old prepupal and 18-day-old adult. |
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