家蚕DNA甲基转移酶功能预测及其在BmNPV感染下的表达特征

Functional prediction of DNA methyltransferase in silkworm and expression characteristics under BmNPV infection

  • 摘要: 【目的】明确家蚕DNA甲基转移酶基因(BmDNMT)生物信息学基本特征及其在家蚕核型多角体病毒(BmNPV)感染前后的表达情况,揭示DNA甲基化在家蚕抗病毒免疫方面的作用机制,为挖掘家蚕抗病毒标志基因和药物靶标蛋白等提供理论依据。【方法】通过OrthoDB、MultiLoc2、SherLoc2、PSORTII、SMART、SWISS-MODEL及Schrödinger等在线生物信息学分析软件进行BmDNMT蛋白氨基酸同源比对、系统进化分析、亚细胞定位、功能结构域预测、三级结构同源建模及小分子配体对接口袋预测,并采用实时荧光定量PCR分析BmDNMT基因在家蚕感染BmNPV后不同时间不同组织中的时空表达特征。【结果】BmDNMT基因包含BmDNMT1BmDNMT2,分别位于家蚕8号染色体和11号染色体上,其推导BmDNMT氨基酸序列均与烟草天蛾DNMT氨基酸序列的亲缘关系最近。BmDNMT1蛋白大量存在于细胞核,少量分布在细胞质;而BmDNMT2蛋白大量存在于细胞质,少量分布在细胞核及线粒体。BmDNMT1蛋白的功能结构域多于BmDNMT2蛋白,二者均含有DNA甲基化酶功能结构域,且具有潜在药物结合口袋。BmNPV感染会影响BmDNMT1BmDNMT2基因在家蚕不同组织中表达差异,BmNPV感染4 h后这2个基因在家蚕中肠的表达差异已非常明显,说明家蚕DNA甲基化可能在病毒感染早期已发生,且中肠可能是最早响应的组织。BmNPV感染后,BmDNMT1BmDNMT2基因的相对表达量和表达趋势并不一致。【结论】BmDNMT1BmDNMT2基因的染色体位置、亚细胞定位及功能结构域等存在差异,BmNPV感染后二者在家蚕不同组组织中的相对表达量和表达趋势也不一致,推测BmDNMT1BmDNMT2基因在家蚕DNA甲基化过程中行使不同的功能。BmDNMT1和BmDNMT2蛋白三维结构具有潜在的药物结合口袋,可能是直接影响DNA甲基化状态的药物靶标。

     

    Abstract: 【Objective】To clarify basic bioinformatics characteristics of Bombyx mori DNA methyltransferase gene(BmDNMT)in silkworm and its expression before and after infected with Bombyx mori nuclearpolyhedrosisvirus(BmNPV)and to reveal mechanism of DNA methylation in antiviral immunity of silkworm,so as to provide theoretical basis for mining antiviral marker genes and drug target proteins of silkworm.【Method】OrthoDB,MultiLoc2,SherLoc2,PSORTII,SMART,SWISS-MODEL and Schrödinger were used to analyze BmDNMT protein amino acid homology,phylogenetic analysis,subcellular localization and functional domain prediction,tertiary structure homology modeling and small molecule ligand docking pocket prediction,and quantitative real-time PCR(qRT-PCR)was used to analyze temporal and spatial expression characteristics of BmDNMT gene in different tissues of silkworm infected with BmNPV at different times.【Result】BmDNMT gene contained BmDNMT1 and BmDNMT2,which were located on chromosome 8 and chromosome11,respectively. Amino acid sequence of BmDNMT was closely related to that in Manduca sexta. A large amount of BmDNMT1 proteins were in the nucleus and a small amount in the cytoplasm. However,BmDNMT2 protein was abundant in cytoplasm and small in nucleus and mitochondria. BmDNMT1 protein had more functional domains than BmDNMT2 protein,both of which contained DNA methylase functional domains and had potential drug-binding pockets. BmNPV infection could affect expression difference of BmDNMT1 and BmDNMT2 genes in different tissues of silkworm. 4 h after BmNPV infection,the expression difference of these 2 genes in the midgut of silkworm was very obvious,suggesting that DNA methylation of silkworm might have occurred in the early stage of virus infection,and the midgut may be the earliest tissue to respond. After BmNPV infection,relative expression and expression trends of BmDNMT1 and BmDNMT2 genes were not consistent.【Conclusion】Differences in the chromosomal location,subcellular location and functional domains of BmDNMT1 and BmDNMT2 genes are found,and relative expression and expression trends of BmDNMT1 and BmDNMT2 genes in different tissues of silkworm after BmNPV infection are also not consistent,suggesting that BmDNMT1 and BmDNMT2 genes play different roles during DNA methylation. The three-dimensional structures of BmDNMT1 and BmDNMT2 proteins have potential drug-binding pockets and may be drug targets that directly affect DNA methylation status.

     

/

返回文章
返回