新生隐球菌毒力差异基因型菌株间部分毒力相关基因分析  

Analysis of virulence-associated gene s in Cryptococcus neoformans va r.grub ii strains oftwo geno-types with different virulence

作　　者：陈玉如[1] 赵亮[1] 金方[1] 李小玲[1] 曹煜[2] 康颖倩[1] 

机构地区：[1]贵阳医学院微生物学教研室,550004 [2]贵阳医学院附属医院皮肤性病科

出　　处：《中华微生物学和免疫学杂志》2014年第8期577-582,共6页Chinese Journal of Microbiology and Immunology

基　　金：国家自然基金地方项目（31060006号及31260029号）;贵州省社会发展科技攻关项目（黔科合sY字[2011]3017号）;贵阳市科技局社会发展与民生计划（筑科合同[2011103]16号）

摘　　要：目的：了解新生隐球菌格鲁比变种（ Cryptococcus neoformans var． grubii）两组毒力差异明显的多位点微卫星型（ multilocus microsatellite typing ， MLMT ）菌株间的差异基因，探讨导致新生隐球菌格鲁比变种间毒力差异的相关基因及这些基因的功能。方法根据已有基因组数据，针对可能为新生隐球菌毒力相关的基因设计特异性引物，扩增并测序，通过序列对比分析每个基因在两组微卫星基因型菌株间存在的单核苷酸多态性（single nucleotide polymorphisms ，SNPs）位点，并对引起差异的蛋白质进行分析和预测。结果在成功测序的11个基因中，除2个基因序列完全相同外，其余9个基因在两组毒力差异明显的基因型菌株间分别存在1～4个规律性SNPs，其中，有2个基因中存在非同义SNPs（ non-synonymous SNPs ，nsSNPs ），这2个基因的编码产物分别为沉默信息调节因子2家族成员HST302和UV切除修复蛋白RAD23。另外，在对RAD23蛋白的系统发育分析发现，与真菌界的其他门类相比，在担子菌纲的多数RAD23蛋白与人类的同源基因亲缘关系更近。结论本研究发现了新生隐球菌两种毒力差异明显的微卫星基因型菌株间具有重要功能的两个差异基因（ Rad23和Hst302），且经预测nsSNPs对蛋白结构和功能均有一定影响，而蛋白的具体功能有待通过各项试验进一步深入研究。Objective To identify non-synonymous single nucleotide polymorphisms （ nsSNPs ） with virulence-associated genes of Cryptococcus neoformans var.grubii strains of two genotypes （ MLMT13 and MLMT36 ） and to investigate their effects on virulence .Methods Specific primers were designed ac-cording to the existing genome database and used to amplify the fragments of virulence -associated genes of Cryptococcus neoformans.The cloned gene fragments were sequenced and aligned with the sequences of Crpy -tococcus neoformans var.grubii strains of MLMT13 and MLMT36 genotypes to analyze SNPs .nsSNPs were screened out to analyze the structure and function of their corresponding proteins .Results Eleven viru-lence-associated genes were sequenced successfully .The alignment analysis showed that besides two identi-cal genes , the other nine genes presented various numbers of SNPs ranging from one to four .nsSNPs were detected in two genes encoding protein HST 302 which was the family member of silent information regular 2 （SIR2） and the UV excision repair protein RAD23, respectively.The phylogenetic analysis demonstrated that most of the RAD23 proteins found in basidiomycetes shared higher homologies with their human orthologs than with those found in other fungal phyla .Conclusion Two virulence-associated genes , Rad23 and Hst302, that determined the virulence difference among Cryptococcus neoformans strains of the two genotypes were detected by using sequencing analysis .The predicated structure and function of their encoded proteins were affected by the existence of nsSNPs to some extent .

关 键 词：新生隐球菌格鲁比变种 微卫星基因分型 毒力 蛋白 

分 类 号：R519.4[医药卫生—内科学]