马尔尼菲篮状菌磷脂酶A基因家族鉴定与表达

Identification and expression of the phospholipase A gene family in Talaromyces marneffei

目的对马尔尼菲篮状菌(Talaromyces marneffei)磷脂酶A(phospholipase A,PLA)基因家族进行鉴定分析,为进一步研究马尔尼菲篮状菌PLA潜在致病作用和机制奠定基础。方法利用生物信息学方法对马尔尼菲篮状菌PLA基因家族成员进行全基因组鉴定,分析其基因结构、染色体定位、蛋白理化性质和结构、保守基序及系统进化关系,并利用实时定量反转录聚合酶链式反应(real-time quantitative reverse transcription polymerase chain reaction,qRT-PCR)方法分析PLA基因在该菌侵染巨噬细胞时的表达水平。结果从马尔尼菲篮状菌基因组中鉴定到8个PLA,不均匀分布在5条染色体上,不存在串联重复和片段复制现象。PLA家族氨基酸数目为577~1546个,相对分子质量为65120~170690,等电点为4.47~9.28,均为亲水蛋白,除PLA3外的其余PLA成员属于不稳定蛋白。系统进化分析表明马尔尼菲篮状菌PLA可分为PLA2和patatin-like磷脂酶2个亚族,成员数量分别为3和5。同一亚族基因结构、蛋白保守基序和结构较为相似,但不同亚族之间具有明显差异。qRT-PCR结果表明,马尔尼菲篮状菌侵染小鼠巨噬细胞时PLA3、PLA6和PLA8表达上调。结论PLA3、PLA6和PLA8可能与马尔尼菲篮状菌能够在巨噬细胞内生存繁殖有关。

Objective To identify and analyze the phospholipase A(PLA)gene family in Talaromyces marneffei(T.marneffei),providing the basis for further research on the potential pathogenic effect and mechanism of T.marneffei PLA.Methods The members of the PLA gene family in T.marneffei were genome-wide identified by using bioinformatics,and their gene structure,chromosome distribution,protein physicochemical property and structure,conserved motif,as well as phyletic evolution,were analyzed.The expression levels of PLA genes in T.marneffei under macrophage treatment were analyzed using the qRT-PCR method.Results A total of eight PLA genes were identified from the genome of T.marneffei,unevenly located on 5 chromosomes without any tandem duplications or fragment duplications.The amino acid residues of PLA proteins ranged from 577 to 1546,molecular weights from 65120 to 170690,and the isoelectric point from 4.47 to 9.28.All PLA proteins were hydrophilic,with all but PLA3 classified as unstable hydrophilic proteins.Phylogenetic analysis revealed that PLA was divided into two subfamilies:PLA2 and patatin-like phospholipase,consisting of 3 and 5 members respectively.Within the same subfamily,gene structures,protein-conserved motifs,and protein structures were relatively similar,whereas significant differences existed between different subfamilies.qRT PCR results showed that the expression levels of PLA3,PLA6,and PLA8 were upregulated in T.marneffei infected with mouse macrophages.Conclusions PLA3,PLA6,and PLA8 may be related to the survival and reproduction of T.marneffei in macrophages.