Antimicrobial functional divergence of the cecropin antibacterial peptide gene family in Musca domestica
Antimicrobial functional divergence of the cecropin antibacterial peptide gene family in Musca domestica
复制标题
家蝇天蚕素抗菌肽基因家族的抗菌功能差异
DOI：
10.1186/s13071-019-3793-0
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发表时间：
2019-11-14
影响因子：
3.2
通讯作者：
Wu，Jianwei
中科院分区：
文献类型：
--
作者：
Peng，Jian;Wu，Zhaoying;Wu，Jianwei
据报道，家蝇中有十多种抗菌肽（amp）属于抗菌肽家族；然而，它们中很少被识别出来，而其他分子的功能也知之甚少。方法：它们的序列。从三龄幼虫总mRNA中反转录cDNA模板，克隆了驯化ecropin家族基因。受到病原体挑战的家养动物。采用DNAMAN综合分析软件进行序列分析，根据成熟肽序列在MEGA v.5.0中采用neighbor - joining法构建cecropin家族分子系统发育树。抗菌活性的研究。采用肉汤微量稀释法检测国产acecropin蛋白，测定最低抑菌浓度（MIC）。对革兰氏阴性菌鲍曼不动杆菌在对数生长和稳定生长阶段进行时间杀伤试验，并通过扫描电镜（SEM）和检测260 nm吸收材料的渗漏来评估其在Cec4处理下的形态变化。结果cec01、cec02和cec1 -9在m的Scaffold18749上的多基因家族中与Cec形式具有同源性。有明显。比较编码的天蚕素蛋白序列，大部分具有天蚕素家族的基本特征，含有19个保守氨基酸残基。据我们所知，这是第一次实验证明Cec家族中的大多数基因都是功能性的。Cec02、Cec1、Cec2、Cec5和Cec7对革兰氏阴性菌的抑菌光谱和抑菌效果相似，而Cec4抗菌活性谱更广，抑菌效果非常强。baumannii。Cec4 eliminatedA。鲍曼菌素快速且呈浓度依赖性，在1× MIC和2× MIC条件下，24 h内均有抑菌效果。SEM分析和260 nm吸波材料渗漏检测表明，Cec4通过破坏细胞膜完整性对细菌进行了杀菌。结论虽然有十多个与toM相关的抗菌肽基因。除Cec4外，部分菌株对sta没有较好的抑菌活性。baumannii。
BackgroundIt has been reported that there are more than ten antimicrobial peptides (AMPs) belonging to the cecropin family inMusca domestica; however, few of them have been identified, and the functions of the other molecules are poorly understood.MethodsSequences of theM. domesticacecropin family of genes were cloned from cDNA template, which was reverse-transcribed from total mRNA isolated from third-instar larvae ofM. domesticathat were challenged with pathogens. Sequence analysis was performed using DNAMAN comprehensive analysis software, and a molecular phylogenetic tree of the cecropin family was constructed using the Neighbour-Joining method in MEGA v.5.0 according to the mature peptide sequences. Antibacterial activity of the syntheticM. domesticacecropin protein was detected and the minimum inhibitory concentration (MIC) values were determined using broth microdilution techniques. Time-killing assays were performed on the Gram-negative bacteria,Acinetobacter baumannii, at the logarithmic or stabilizing stages of growth, and its morphological changes when treated with Cec4 were assessed by scanning electron microscopy (SEM) and detection of leakage of 260 nm absorbing material.ResultsEleven cecropin family genes, namelyCec01,Cec02andCec1-9, show homology to the Cec form in a multigene family on the Scaffold18749 ofM. domestica. In comparing the encoded cecropin protein sequences, most of them have the basic characteristics of the cecropin family, containing 19 conservative amino acid residues. To our knowledge, this is the first experimental demonstration that most genes in the Cec family are functional. Cec02, Cec1, Cec2, Cec5 and Cec7 have similar antibacterial spectra and antibacterial effects against Gram-negative bacteria, while Cec4 displays a more broad-spectrum of antimicrobial activity and has a very strong effect onA. baumannii. Cec4 eliminatedA. baumanniiin a rapid and concentration-dependent manner, with antibacterial effects within 24 h at 1× MIC and 2× MIC. Furthermore, SEM analysis and the leakage of 260 nm absorbing material detection indicated that Cec4 sterilized the bacteria through the disruption of cell membrane integrity.ConclusionsAlthough there are more than ten cecropin genes related toM. domestica, some of them have no preferred antibacterial activity other than Cec4 againstA. baumannii.