Mycoplasma gallisepticum (HS strain) surface lipoprotein pMGA interacts with host apolipoprotein A-I during infection in chicken
Mycoplasma gallisepticum (HS strain) surface lipoprotein pMGA interacts with host apolipoprotein A-I during infection in chicken
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鸡毒支原体（HS 株）表面脂蛋白 pMGA 在鸡感染过程中与宿主载脂蛋白 A-I 相互作用
DOI：
10.1007/s00253-015-7117-9
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发表时间：
2016-02-01
影响因子：
5
通讯作者：
Peng, Xiuli
中科院分区：
文献类型：
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作者：
Hu, Fuli;Zhao, Chengcheng;Peng, Xiuli
鸡毒支原体（HS株）的粘附素蛋白pMGA 1.2是鸡毒支原体（M.鸡败血症（MG）感染鸡。然而，与pMGA1.2相互作用的宿主因子尚不清楚。本研究从鸡胚气管上皮细胞中分离出细胞膜组分。使用改进的病毒覆盖蛋白印迹分析（VOPBA）和谷胱甘肽S-转移酶（GST）下拉分析，我们发现pMGA1.2特异性结合约30 kDa的宿主蛋白。该宿主蛋白通过质谱进一步鉴定为鸡载脂蛋白A-I（ApoA-I）。我们在大肠杆菌中表达并纯化了重组ApoA-I蛋白，并在体外证实其与纯化的pMGA1.2蛋白结合。瞬时表达的pMGA1.2和ApoA-I在HeLa细胞中共定位。最后，我们设计了小干扰RNA（siRNA）分子，敲低ApoA-I或pMGA1.2的表达，从而抑制MG诱导的鸡胚成纤维细胞系（DF-1）细胞周期紊乱。同样，敲低ApoA-I抑制MG感染鸡气管细胞的纤毛损失和损伤。因此，ApoA-I可能通过与pMGA1.2相互作用而成为MG感染的重要宿主因子。
The adhesin protein from Mycoplasma gallisepticum (HS strain), namely pMGA1.2, is required for M. gallisepticum (MG) infection in chicken. However, the host factor(s) that interact with pMGA1.2 is not known. In this study, we prepared the membrane fraction of trachea epithelial cells from chicken embryos. Using an improved virus overlay protein blot assay (VOPBA) and glutathione S-transferase (GST) pull-down assay, we found that pMGA1.2 specifically bound to a ∼30 kDa host protein. This host protein was further identified by mass spectrometry as chicken apolipoprotein A-I (ApoA-I). We expressed and purified the recombinant ApoA-I protein in Escherichia coli and confirmed that it bound to the purified pMGA1.2 protein in vitro. Transiently expressed pMGA1.2 and ApoA-I were colocalized in HeLa cells. Finally, we designed small interfering RNA (siRNA) molecules to knock down the expression of either ApoA-I or pMGA1.2, which inhibited the MG-induced cell cycle disruption in cells of chicken embryo fibroblast cell line (DF-1). Similarly, knockdown of ApoA-I inhibited the cilia loss and damage in chicken trachea cells in MG infection. In summary, ApoA-I may be an essential host factor in MG infection through interacting with pMGA1.2.