Rodriguez J M, Salas M L, Santarén J F
Centro de Biologia Molecular Severo Ochoa, Universidad Autónoma de Madrid, Spain.
Proteomics. 2001 Nov;1(11):1447-56. doi: 10.1002/1615-9861(200111)1:11<1447::AID-PROT1447>3.0.CO;2-Y.
High-resolution two-dimensional electrophoresis followed by computer analysis has been used to study quantitatively the patterns of protein synthesis produced in porcine alveolar macrophages and in Vero cells infected with African swine fever virus (ASFV). Initially, a protein database for each cell type was constructed. The porcine alveolar macrophage database includes 995 polypeptides (818 acidic, isoelectric focusing (IEF) and 177 basic, nonequilibrium pH gradient electrophoresis (NEPHGE)) whereas the Vero database contains 1,398 polypeptides (1,127 acidic, IEF and 271 basic, NEPHGE). Taking these databases as reference, ASFV highly virulent strain E70 induces 57 acid and 43 basic polypeptides in porcine alveolar macrophages, which account for most of the information content of the virus DNA. The kinetics of synthesis of the virus-induced polypeptides showed the existence of three classes of proteins: one whose synthesis starts early after infection, continues for a period and then switches off; another whose synthesis also starts early but continues for prolonged periods; and a third which requires DNA replication. The attenuated, cell adapted, strain BA71V induces 92 acidic and 37 basic proteins in Vero cells. Significant differences were observed when comparing the patterns of polypeptides induced by the two viral strains. In both cell systems studied, ASFV infection produces a general shutoff of protein synthesis that affects up to 65% of the cellular proteins. Interestingly, 28 proteins of porcine alveolar macrophages and 48 proteins of Vero cells are stimulated at least two times by ASFV infection.
高分辨率二维电泳结合计算机分析已被用于定量研究猪肺泡巨噬细胞以及感染非洲猪瘟病毒（ASFV）的非洲绿猴肾细胞（Vero细胞）中产生的蛋白质合成模式。最初，构建了每种细胞类型的蛋白质数据库。猪肺泡巨噬细胞数据库包含995种多肽（818种酸性多肽，采用等电聚焦（IEF）法；177种碱性多肽，采用非平衡pH梯度电泳（NEPHGE）法），而Vero细胞数据库包含1398种多肽（1127种酸性多肽，IEF法；271种碱性多肽，NEPHGE法）。以这些数据库为参考，ASFV强毒株E70在猪肺泡巨噬细胞中诱导产生57种酸性多肽和43种碱性多肽，这些多肽占病毒DNA信息含量的大部分。病毒诱导多肽的合成动力学表明存在三类蛋白质：一类在感染后早期开始合成，持续一段时间后停止；另一类也在早期开始合成，但持续较长时间；第三类需要DNA复制。减毒的、适应细胞的BA71V毒株在Vero细胞中诱导产生92种酸性蛋白质和37种碱性蛋白质。比较两种病毒毒株诱导的多肽模式时观察到显著差异。在所研究的两种细胞系统中，ASFV感染都会导致蛋白质合成普遍停止，影响多达65%的细胞蛋白质。有趣的是，ASFV感染至少两次刺激猪肺泡巨噬细胞中的28种蛋白质和Vero细胞中的48种蛋白质。
