Yugawa T, Amanuma H
Laboratory of Gene Technology and Safety, Tsukuba Life Science Center, The Institute of Physical and Chemical Research (RIKEN), Ibaraki, Japan.
J Virol. 1998 Mar;72(3):2272-9. doi: 10.1128/JVI.72.3.2272-2279.1998.
We previously reported (N. Watanabe, M. Nishi, Y. Ikawa, and H. Amanuma, J. Virol. 65:132-137, 1991) that the mutant Friend spleen focus-forming virus (F-SFFV(MS)), which encodes a mutant gp55 membrane glycoprotein with an ecotropic env gp70 sequence, was nonpathogenic. Here we injected the F-SFFV(MS)-Friend murine leukemia virus (F-MuLV) clone 57 complex into newborn DBA/2 mice. We obtained four groups of pathogenic variant F-SFFV complexes, each showing a different degree of pathogenicity in adult mice and a different gp55 profile. Of these, group 1 variant F-SFFV was particularly interesting, because it was the most frequently obtained and because it produced doublet bands of gp55 (59 and 57 kDa), neither of which reacted with the nonecotropic gp70-specific monoclonal antibody, and because its DNA intermediate did not hybridize with the nonecotropic env-specific probe. Cloning and DNA sequence analysis of the env region of one isolate of the group 1 variant F-SFFV revealed that this virus consisted of two distinct F-SFFV genomes; one (clone 117) differed from the other (clone 118) due to the presence of a 39-bp in-frame deletion. Reconstitution to full-length F-SFFV genomes and a pathogenicity assay showed that each reconstituted F-SFFV was pathogenic, with clone 117 showing a higher degree of pathogenicity than clone 118. Both reconstituted F-SFFVs caused activation of the mouse erythropoietin receptor in the factor-independent cell proliferation assay, although much less efficiently than the wild-type polycythemia-inducing isolate F-SFFVp. Clone 118 produced a gp55 of 59 kDa, while clone 117 produced one of 57 kDa. Clone 118 had a substitution by the F-MuLV clone 57 gp70 sequence, indicating that it was derived from the F-SFFV(MS) env gene by a homologous recombination with the F-MuLV clone 57 env gene. The site of the 39-bp deletion in clone 117 corresponded to the portion of the clone 118 sequence which was unique to the ecotropic env genes. These results indicated the importance for the biological activity of gp55 of the sequences in the gp70 differential region, which are contained in both polytropic and ecotropic env genes.
我们之前报道过（N. 渡边、M. 西、Y. 池川和 H. 天沼，《病毒学杂志》65:132 - 137，1991），编码具有嗜亲性 env gp70 序列的突变型 gp55 膜糖蛋白的突变型弗瑞德脾集落形成病毒（F - SFFV(MS)）无致病性。在此，我们将 F - SFFV(MS) - 弗瑞德鼠白血病病毒（F - MuLV）克隆 57 复合物注射到新生的 DBA/2 小鼠体内。我们获得了四组致病性变异的 F - SFFV 复合物，每组在成年小鼠中表现出不同程度的致病性以及不同的 gp55 图谱。其中，第 1 组变异型 F - SFFV 特别有趣，因为它是最常获得的，并且它产生了 gp55 的双峰带（59 和 57 kDa），这两条带均不与非嗜亲性 gp70 特异性单克隆抗体发生反应，还因为其 DNA 中间体不与非嗜亲性 env 特异性探针杂交。对第 1 组变异型 F - SFFV 的一个分离株的 env 区域进行克隆和 DNA 序列分析表明，该病毒由两个不同的 F - SFFV 基因组组成；其中一个（克隆 117）与另一个（克隆 118）不同，原因是存在一个 39 碱基对的框内缺失。将其重构成全长 F - SFFV 基因组并进行致病性测定表明，每种重构的 F - SFFV 都具有致病性，克隆 117 的致病性高于克隆 118。在非依赖因子的细胞增殖试验中，两种重构的 F - SFFV 均能激活小鼠促红细胞生成素受体，尽管效率远低于野生型诱导红细胞增多症的分离株 F - SFFVp。克隆 118 产生 59 kDa 的 gp55，而克隆 117 产生 57 kDa 的 gp55。克隆 118 具有由 F - MuLV 克隆 57 的 gp70 序列替代的情况，表明它是通过与 F - MuLV 克隆 57 的 env 基因同源重组从 F - SFFV(MS) 的 env 基因衍生而来。克隆 117 中 39 碱基对缺失位点对应于克隆 118 序列中嗜亲性 env 基因特有的部分。这些结果表明，多嗜性和嗜亲性 env 基因中所含的 gp70 差异区域的序列对 gp55 的生物学活性很重要。
