Thomas A, Gray K D, Roth M J
Department of Biochemistry, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway 08854, USA.
Virology. 1997 Jan 20;227(2):305-13. doi: 10.1006/viro.1996.8333.
The role of the cytoplasmic tail of the Moloney murine leukemia virus transmembrane protein in the regulation of syncytia was examined. Three mutations within the cytoplasmic tail were studied. Linker-insertion in7705-12a is within the viral-associated cytoplasmic tail, linker-insertion in7748-12a is within the R peptide, and a third mutation expresses TM lacking the R peptide (Env R-). The Env R- construct was nonviable in Rat1 cells, however, rapidly reverted to a form containing the R peptide when passaged in NIH/3T3 cells. in7705-12a was temperature-sensitive in Rat1 cells, as previously characterized, but was viable at either temperature in NIH/3T3 cells. in7748-12a was comparable with wild-type M-MuLV. The ability of the env constructs to form large multinucleated syncytia with NIH/3T3 and XC cells were examined using transient expression assays, eliminating reversion events due to viral passage and reverse transcription. The Env R- constructs formed syncytia with NIH/3T3 cells. in7705-12a displays enhanced proteolytic cleavage of the R peptide. Neither linker-insertion mutation in7705-12a or in7748-12a activated fusion with NIH/3T3, despite the abundance of processed TM with in7705-12a. All three mutants were fusion competent with Rat XC cells, even in the absence of any cleavage of the R peptide. These results provide insights regarding steric and the temporal affects of cleavage of the R peptide and the assembly of a fusion competent oligomer.
研究了莫洛尼鼠白血病病毒跨膜蛋白的细胞质尾部在合胞体调控中的作用。对细胞质尾部的三个突变进行了研究。7705 - 12a中的接头插入位于病毒相关的细胞质尾部内，7748 - 12a中的接头插入位于R肽内，第三个突变表达缺失R肽的跨膜蛋白（Env R-）。Env R-构建体在大鼠1细胞中无法存活，然而，在NIH/3T3细胞中传代时会迅速回复为含有R肽的形式。7705 - 12a在大鼠1细胞中是温度敏感的，如先前所述，但在NIH/3T3细胞中在任何一个温度下都能存活。7748 - 12a与野生型M - MuLV相当。使用瞬时表达测定法检测了env构建体与NIH/3T3和XC细胞形成大型多核合胞体的能力，消除了由于病毒传代和逆转录导致的回复事件。Env R-构建体与NIH/3T3细胞形成了合胞体。7705 - 12a显示出R肽的蛋白水解切割增强。尽管7705 - 12a中有大量加工后的跨膜蛋白，但7705 - 12a或7748 - 12a中的接头插入突变均未激活与NIH/3T3的融合。即使在没有R肽任何切割的情况下，所有三个突变体与大鼠XC细胞都具有融合能力。这些结果提供了关于R肽切割的空间和时间影响以及融合活性寡聚体组装的见解。
