Holland J J, Villarreal L P, Breindl M
J Virol. 1976 Mar;17(3):805-15. doi: 10.1128/JVI.17.3.805-815.1976.
Previous indications that cloned B virions might be genetically predisposed to generate a particular defective T particle are shown to be inaccurate. T particle generation was found to be a much more random process than was previously believed. We show that the previously observed generation of particular sizes of T particles by B virion pools is due to the random generation of T particles during preparation of first-passage pools of cloned B virions, and these breed true during the additional passages needed to produce visible quantities of T particles. It is also shown that different host cell lines selectively amplify different T particles, suggesting a strong role of host cell factors in T particle replication. Surprisingly, our line of HeLa cells did not generate or replicate detectable T particles of vesicular stomatitis virus (VSV) Indiana after either serial undiluted passage or direct addition of T particles, even though the added T particles strongly interfered with B virion replication. In contrast to VSV, rabies virus generates large amounts of T particles during the first passage of cloned B virions, and every rabies-infected baby hamster kidney-21 cell culture evolves into a persistent carrier state. We find that T particle RNA is biologically inactive although T particle nucleocapsid ribonucleoprotein replicates and interferes in cells coinfected with B virions. Efforts to study the mechanism of T particle generation by in vitro attempts to generate T particles or modify their size (using sheared ribonucleoprotein or chemical or UV mutagenesis) were unsuccessful. The kinetics of UV and nitrous acid inactivation of T particles indicate a smaller target size relative to B virions, even after correcting for lengths of RNA molecules. The intercalating dye proflavine does not photosensitize VSV B virions or T particles when present during replication, indicating that there is little or no RNA base pairing in the helical nucleocapsids of either.
先前有关克隆的B病毒粒子可能在遗传上易于产生特定缺陷T粒子的迹象被证明是不准确的。发现T粒子的产生是一个比先前认为的更为随机的过程。我们表明，先前观察到的B病毒粒子库产生特定大小的T粒子是由于在克隆B病毒粒子的第一代培养物制备过程中T粒子的随机产生，并且在产生可见数量的T粒子所需的后续传代过程中这些粒子能够稳定遗传。还表明不同的宿主细胞系选择性地扩增不同的T粒子，这表明宿主细胞因子在T粒子复制中起重要作用。令人惊讶的是，我们的HeLa细胞系在连续未稀释传代或直接添加T粒子后，均未产生或复制可检测到的水疱性口炎病毒（VSV）印第安纳株的T粒子，尽管添加的T粒子强烈干扰了B病毒粒子的复制。与VSV相反，狂犬病病毒在克隆B病毒粒子的第一代传代过程中产生大量T粒子，并且每个感染狂犬病的幼仓鼠肾-21细胞培养物都会演变成持续携带状态。我们发现T粒子RNA在生物学上无活性，尽管T粒子核衣壳核糖核蛋白在与B病毒粒子共感染的细胞中复制并产生干扰。通过体外尝试产生T粒子或改变其大小（使用剪切的核糖核蛋白或化学或紫外线诱变）来研究T粒子产生机制的努力均未成功。T粒子的紫外线和亚硝酸灭活动力学表明，即使校正了RNA分子的长度，其靶标大小相对于B病毒粒子也较小。在复制过程中存在嵌入染料原黄素时，它不会使VSV B病毒粒子或T粒子发生光致敏，这表明两者的螺旋核衣壳中几乎没有或不存在RNA碱基配对。
