Masuda T, Harada S
Department of Biodefence and Medical Virology, Kumamoto University Medical School, Japan.
J Virol Methods. 1990 Jun;28(3):219-33. doi: 10.1016/0166-0934(90)90116-w.
Retroviral RNA is copied into DNA by reverse transcriptase when the viral genome enters into its life cycle. In the case of human immunodeficiency virus (HIV), massive amounts of unintegrated viral DNA reportedly appear in the early phase of primary infection. However, the relationship between the accumulation of this DNA and the cytopathic effect (CPE) remains obscure. In an attempt to delineate this association, we examined the appearance of the unintegrated viral DNA by means of two experimental systems: (1) primary infection of highly susceptible MOLT-4#8 cells and (2) induction of CPE by cell-fusion of persistently infected MOLT-4#8 cells. A correlation was observed between the accumulation of unintegrated viral DNA and the appearance of CPE, both when MOLT-4#8 cells were infected with cell-free virus and when persistently infected MOLT-4#8 cells were co-cultured with uninfected cells. Persistently infected cells did not fuse spontaneously in culture, because they lack the CD4-molecule on their surfaces. However, when treated with polyethylene glycol (PEG), the cells fused, exhibited ballooning degeneration, and released fewer viruses. After PEG treatment, unintegrated viral DNA also appeared. Since such DNA is generally not detected in persistently infected cells, it is possible that some cellular mechanism exists to suppress the synthesis of viral DNA and that the fusion induced by PEG treatment cancels the suppression. Treatment of persistently infected cells with Ca2+ ionophore and Ca2+ antagonist also resulted in the accumulation of unintegrated viral DNA and inhibited virus release. These findings suggest that the induction of unintegrated HIV DNA may be an effective strategy for reducing the release of the virus.
当病毒基因组进入其生命周期时，逆转录酶会将逆转录病毒RNA复制成DNA。就人类免疫缺陷病毒（HIV）而言，据报道在初次感染的早期阶段会出现大量未整合的病毒DNA。然而，这种DNA的积累与细胞病变效应（CPE）之间的关系仍不清楚。为了阐明这种关联，我们通过两个实验系统研究了未整合病毒DNA的出现情况：（1）对高度易感的MOLT-4#8细胞进行初次感染，以及（2）通过持续感染的MOLT-4#8细胞与未感染细胞的细胞融合诱导CPE。当MOLT-4#8细胞用无细胞病毒感染时，以及当持续感染的MOLT-4#8细胞与未感染细胞共培养时，均观察到未整合病毒DNA的积累与CPE的出现之间存在相关性。持续感染的细胞在培养中不会自发融合，因为它们表面缺乏CD4分子。然而，用聚乙二醇（PEG）处理后，细胞发生融合，出现气球样变性，并释放出较少的病毒。PEG处理后，未整合的病毒DNA也出现了。由于在持续感染的细胞中通常检测不到这种DNA，因此可能存在某种细胞机制来抑制病毒DNA的合成，并且PEG处理诱导的融合消除了这种抑制作用。用Ca2+离子载体和Ca2+拮抗剂处理持续感染的细胞也导致未整合病毒DNA的积累并抑制病毒释放。这些发现表明，诱导未整合的HIV DNA可能是减少病毒释放的一种有效策略。
