Influence of Mixed Retrovirus Infections on Leukemia and Neurological disease

小鼠白血病病毒(MULV)引起小鼠多种疾病，涉及到称为多嗜性MULV的逆转录病毒的参与。这包括导致增殖性、免疫性和神经性疾病。多向性MuLV是近交系小鼠基因组中存在的外源性生态MULV与内源性包膜序列重组形成的，导致病毒利用不同的细胞表面受体进行感染。生态型MuLV的感染宿主范围仅限于小鼠；然而，用生态型MuLV接种小鼠后产生的重组多角体病毒能够感染许多其他物种以及小鼠。因此，多嗜性病毒的产生导致具有不同感染特性的病毒的混合逆转录病毒感染。我们早期的研究有力地表明，宿主中生态型和多向性MuLV之间的相互作用在促进肿瘤发生中发挥了作用。最近，我们研究了逆转录病毒在体内混合感染中的相互作用，方法是将多嗜性MuLV分离株和生态性MuLV株混合接种小鼠。与感染单一逆转录病毒的小鼠相比，感染特定逆转录病毒混合物的小鼠表现出显著的病理变化。这些包括一种多嗜性MuLV在引起增殖性疾病方面的非常显著的延迟，以及一种深刻的协同效应，导致与另一种多嗜性分离株突然发展成一种神经系统疾病。在这两种情况下，联合接种的小鼠的多嗜性病毒载量显着增加，而生态型MuLV的水平没有变化。此外，在联合接种的小鼠中，多嗜性MuLV几乎完全是生态型病毒粒子内的假型。 有许多可能的机制可以促进多嗜性病毒在体内的深刻扩增，包括由于生态型病毒粒子内的伪分型而促进病毒的传播，或者可能是多嗜性病毒在共感染细胞中的反式激活。为了在一个不那么复杂的系统中检查这些问题，我们将这些研究扩展到检查体外细胞系的混合逆转录病毒感染。我们发现，多嗜性MuLV与生态型或两性型病毒混合感染后，其扩增和伪分型特征与我们在体内观察到的非常相似。混合感染细胞释放的多向性感染性显著增加，而生态性感染性保持不变。此外，传染性的增加伴随着生态型病毒粒子内多变基因组的广泛伪分型。这种观察延伸到来自不同组织和不同小鼠的细胞系，表明这是混合感染的成分的特征，而不是细胞的特性。从混合感染细胞释放的多嗜性MuLV感染力的提高可能是由于释放的多嗜性基因组水平的增加。或者，观察到的增加可能反映了与多嗜性病毒粒子相比，生嗜性或两性嗜性病毒粒子的特定传染性要高得多。我们已经发现，在具有生性或两性MuLV的多嗜性MuLV混合物中，多嗜性病毒滴度的增加至少有一部分可以归因于共同感染细胞的多嗜性基因组包装和释放效率的提高。 对释放不同水平的多嗜性病毒的克隆细胞的分析表明，当这些克隆细胞与一种生态病毒共感染时，每个克隆细胞都可以被诱导释放类似的高水平的多嗜性病毒。这些结果进一步表明，与一种生态病毒的混合感染促进了多角体基因组的包装和释放，这可能反映了多角体包膜的固有缺陷。在这方面，与生态型和两性型病毒不同，重组多角体病毒是嵌合病毒，其中病毒的包膜没有与其他结构基因共同进化。因此，多嗜性病毒的包膜蛋白在包装和释放后代病毒时可能不那么有效。 2014年，我们继续对克隆细胞株进行研究，这些克隆细胞株释放非常低水平的接近潜伏感染的传染性多嗜性病毒。我们已经证实，重叠感染生态型MuLV后释放的传染性显着增加，然而释放的病毒粒子RNA水平仅适度增加，仅占传染性大幅增加(10,000倍)的一小部分。从每个克隆细胞系中获得的病毒在传到新细胞上时表现出正常的感染性，这表明病毒释放的低水平并不是由于每个克隆中存在有缺陷的病毒分离物。与生态型MuLV的重叠感染相反，克隆细胞系与第二种多嗜性病毒重叠感染不会导致感染性的显著增加。此外，在感染的克隆细胞系中，与正常细胞感染相比，重叠感染的多嗜性病毒的复制大大减少。这些结果表明，克隆细胞系的低水平复制是细胞本身的一种特性，并仅限于多向类MuLV。生态型MuLV的重叠感染绕过了这一限制。发生这种情况的机制(S)正在进一步调查中。

The induction of many diseases in mice by murine leukemia viruses (MuLVs), involves the participation of variant retroviruses termed polytropic MuLVs. These include the induction of proliferative, immunological and neurological disorders. Polytropic MuLVs are formed by recombination of exogenous ecotropic MuLVs with endogenous envelope sequences present in the genomes of inbred mouse strains resulting in viruses which utilize a distinct cell-surface receptor for infection. The infectious host range of ecotropic MuLVs is limited to mice; however the recombinant polytropic viruses generated after inoculation of mice with ecotropic MuLVs are capable of infecting a number of other species as well as mice. Thus, the generation of polytropic viruses results in a mixed retrovirus infection of viruses with different infectious properties. Our earlier studies strongly suggest that the interactions of ecotropic and polytropic MuLVs in the host play a role in facilitating oncogenesis. More recently we have investigated the interactions of retroviruses in mixed infections in vivo by co-inoculation of mice with mixtures of polytropic MuLV isolates and ecotropic MuLVs. Mice infected with defined mixtures of retroviruses exhibit dramatically altered pathology compared to infection with the individual viruses of the mixture. These included a highly significant delay in the induction of proliferative disease with one polytropic MuLV and a profound synergistic effect resulting in the abrupt development of a neurological disease with another polytropic isolate. In both instances the polytropic virus load in the co-inoculated mice was markedly enhanced while the level of the ecotropic MuLV was unchanged. Furthermore, the polytropic MuLV was nearly completely pseudotyped within ecotropic virions in co-inoculated mice. There are a number of possible mechanisms which could facilitate the profound in vivo amplification of the polytropic MuLVs including enhanced spread of the virus due to pseudotyping within ecotropic virions or possibly transactivation of the polytropic virus in co-infected cells. To examine these questions in a less complex system we have extended these studies to examine mixed retrovirus infections of an in vitro cell line. We have found that co-infection of polytropic MuLVs with ecotropic or amphotropic viruses results in amplification and pseudotyping characteristics remarkably similar to what we have observed in vivo. The polytropic infectivity released from co-infected cells is markedly increased while the ecotropic infectivity remains unaltered. Further, the increase in infectivity is accompanied by extensive pseudotyping of the polytropic genome within ecotropic virions. This observation extended to cell lines from different tissues and different mice indicating that it was a feature of the components of the mixed infection rather than a property of the cells. The elevation of polytropic MuLV infectivity released from co-infected cells could have resulted from an increase in the level of polytropic genomes released. Alternatively, the observed increase could reflect a much higher specific infectivity of ecotropic or amphotropic virions compared to polytropic virions. We have found in polytropic MuLV mixtures with either ecotropic or amphotropic MuLVs, that at least some of the increase in polytropic virus titer can be attributed to an increase in the efficiency of packaging and release of the polytropic genome from co-infected cells. Analyses of clonal cell lines releasing different levels of polytropic viruses indicated that each of these lines could be induced to release similar high levels of polytropic virus upon co-infection of these cells with an ecotropic virus. These results further suggest that co-infection with an ecotropic virus facilitates the packaging and release of the polytropic genome, possibly reflecting an inherent defectiveness of tthe polytropic envelope. In this regard, unlike ecotropic and amphotropic viruses, recombinant polytropic viruses are chimeric viruses in which the envelope of the virus has not co-evolved with the other structural genes. Thus the envelope protein of polytropic viruses may not function as efficiently in packaging and release of progeny viruses. In 2014 we have continued studies with clonal cell lines releasing very low levels of infectious polytropic viruses that approach a latent infection. We have confirmed a remarkable increase in infectivity released upon superinfection with ecotropic MuLVs however the level of released virion RNA is only moderately increased and accounts for only a small portion of the profound increase in infectivity (10,000-fold). Virus harvested from each of the clonal cell lines exhibit normal infectivity when passaged onto new cells indicating that the low level of virus release is not due to the presence of a defective virus isolate in each of the clones. In contrast to superinfection by an ecotropic MuLV, superinfection of the clonal cell lines with a second polytropic virus does not result in a substantial increase in infectivity. Moreover, the replication of the superinfected polytropic virus is greatly diminished in the infected clonal cell lines compared to infection of normal cells. These results suggest that the low level of replication of the clonal cell lines is a property of the cells themselves and is restricted to the polytropic class of MuLVs. Superinfection with ecotropic MuLV circumvents this restriction. The mechanism(s) by which this occurs is under further investigation.