Osterrieder N, Neubauer A, Brandmüller C, Kaaden O R, O'Callaghan D J
Institute for Medical Microbiology, Infectious and Epidemic Diseases, Ludwig-Maximilians-University Munich, Germany.
Virology. 1996 Dec 15;226(2):243-51. doi: 10.1006/viro.1996.0652.
The diploid IR6 gene (ORF 67) of equine herpesvirus type 1 (EHV-1) is absent in the modified live EHV-1 vaccine strain RacH and is present in a mutated form in the avirulent EHV-1 strains RacM24 and RacM36, such that the IR6 protein fails to form the typical rod-like structures observed for wild-type EHV-1 RacL11. To assess the role of the IR6 protein in EHV-1 replication and virulence, two recombinant RacH viruses, HIR6-1 and HIR6-2, that harbor a single copy of the wild-type IR6 gene were engineered and characterized. It was shown that: (i) HIR6-1 or HIR6-2 virus encoded for an IR6 protein that was capable of forming the rod-like structures typical of cells infected with the wild-type virulent virus strain RacL11. (ii) Whereas the avirulent EHV-1 strains RacH and RacM36 are temperature-sensitive (in that virus replication at 40 degrees versus that at 37 degrees was reduced by as much as 7,500-fold), the HIR6-1 and HIR6-2 viruses, like RacL11 virus, were capable of significant replication at the elevated temperature. (iii) Electron microscopic analyses revealed that cells infected with the HIR6-1 or HIR6-2 virus, like those infected with virulent RacL11 virus, produced and released comparable numbers of virus particles at both 37 and 40 degrees. In cells infected with the RacH virus at 40 degrees, however, release of extracellular particles was inhibited by greater than 90% and relatively few of the particles were enveloped. (iv) Infections of BALB/cA mice revealed that both the HIR6-1 and HIR6-2 viruses, unlike the parent RacH virus, were as virulent as the wild-type RacL11 strain as judged by the criteria of body weight loss, development of clinical signs of EHV-1 infection, virus titers in the lung, and ability to cause viremia. These findings and those of our recent studies indicate that the IR6 protein is a major determinant of EHV-1 virulence and that the IR6 protein may play a role in virus maturation and/or egress.
1型马疱疹病毒（EHV-1）的二倍体IR6基因（开放阅读框67）在改良活EHV-1疫苗株RacH中不存在，而在无毒力的EHV-1株RacM24和RacM36中以突变形式存在，因此IR6蛋白无法形成野生型EHV-1 RacL11所观察到的典型杆状结构。为了评估IR6蛋白在EHV-1复制和毒力中的作用，构建并鉴定了两种携带单拷贝野生型IR6基因的重组RacH病毒HIR6-1和HIR6-2。结果表明：（i）HIR6-1或HIR6-2病毒编码的IR6蛋白能够形成野生型强毒病毒株RacL11感染细胞典型的杆状结构。（ii）无毒力的EHV-1株RacH和RacM36对温度敏感（在40℃时病毒复制与37℃时相比减少多达7500倍），而HIR6-1和HIR6-2病毒与RacL11病毒一样，在升高的温度下能够大量复制。（iii）电子显微镜分析显示，感染HIR6-1或HIR6-2病毒的细胞与感染强毒RacL11病毒的细胞一样，在37℃和40℃时产生并释放相当数量的病毒颗粒。然而，在40℃感染RacH病毒的细胞中，细胞外颗粒的释放受到超过90%的抑制，并且相对较少的颗粒被包膜。（iv）对BALB/cA小鼠的感染显示，与亲本RacH病毒不同，HIR6-1和HIR6-2病毒在体重减轻、EHV-1感染临床症状的发展、肺中的病毒滴度以及引起病毒血症的能力等标准方面与野生型RacL11株一样具有毒力。这些发现以及我们最近研究的结果表明，IR6蛋白是EHV-1毒力的主要决定因素，并且IR6蛋白可能在病毒成熟和/或释放中发挥作用。
