RESEARCH TRAINING IN VIRUS ENTRY STUDIES

细小病毒会导致包括人类在内的许多动物患病。B19人类细小病毒导致儿童疾病第五种疾病，以及更罕见但更严重的成人和胎儿疾病。细小病毒的感染性细胞进入途径尚未被描述。这些研究的目的是确定犬细小病毒(CPV)进入组织培养细胞的感染途径，并利用这一信息来了解宿主范围限制的机制。CPV的侵染性进入途径将通过GTP酶的显性干扰突变体的过度表达而被剖析，GTP酶在正常的内吞运输过程中调节特定的步骤。此外，阻断这些步骤的抑制性抗体将被显微注射到细胞中，并对病毒感染性的影响进行检测。CPV是最近出现的一种致病性细小病毒，它是由猫病毒的衣壳蛋白自然突变而产生的。CPV衣壳的三个表面区控制着犬的宿主范围，许多在这些区域内氨基酸发生变化的突变体失去了感染犬细胞的能力，同时仍然感染猫细胞。这些突变体的感染在病毒进入细胞结合后但在病毒DNA复制之前的某个时间点被阻止。对CPV及其宿主范围突变株在允许或限制性细胞系中的感染进入途径的比较可以解释这些衣壳区域如何在不同的宿主中发挥作用。

Parvoviruses cause disease in many animals, including humans. The B19 human parvovirus causes the childhood disease Fifth Disease, as well as rarer, but more severe diseases of adults and fetuses. The infectious cellular entry pathway has not been described for any parvovirus. The aims of these studies are to define the infectious entry pathway of canine parvovirus (CPV) into tissue culture cells, with the longer range goal of using this information to understand the mechanisms of host range restriction. The infectious entry pathway of CPV will be dissected by overexpression of dominant interfering mutants of GTPases which regulate specific steps during normal endocytic trafficking. Additionally inhibitory antibodies which block these steps will be microinjected into cells and the effect on viral infectivity assayed. CPV is a recently emerged pathogenic parvovirus that arose by natural mutation of the capsid protein of a cat virus, feline panleukopenia virus (FPV). Three surface regions on the CPV capsid control canine host range, and many mutants with amino acid changes within these regions lose the ability to infect canine cells while still infecting feline cells. Infection by these mutants is blocked during viral entry at a point after cell binding, but prior to viral DNA replication. A comparison of the infectious entry pathways of CPV and its host range mutants in permissive or restrictive cell lines may explain how these regions of the capsid function in different hosts.