Investigation of SV5 Budding Mechanisms

描述（由申请人提供）：包膜病毒是由病毒成分在细胞膜上组装后发生的出芽过程形成的。在许多情况下，导致膜裂变和颗粒释放的后期步骤似乎是通过操纵宿主机制来完成的，宿主机制被招募到病毒组装位点。由于副粘病毒缺乏与许多其他包膜病毒（如逆转录病毒）用于宿主因子募集的相同的晚期结构域序列，导致副粘病毒颗粒释放的步骤知之甚少。最近，我们在副粘病毒副流感病毒5 （SV5）的基质(M)蛋白中发现了一个替代序列FPIV，它可以在功能上替代人类免疫缺陷病毒1型（HIV-1）的PTAP晚期结构域出芽。在此，我们提出了进一步研究副粘病毒操纵宿主机制以促进病毒出芽的实验。我们的第一个目的是通过测试SV5 M蛋白是泛素分子附着的直接靶标的可能性，来研究泛素-蛋白酶体途径在SV5出芽中的作用。我们将尝试通过去除M蛋白FPIV序列附近的赖氨酸残基来防止这种泛素化的发生。我们的第二个目标是确定对SV5出芽很重要的单个E类蛋白的子集，并将其招募到SV5组装位点。我们的第三个目标是鉴定与SV5 M蛋白结合的宿主蛋白，测试M相互作用蛋白对SV5出芽的重要性，并评估结合是否以依赖于FPIV序列的方式发生。我们的第四个目标是研究SV5 NP蛋白在病毒样颗粒出芽中的独特作用。与公共卫生的相关性：副粘病毒是影响人类和动物的一系列广泛疾病的罪魁祸首，包括麻疹和腮腺炎。更好地了解副粘病毒出芽将为未来开发针对病毒生命周期这一阶段的抗病毒药物提供基础。

DESCRIPTION (provided by applicant): Enveloped viruses are formed by a budding process that occurs following the assembly of viral components at cellular membranes. The late steps leading up to membrane fission and particle release appear to be accomplished in many cases through manipulation of host machinery that is recruited to virus assembly sites. As paramyxoviruses lack the same late domain sequences used by many other enveloped viruses, such as retroviruses, for host factor recruitment, the steps leading up to the release of paramyxovirus particles are poorly understood. Recently, we identified an alternative sequence, FPIV, within the matrix (M) protein of the paramyxovirus parainfluenza virus 5 (SV5) that can functionally substitute for the PTAP late domain of human immunodeficiency virus type 1 (HIV-1) for budding. Here, we propose experiments to further investigate mechanisms by which paramyxoviruses manipulate host machinery to facilitate virus budding. Our first aim is to investigate the role of the ubiquitin-proteasome pathway in SV5 budding by testing the possibility that SV5 M protein is a direct target for the attachment of ubiquitin molecules. We will attempt to prevent this ubiquitination from occurring through removal of lysine residues near the FPIV sequence of M protein. Our second aim is to define the subset of individual Class E proteins that is important for SV5 budding and that is recruited to SV5 assembly sites. Our third aim is to identify host proteins that bind to SV5 M protein, test the importance of M-interacting proteins for SV5 budding, and assess whether binding occurs in a way that is dependent on the FPIV sequence. Our fourth aim is to investigate the unique role of SV5 NP protein in the budding of virus-like particles. Relevance to public health: Paramyxoviruses are responsible for a wide range of diseases that affect both humans and animals, including measles and mumps. A better understanding of paramyxovirus budding will provide a foundation for future efforts towards the development of antiviral drugs that target this step of the virus lifecycle.