Ali Ayub, Realegeno Susan, Yang Otto O, Lewis Martha J
Division of Infectious Diseases, Department of Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095, USA.
J Virol Methods. 2009 Nov;161(2):297-304. doi: 10.1016/j.jviromet.2009.07.006. Epub 2009 Jul 28.
The HIV-1 Nef protein plays a key role in pathogenesis, as demonstrated by strong selective pressure to maintain its open reading frame, and disease attenuation when it is deleted. Among myriad cellular effects attributed to Nef, downregulation of cell surface CD4 and major histocompatibility complex class I (MHC-I) proteins are the best documented. However, few data regarding primary isolate Nef functions are available, and most studies have been performed using transient transfections to express Nef driven by a non-physiologic promoter. A novel assay system to measure simultaneously the downregulation of CD4 and MHC-I by primary HIV-1 nef in a more physiologic viral genomic context is presented. Examination of plasma nef mixtures allowed comprehensive profiling of these Nef functions within the quasispecies in vivo. Subsets within the circulating nef population were observed that are either fully functional or non-functional. These data demonstrated that this assay system allows rapid characterization of bulk and clonal Nef functional profiles that can be used in pathogenesis studies to define further its important role in pathogenesis.
HIV-1 Nef蛋白在发病机制中起关键作用，这一点已通过维持其开放阅读框的强大选择压力以及缺失该蛋白时疾病的减轻得到证明。在归因于Nef的众多细胞效应中，细胞表面CD4和主要组织相容性复合体I类（MHC-I）蛋白的下调是记录最充分的。然而，关于原代分离株Nef功能的数据很少，并且大多数研究是使用瞬时转染来表达由非生理性启动子驱动的Nef进行的。本文介绍了一种新的检测系统，该系统能够在更生理性的病毒基因组背景下同时测量原代HIV-1 nef对CD4和MHC-I的下调作用。对血浆nef混合物的检测能够在体内对准种内的这些Nef功能进行全面分析。观察到循环nef群体中的亚群要么具有完全功能，要么没有功能。这些数据表明，该检测系统能够快速表征大量和克隆的Nef功能谱，可用于发病机制研究，以进一步确定其在发病机制中的重要作用。
