Entry and replication of positive-sense, RNA viruses

丙型肝炎病毒是一个重大的全球公共卫生问题，全世界约有7000万人感染。目前还没有批准的抗丙肝病毒感染的疫苗。疾病控制中心估计，仅在美国，每年就有4万多例新感染病例，而世界其他地区的新感染病例则为150万例。慢性丙型肝炎病毒感染可以通过一种有效但昂贵的抗病毒治疗治愈。然而，药物治疗并不是在世界范围内根除丙型肝炎病毒感染的可行途径，因为这样做的费用太高。此外，成功治疗感染一种病毒基因型的患者并不排除再次感染另一种病毒基因型。药物治疗方法也因大多数受影响的个人不知道自己被感染而变得复杂，许多人从事危险行为，例如静脉注射药物。简而言之，最好的长期解决办法是投入大量智力和财力资源，发现和开发一种多价疫苗，有效对抗大多数（如果不是全部的话）丙型肝炎病毒基因型。

HCV represents a major global public health problem, infecting approximately 70 million people worldwide. There is currently no approved vaccine to counter HCV infection. The Center for Disease Control estimates more than 40,000 new infections annually in the US alone with an additional 1.5 million new infections in the rest of world. Chronic HCV infection is curable by an effective, albeit expensive, antiviral therapy. Drug treatment is not, however, a feasible route to worldwide eradication of HCV infection, as the cost of doing so would be prohibitive. Moreover, successful treatment of a patient infected with one viral genotype does not preclude re-infection with another. The drug treatment approach is also complicated by the fact that most affected individuals are unaware that they are infected, and many engage in risky behaviors, such as intravenous drug use. Simply put, the best long term solution is to invest considerable intellectual and financial resources in discovery and development of a polyvalent vaccine effective against most, if not all, HCV viral genotypes. HCV enters hepatocytes through a multistep process requiring a series of host cellular factors and the viral envelope glycoproteins E1 and E2. The HCV glycoproteins mediate cell targeting, endocytosis, and membrane fusion ultimately stimulated by endosomal acidification. At least four cellular factors are critical for HCV attachment and entry: CD81, scavenger receptor class B type I (SRBI), claudin-1 (CLDN), and occludin (OCLN). CD81 is ubiquitously expressed on a variety of cell lines, indicating a role secondary to hepatocyte-specific receptor binding. CD81 translocates with the virion to tight junctions and engages with late entry factors, CLDN and OCLN, in the endosome for acidification and entry. The majority of broadly neutralizing antibodies preclude interaction between HCV E2 and the large extracellular loop (LEL) of the cellular receptor CD81. We observed that low pH enhances CD81-LEL binding to E2 and determined the crystal structure of E2/CD81-LEL. Upon binding CD81, E2 extends an internal loop along the surface of CD81. Docking of the E2/CD81-LEL complex onto a membrane embedded, full length CD81 places the loop proximal to the membrane. Liposome flotation assays demonstrate that low pH and CD81-LEL increase E2 interaction with membranes, while structure-based mutants of highly conserved, hydrophobic amino acids abolish membrane binding. These data support a model that acidification and receptor binding result in a conformation change in E2 in preparation for membrane fusion.The molecular mechanisms thereafter, for mediating cell entry and membrane fusion for HCV, remain undefined. These comprehensive structural, biochemical, and biophysical results have established a foundation to better define the functional roles of the envelope glycoproteins in HCV infection for vaccine design.