Host Genetic Factors to Combat Lassa Hemorrhagic Fever

描述（由申请人提供）：该基金的重点是沙粒病毒的发病机制（病毒如何引起疾病），以及为什么在感染过程中，一些宿主会严重患病并死亡，而另一些宿主则相对毫发无损。从淋巴细胞性脉络丛脑膜炎病毒（LCMV）中吸取的经验教训适用于拉沙热病毒（LASV）。 LASV每年感染30多万人，导致2万多人死亡。LASV已经被旅行者从西非运到美国。其作为生物战武器的潜力令全国关切。LASV被列入A类微生物菌剂清单。我的实验室鉴定出α-肌营养不良蛋白聚糖（<$-DG）作为LCMV和LASV的宿主细胞受体，并证明<$-DG优先定位于树突状细胞（DC）上。 然后，我们表明，糖基转移酶LARGE对<$-DG的翻译后修饰对于<$-DG作为LASV和LCMV结合、进入和复制的受体的功能成熟是绝对关键的。P. Sabeti及其同事发现LARGE突变在LASV感染流行的尼日利亚和塞拉利昂的西非人群中很常见，但在LASV不流行的西非人群中不存在。LARGE等位基因的多态性存在于约35%的LASV流行人群中，其中约45%的个体为杂合型，16%为纯合型。LARGE突变见于LASV存活者。由于高病毒血症（> 9 log空斑形成单位[PFU/ml血清]）通常与严重LASV感染和死亡的体征相关，而较低的病毒血症（< 8 log PFU/ml或更低）与恢复相关，并且<$-DG/LARGE复合物控制LASV进入和随后的复制，因此LARGE中的突变很可能通过有利于宿主对抗该病毒的存活而进化。限制产生的病毒量为宿主的免疫系统提供了一个机会窗口，以产生有效的抗LASV免疫应答，从而预防严重疾病并促进病毒清除。该基金的目的是确定LARGE突变是否是LASV感染后存活的原因。我们将在功能性LARGE酶试验中测定含有LARGE突变的LASV感染容许细胞（DC/单核细胞），并使用LASV Gp/LCMV重组病毒测定病毒结合、进入和复制。该重组病毒可用于BSL/2实验室，并已显示在允许的人和小鼠细胞中结合、进入和复制。此后，将在CDC用wtLASV检测来自具有LARGE突变并显示LASV结合、进入或复制或酶活性改变的个体的DC和单核细胞。这些研究的成果提供了机会，以确定力量塑造人类进化的个体拥有大突变等位基因可能选择抗LASV。 此外，这些研究还可以确定替代标记物，以分类对这种病毒感染敏感或耐药的个体。

DESCRIPTION (provided by applicant): This grant focuses on arenavirus pathogenesis (how viruses cause disease) and why during infection some hosts become severely ill and die while others remain relatively unscathed. Lessons learned from lymphocytic choriomeningitis virus (LCMV) are applied to Lassa fever virus (LASV). LASV infects over 300,000 individuals per year resulting in over 20,000 deaths. LASV has been transported by travelers from Western Africa to the USA. Its potential as a weapon of biowarfare is a source of national concern. LASV is placed on the Category A list of microbial agents. My laboratory identified alpha-dystroglycan (¿-DG) as the host cell receptor for LCMV and LASV and demonstrated preferential localization of ¿-DG on dendritic cells (DCs). We then showed that post- translational modification of ¿-DG by the glycosyltransferase LARGE is absolutely critical for the functional maturation of ¿-DG as a receptor for LASV and LCMV binding, entry and replication. P. Sabeti and colleagues found that mutations in LARGE were common in West African populations of Nigeria and Sierra Leone where LASV infection is endemic, but absent in those populations in West Africa where LASV is not endemic. Polymorphisms of the LARGE allele are present in about 35% of the population where LASV is endemic with about 45% of individuals heterozygous and 16% homozygous for it. Mutations in LARGE are found in LASV survivors. Since high viremia (> 9 logs of plaque forming units [PFU/ml of sera]) is routinely associated with signs of severe LASV infection and death, whereas lower viremia (< 8 logs of PFU/ml or less) with recovery, and the ¿-DG/LARGE complex controls LASV entry and subsequent replication, it is likely that mutations in LARGE shape evolution by favoring survival of the host against this virus. Limiting the amount of virus made provides the host's immune system a window of opportunity to mount an effective anti-LASV immune response thereby preventing severe disease and promoting virus clearance. This grant's purpose is to determine if LARGE mutations are responsible for survival to LASV infection. We will assay cells permissive to LASV infection (DCs/monocytes) that contain LARGE mutations in a functional LARGE enzyme assay, and for virus binding, entry and replication using a LASV Gp/LCMV recombinant virus. The recombinant virus can be used in BSL/2 laboratory and has been shown to bind, enter and replicate in permissive human and mouse cells. Thereafter those DCs and monocytes from individuals having LARGE mutations and displaying altered LASV binding, entry or replication or enzymatic activity will be tested at CDC with wtLASV. The fruits of these studies provide the opportunity to identify forces shaping human evolution in that individuals possessing LARGE mutant alleles are likely selected for resistance to LASV. Further, these studies may also identify surrogate markers to classify susceptible or resistant individuals to this viral infection.