Dissecting Structural Details of Hepadnavirus Subviral Particles

乙型肝炎病毒（HBV）感染是一个全球公共卫生问题；然而，目前批准的治疗方法很少能完全治愈。乙型肝炎表面抗原（HBsAg）是乙型肝炎病毒感染的血清学标志物。它的特点是三个跨膜蛋白具有相同的c端，在n端具有可变的框内延伸。自然地，HBsAg蛋白被整合到病毒包膜中或与宿主细胞的脂质膜组装成无基因组和无衣壳的亚病毒颗粒（SVPs）。最近，通过阻断svp的组装和释放的一种新的治疗方法在快速降低慢性HBV感染患者的HBsAg水平方面显示出有希望的结果。尽管HBsAg具有潜在的临床应用价值，但目前还没有关于HBsAg的结构信息，并且已经提出了不同的svp模型。在本研究中，我们主要利用低温电子显微镜（cryo-EM）从不同的模型体系中获得svp的高分辨率结构。我们将检验中心假设，即SVPs结构组织的变化将对其分泌途径产生重要影响。在目的1中，我们将确定土拨鼠肝炎病毒（WHV） SVP的结构。土拨鼠感染WHV是研究HBV感染发病机制的重要实验模型。在Aim 2中，我们将阐明来自携带者血清和肝癌细胞系的HBV SVP的结构细节。我们将使用抗体对HBsAg的不同区域进行空间标记。最后，将根据本方案确定的所有结构进行比较，并利用肝病毒表面抗原的详细结构信息构建SVP的完整图谱。该项目的成功完成将揭示svp的新结构信息，并将加深我们对HBV颗粒形态发生的理解，从而促进目前寻找HBV感染治疗方法的努力。

Hepatitis B virus (HBV) infection is a global public health concern; however, current approval therapies rarely lead to a complete cure. Hepatitis B surface antigen (HBsAg) is the serological marker for HBV infection. It is characterized by three transmembrane proteins having the same C-termini with variable in-frame extension at the N-terminal ends. Naturally, HBsAg proteins are incorporated in the viral envelope or assembled into genome-free and capsid-free subviral particles (SVPs) with the lipid membrane derived from the host cells. Recently, a new therapeutic approach by blocking assembly and release of SVPs has shown promising results in rapidly reducing HBsAg level in chronic HBV- infected patients. Despite its potential clinically relevant applications, there is no structural information available for HBsAg and yet different models of SVPs have been proposed. In this proposal we focus on obtaining the high resolution structures of SVPs from different model systems using cryo-electron microscope (cryo-EM). We will test the central hypothesis that changes in the structural organizations of SVPs will have important implications on their secretion pathway. In Aim 1, we will determine the structures of SVP from woodchuck hepatitis virus (WHV). Woodchuck infected with WHV is a critical experimental model for studying pathogenesis of HBV infection. In Aim 2, we will elucidate the structural details of HBV SVP from carrier's sera and hepatoma cell line. We will use antibodies to spatially label different regions of HBsAg. Finally, all structures determined from this proposal will be compared and used to build a complete picture of SVP with the detailed structural information of Hepadnavirus surface antigen. Successful completion of the proposed project will reveal new structural information of SVPs and will deepen our understanding of HBV particles morphogenesis to facilitate current efforts in finding a cure for HBV infection.