Gammaherpesvirus pathogenesis: role of heparan sulfate in vivo

描述(申请人提供)：人类伽玛疱疹病毒卡波西氏肉瘤相关疱疹病毒(KSHV)是皮肤癌、卡波西氏肉瘤和某些艾滋病相关淋巴增生性疾病的病原体。MHV68是一种小鼠伽马疱疹病毒，具有与KSHV相同的生物学特性和序列同源性，为研究宿主-伽马疱疹病毒相互作用提供了一个很好的小鼠模型。与KSHV一样，MHV68严重依赖于细胞表面硫酸乙酰肝素(HS)的存在来感染其靶细胞。令人惊讶的是，虽然B细胞是体内KSHV和MHV68的主要病毒库，但这些细胞表面不表达HS。这些病毒如何感染B细胞，而这种细胞类型缺乏主要的病毒辅助受体？我们发现，在小鼠注射MHV68后的最初几个小时内，HS在B细胞上迅速上调。这种作用是由I型干扰素(干扰素-I)的作用所介导的，干扰素是一种在感染早期产生的细胞因子，是先天免疫反应的一部分。我们的发现提出了以下模型：MHV68首先与树突状细胞相互作用，树突状细胞产生干扰素-I；干扰素-I诱导B细胞表面HS上调，使其对感染具有容许性。事实上，病毒可以利用干扰素的反应，这是一个全新的、意想不到的发现。在这里，我们提出实验来验证我们的模型(AIM1)，并描述暴露于I型干扰素(AIM2)后HS上调B细胞的分子机制。发现在B细胞中诱导HS的分子机制可能会为设计旨在限制KSHV在免疫低下患者中传播的药物提供有用的信息。尽管已知许多病毒利用HS感染其靶细胞，但据我们所知，我们的工作是第一次研究HS在病毒体内致病机制中的作用，特别是证明HS在感染背景下的表达受到严格调控。重要的是，干扰素-I在大多数病毒和细菌感染的病例中产生，因此我们的发现，即HS在B细胞上上调，几乎可以肯定扩展到许多病原体。硫酸乙酰肝素能结合生长因子、细胞因子和趋化因子，影响多种生物过程。因此，我们认为B细胞上HS对干扰素-I产生的上调对这些细胞的生物学以及最终对建立强大的免疫反应的能力有重要的影响。我们的长期目标是研究我们的发现对免疫学和传染病领域的直接影响。 公共卫生相关性：卡波西肉瘤相关疱疹病毒(KSHV)是一种伽玛疱疹病毒，可导致人类癌症。这种病毒不会感染小鼠，也不会有效地感染培养中的细胞。小鼠伽马疱疹病毒68(MHV68)是一种密切相关的病毒，在细胞培养系统和小鼠体内都被用作研究伽马疱疹病毒致病机制的模型。像许多病毒一样，KHSV和MHV68使用硫酸乙酰肝素(HS)作为辅助受体进入它们的靶细胞。然而，虽然B细胞是体内感染的主要细胞类型，但我们观察到这些细胞在其表面通常不表达HS。我们工作的目的是了解这些病毒如何感染B细胞，而这种细胞类型缺乏病毒进入所需的主要分子。在我们的初步研究中，我们发现MHV68发现了一种非常意想不到的方式来触发B细胞表面HS的表达。在注射MHV68后的头几个小时，小鼠产生I型干扰素，作为正常抗病毒反应的一部分。我们发现这种I型干扰素的产生触发了B细胞表面HS的表达，从而使这些细胞对感染具有潜在的容许性。在这里，我们建议进行实验来验证我们的模型，并了解控制这些事件的分子机制。

DESCRIPTION (provided by applicant): The human gammaherpesvirus Kaposi's Sarcoma associated Herpesvirus (KSHV) is the causative agent of a skin cancer, Kaposi's Sarcoma, and certain AIDS-associated lymphoproliferative disorders. MHV68 is a murine gammaherpesvirus that shares biological features and sequence homology with KSHV and provides an excellent mouse model in which host-gammaherpesvirus interactions can be studied. Like KSHV, MHV68 is critically dependent on the presence of heparan sulfate (HS) at the cell surface to infect its target cells. Surprisingly, while B cells represent the main viral reservoir for KSHV and MHV68 in vivo, these cells do not express HS at their surface. How can these viruses infect B cells, while this cell type lacks a major viral coreceptor? We have found that HS was rapidly upregulated on B cells in the first hours following MHV68 injection in mice. This effect was mediated by the action of type-I interferon (IFN-I), a cytokine produced early upon infection as part of the innate immune response. Our findings suggest the following model: MHV68 first interacts with dendritic cells, which in turn produce IFN-I; IFN-I induces an upregulation of HS at the surface of B cells, rendering them permissive to infection. The fact that a virus can take advantage of the interferon response is a completely novel and unexpected finding. Here we propose experiments to validate our model (Aim1) and characterize the molecular mechanisms of HS upregulation on B cells upon exposure to type-I interferon (Aim2). Finding the molecular mechanisms of HS induction in B cells might provide useful information for the design of drugs aimed at limiting KSHV propagation in immunocompromised patients. Although many viruses are known to use HS to infect their target cells, our work is, to our knowledge, the first study examining the role of HS in viral pathogenesis in vivo, and in particular, to show that HS expression is tightly regulated in the context of an infection. Importantly, IFN-I is produced in most cases of viral and bacterial infections and thus our findings that HS is upregulated on B cells can almost certainly be extended to numerous pathogens. Heparan sulfate binds growth factors, cytokines and chemokines, influencing numerous biological processes. Thus, we believe that the upregulation of HS on B cells upon IFN-I production has important consequences on the biology of these cells and ultimately on the ability to mount a robust immune response. Our long-term objectives are to study the direct impact of our findings on the field of immunology and infectious disease. PUBLIC HEALTH RELEVANCE: Kaposi's Sarcoma associated Herpesvirus (KSHV) is a gammaherpesvirus that causes cancers in humans. This virus does not infect mice nor does it infect cells in culture efficiently. Murine gammaherpesvirus 68 (MHV68) is a closely related virus that is used as a model to study gammaherpesvirus pathogenesis both in cell culture systems and in mice. Like many viruses, KHSV and MHV68 use heparan sulfate (HS) as a coreceptor to enter into their target cells. However, while B cells are the main type of cells infected in vivo, we observed that these cells do not normally express HS at their surface. The purpose of our work is to understand how these viruses can infect B cells while this cell type lacks a major molecule necessary for viral entry. In our preliminary study, we found that MHV68 has found a very unexpected way to trigger HS expression at the B cell surface. In the first hours following MHV68 injection, mice produce type-I interferon as part of the normal antiviral response. We found that this type-I interferon production triggers HS expression at the B cell surface, thus rendering these cells potentially permissive to the infection. Here we propose experiments to validate our model and understand the molecular mechanisms that control these events.