Human endogenous retrovirus superantigen HERV-K18

描述（由申请人提供）：发现人内源性逆转录病毒（HERV）K18编码超抗原（SAg），其特征在于能够激活携带特定TCR Vp元件的T细胞。与已充分表征的细菌SAgs相比，对病毒SAgs的了解较少。HERV SAg可能是建立EB病毒（EBV）长期持续感染和1型糖尿病发病机制所必需的。K18 SAg代表TCR的独特类型的配体，不同于常规肽抗原和其他已知的SAg。这些HERV SAg通常是休眠的，并且可以被干扰素-α和EBV转录激活。与仅激活特定T细胞亚群的其他SAg相比，EBV激活的HERV-K18 SAg最初以Vp 13特异性方式刺激T细胞，但随后快速多克隆激活，其分子机制尚不清楚。细菌SAg-受体复合物的晶体结构显示每个SAg与宿主受体不同地结合。我们假设HERV SAg的相互作用不同于细菌SAg，和/或具有不同于细菌SAg的宿主受体结合动力学。该申请的长期目标是确定HERV SAg的三维结构及其与TCR和MHC分子的相互作用。这将需要相对大量的可溶性病毒SAg的可用性。然而，可溶性和功能性病毒SAgs的生产一直是一个具有挑战性的问题。因此，我们建议进行以下研究：目的1：开发一种使用GFP折叠报告系统表达可溶性HERV-K18 SAg的高效策略，其中GFP荧光提供关于融合靶蛋白折叠成功或失败的信息;目的2。HERV-K18 SAg的可溶性变体及其与TCR和II类MHC分子的复合物的功能表征和结晶。第一轮诱变已经产生了几种具有增强的荧光的K18 SAg变体，这表明溶解性改善。此R21应用旨在为后续R 01应用奠定基础。当前申请的探索/开发性质与R21机制一致。这些HERV SAgs及其与宿主受体复合物的详细结构和功能表征将有助于我们了解病毒SAgs如何执行其生物学功能，并可能揭示我们如何在不受伤的情况下与敌人睡觉。

DESCRIPTION (provided by applicant): Human endogenous retrovirus (HERV) K18 was found to encode superantigens (SAgs) which are characterized by the ability to activate T-cells bearing particular TCR Vp elements. Compared to the well characterized bacterial SAgs, viral SAgs are less understood. The HERV SAgs may be essential for establishing the long-term persistent infection of Epstein-Barr virus (EBV) and for the pathogenesis of type-l diabetes. The K18 SAg represents a unique type of ligand for TCRs, distinct from both conventional peptide antigens and other known SAgs. These HERV SAgs are normally dormant and can be transcriptionally activated by interferon-alpha and by EBV. Compared to other SAgs which only activate specific subsets of T cells, the EBV-activated HERV-K18 SAgs stimulates T cells initially in a Vp13-specific manner, but followed by a rapid polyclonal activation, for which the molecular mechanism is unknown. The crystal structures of bacterial SAgs-receptor complexes revealed that each SAg binds distinctly to the host receptors. We hypothesize that the HERV SAg interacts differently than, and/or has host receptor binding kinetics different from those of, the bacterial SAgs. The long-term objective of the application is to determine the three-dimensional structures of the HERV SAgs and their interactions with TCR and MHC molecules. This will require the availability of relatively high quantities of soluble viral SAgs. However, production of soluble and functional viral SAgs has been a challenging problem. Therefore, we propose to perform the following studies: Aim 1: Development of a highly efficient strategy to express soluble HERV-K18 SAgs using a GFP folding reporter system, in which the GFP fluorescence provides information regarding the folding success or failure of the fusion target protein; Aim 2. Functional characterization and crystallization of soluble variants of the HERV-K18 SAgs and their complexes with TCR and class II MHC molecules. A first round of mutagenesis has generated several K18 SAg variants with enhanced fluorescence, suggesting improved solubility. This R21 application is intended to lay the ground work for a subsequent R01 application. The exploratory/development nature of the current application is consistent with the R21 mechanism. A detailed structural and functional characterization of these HERV SAgs and their complexes with the host receptors will help us understand how viral SAgs perform their biological functions and may shed light on how we manage to sleep with the enemy without getting hurt.