SARS CoV-2 Immune Evasion Mechanisms

我们建议表征SARS-CoV-2与人类T细胞的相互作用，并研究这种相互作用是如何影响的 适应性免疫和体液免疫。SARS-CoV-2是新冠肺炎的病原体，居世界第三位。 致病性冠状病毒自2000年以来作为人类病原体出现。冠状病毒2型失调致病 宿主炎症并可能导致严重的肺部炎症、“细胞因子风暴”等临床表现 结果。尽管有这种免疫介导的发病机制，严重新冠肺炎的患者却矛盾地 未被激活的外周血液T细胞。基于SARS CoV-1和MERS，T细胞反应是 发育迟缓，在感染期间程度相对较低。这一点，以及明确的证据表明 CoV-2有能力或再次感染并在接种疫苗的个人中引起“突破性”感染 关于冠状病毒2型S诱导记忆T细胞反应能力的问题。T细胞受体(TCR)必须相互作用 与MHC I类或II类背景中的抗原一起启动复杂的信号事件级联。跟随 TCR参与，信号导致多种结果，包括T细胞增殖，发展 效应器的功能和分化。TCR刺激后的TCR信号强度(亲和度和持续时间)为 对驱动T细胞命运至关重要，较低的TCR信号强度会导致T细胞活化、增殖减少 以及与较高的信号强度相比，长期记忆单元减少的效应器响应 互动。低TCR信号强度也促进与自身免疫相关的Th17表型 发炎。许多病毒已经进化出干扰T细胞功能的机制，有助于 复制和免疫逃避。我们的实验室研究了病毒颗粒和结构蛋白如何 在体外和体内干扰TCR信号。虽然T细胞的增殖、分化、细胞因子的释放 而最终T细胞的命运是由TCR信号强度决定的，病毒干扰在下游信号传递中的作用 还没有得到很好的研究。CD4+T细胞信号平衡也支持高亲和力B细胞的发展， 而TCR信号的失调可能导致低亲和力B细胞比例的增加，导致 降低了B细胞的保护。我们有数据表明，人类原代T细胞与CoV2颗粒孵育 Or Spike(S)蛋白失调TcR信号导致T细胞分化偏离Th1 细胞因子反应。这至少部分是由S蛋白激活ERK激酶介导的，其结果是 减少T细胞刺激后的钙流动和LAT聚集。ERK在调节命运中起着重要作用 因此，我们的发现为自体炎症细胞因子风暴提供了新的见解 在CoV2感染期间诱导。因为再感染和突破性感染导致了持续的 大流行，旨在了解SARS CoV-2失调的T细胞功能如何表现的研究 这是急需的。我们假设SARS CoV-2颗粒和Spike蛋白干扰TCR 信令和下游TCR干扰降低了T细胞反应并扭曲了T细胞 分化、细胞因子反应和记忆取决于信号干扰的部位(S)。我们 进一步表明，病毒CoV-2 T细胞调节失调有助于改变抗体谱和 免疫发病机制和逃避。我们将从三个方面阐述这一假说。首先，我们将描述 SARS冠状病毒2型颗粒和S蛋白调节的T细胞信号通路 途径研究和使用无偏见的转录组学方法。第二，我们将检测人类抗体 COVID或疫苗接种后的曲目，并将其与T细胞功能读数相关联。此外，我们还将 从变异的尖峰序列中确定免疫逃逸的风险。最后，我们将研究CoV2和 S蛋白扭曲T细胞分化，评估疾病严重程度和疫苗配方对T细胞分化的影响 感染和接种后的TCR和记忆反应。这些研究将提供新的见解 CoV2的致病机制和免疫逃避，并确定新的抗病毒靶点的潜在靶点。

We propose to characterize SARS-CoV-2 interactions with human T cells and examine how this influences adaptive and humoral immunity. SARS-CoV-2 (CoV2) is the etiologic agent of COVID-19, the third highly pathogenic coronavirus to emerge as a human pathogen since 2000. CoV-2 causes disease by dysregulating host inflammation and may lead to severe pulmonary inflammation, “cytokine storm” and other clinical outcomes. Despite this immune-mediated pathogenesis, patients with severe COVID-19 paradoxically have peripheral blood T cells that are not activated. Based on SARS CoV-1 and MERS, T cell responses are delayed in development and relatively low in magnitude during infection. This, along with clear evidence that CoV-2 is capable or reinfection and of causing “breakthrough” infection in vaccinated individuals raise questions regarding CoV-2’s ability to elicit memory T cell responses. The T cell receptor (TCR) must interact with antigens in the context of MHC class I or II to initiate a complicated cascade of signaling events. Following TCR engagement, signaling leads to a variety of outcomes including T cell proliferation, development of effector functions and differentiation. TCR signal strength (avidity and duration) following TCR stimulation is critical for driving T cell fate, and lower TCR signal strength results in reduced T cell activation, proliferation and effector responses with a reduction in long-term memory cells compared to higher signal strength interactions. Low TCR signal strength also promotes a Th17 phenotype associated with autoimmunity and inflammation. Many viruses have evolved mechanisms to interfere with T cell functions contributing to replication and immune evasion. Our laboratory has studied how virus particles and structural proteins interfere with TCR signaling in vitro and in vivo. Although T cell proliferation, differentiation, cytokine release and ultimate T cell fate are driven by TCR signal strength, the role of viral interference in downstream signaling has not been well studied. CD4+ T cell signaling equilibrium also supports development of high affinity B cells, and dysregulated TCR signaling may contribute to an increase the proportion of low-affinity B cells leading to reduced B cell protection. We have data showing that incubation of primary human T cells with CoV2 particles or spike (S) protein dysregulates TCR signaling leading to skewing of T cell differentiation away from a Th1 cytokine response. This is mediated at least in part by the S protein activating ERK kinase, with resultant reduced calcium flow and LAT aggregation following T cell stimulation. ERK is important in regulating the fate of T cell differentiation, thus our findings provide novel insights into the autoinflammatory cytokine storm induced during CoV2 infection. Since reinfection and breakthrough infections contribute to the ongoing pandemic, studies designed to understand how SARS CoV-2 dysregulates T cell function represent are critically needed. We hypothesize that SARS CoV-2 particles and spike protein interfere with TCR signaling and that downstream TCR interference reduces T cell responses and skews T cell differentiation, cytokine response, and memory depending upon the site(s) of signal interference. We further suggest that viral Cov-2 T cell dysregulation contributes to altered antibody repertoire and immune pathogenesis and evasion. We will address this hypothesis in three aims. First, we will characterize T cell signaling pathways regulated by SARS CoV-2 particles and S protein in both focused TCR signaling pathway studies and using unbiased transcriptomics approaches. Second, we will examine human antibody repertoire following COVID or vaccination and correlate this with T cell functional readouts. Further, we will determine the risk of immune evasion from variant Spike sequences. Finally, we will examine how CoV2 and S protein skew T cell differentiation and evaluate the impact of disease severity and vaccine formulation on TCR and memory responses following infection and vaccination. These studies will offer new insights into CoV2 pathogenesis and immune evasion, and identify potential targets for novel antiviral targets.