Cargo, biogenesis and functions of extracellular vesicles released during HSV-1 infection

HSV-1感染期间释放的细胞外囊泡的货物、生物发生和功能

• 批准号: 10652535
• 负责人: Maria Kalamvoki
• 金额: $ 48.66万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10652535
• 关键词: Afferent Neurons; Alzheimer' s Disease; Benign; Biogenesis; Capsid Proteins; Cell physiology; Cells; Communication; Complex; Cytosol; DNA; Data; Disease; Encephalitis; Endosomes; Epithelial Cells; Gene Expression; Gene Silencing; Glycoproteins; HIV-1; Herpes Labialis; Herpes Simplex Infections; Herpesvirus 1; Human; Immune Evasion; Immune response; Infection; Innate Immune Response; Integration Host Factors; Light; Lytic Phase; Mass Spectrum Analysis; Mediating; Mucous Membrane; Names; Natural Immunity; Neurodegenerative Disorders; Neurons; Pathogenesis; Pathway interactions; Population; Primary Infection; Production; Productivity; Proteins; Reporting; Role; Sensory Ganglia; Severities; Sorting; Stimulator of Interferon Genes; Stress; Testing; VP 16; Viral; Viral Genes; Viral Genome; Viral Proteins; Virion; Virus; Virus Replication; cell type; design; env Gene Products; extracellular vesicles; in vivo; insight; intercellular communication; mutant; particle; pathogen; response; sensor; vesicular release

Following productive, lytic infection in mucosal epithelial cells located at the portal of entry in the body, herpes simplex virus-1 (HSV-1) establishes a lifelong, silent infection in sensory neurons. The virus is occasionally reactivated due to weakened immune response or stress causing diseases that range in severity from benign cold sores to encephalitis. HSV-1 contributes to exacerbation of neurodegenerative diseases such as Alzheimer's and facilitates infection by other pathogens such as HIV-1. Extracellular vesicles (EVs) are released by all types of cells, as they constitute a major mechanism of intercellular communication, and they can influence recipient cell functions. During HSV-1 infection two populations of EVs are readily detectable. The first population is produced through a tetraspanins biogenesis pathway. These are the most abundant EVs released from HSV-1 infected cells, which are enriched in CD63 tetraspanin and carry host factors such as the STimulator of INterferon Genes (STING), a sensor of DNA in the cytosol. These CD63+ EVs can activate innate immune responses in uninfected recipient cells and suppress a subsequent HSV-1 infection. The second population of EVs is produced through the endosomal sorting complex required for the transport (ESCRT) pathway, thus we named them ESCRT+ EVs. These EVs carry ESCRT components along with selected viral tegument and envelope protein and they appear to have a proviral role. In addition to these two population of EVs, earlier studies have reported the production of capsidless particles during HSV-1 infection composed of tegument and envelope proteins but lacking viral genome. These particles are lighter than HSV-1 virions and are known as L-particles. L-particles resemble the ESCRT+ EVs in that they carry viral proteins and have a proviral effect but are denser than ESCRT+ EVs and enriched in viral proteins. We hypothesize that distinct populations of EVs are released during HSV- 1 infection with different effects on the infection. To test our hypothesis, we have formulated three Aims. In Aim 1, we propose to compare the cargo of ESCRT+ EVs and CD63+ EVs with L-particles produced during HSV-1 infection using a mass spectrometry approach. The results of this analysis will generate important information about messages communicated by infected cells. In Aim 2, we propose to determine EV biogenesis pathways involved in the production of ESCRT+ EVs, CD63+ EVs and L-particles during HSV-1 infection. For this, we will use a gene silencing approach to compromise selected EV biogenesis pathways or we will use mutant viruses to determine the impact of selected viral genes on EV biogenesis. In Aim 3, we propose to determine the type of responses that CD63+ EVs, ESCRT+ EVs and L-particles trigger in recipient cells and determine the consequences to the infection. These studies will provide important information on the biogenesis, cargo, and functions of EVs released by HSV-1 infected cells, which are essential to determine their impact on HSV-1 pathogenesis.

位于体内入口处的粘膜上皮细胞发生生产性裂解性感染后，1 型单纯疱疹病毒 (HSV-1) 会在感觉神经元中建立终生的、无声的感染。由于免疫反应减弱或引起压力的疾病（严重程度从良性唇疱疹到脑炎），病毒偶尔会重新激活。 HSV-1 会加剧阿尔茨海默病等神经退行性疾病，并促进 HIV-1 等其他病原体的感染。细胞外囊泡（EV）由所有类型的细胞释放，因为它们构成细胞间通讯的主要机制，并且可以影响受体细胞的功能。在 HSV-1 感染期间，很容易检测到两个 EV 群体。第一个群体是通过四跨膜蛋白生物发生途径产生的。这些是 HSV-1 感染细胞释放的最丰富的 EV，富含 CD63 四跨膜蛋白，并携带宿主因子，例如干扰素基因 ST 刺激器 (STING)（细胞质中的 DNA 传感器）。这些 CD63+ EV 可以激活未感染的受体细胞中的先天免疫反应，并抑制随后的 HSV-1 感染。第二组 EV 是通过运输 (ESCRT) 途径所需的内体分选复合体产生的，因此我们将它们命名为 ESCRT+ EV。这些 EV 携带 ESCRT 成分以及选定的病毒外皮和包膜蛋白，它们似乎具有前病毒作用。除了这两个 EV 群体之外，早期的研究还报道了 HSV-1 感染期间产生的无衣壳颗粒，由外皮和包膜蛋白组成，但缺乏病毒基因组。这些颗粒比 HSV-1 病毒颗粒轻，被称为 L 颗粒。 L-颗粒与 ESCRT+ EV 类似，因为它们携带病毒蛋白并具有前病毒作用，但比 ESCRT+ EV 密度更大，并且富含病毒蛋白。我们假设 HSV-1 感染期间释放出不同的 EV 群体，对感染产生不同的影响。为了检验我们的假设，我们制定了三个目标。在目标 1 中，我们建议使用质谱方法将 ESCRT+ EV 和 CD63+ EV 的货物与 HSV-1 感染期间产生的 L 颗粒进行比较。该分析的结果将生成有关受感染细胞传达的消息的重要信息。在目标 2 中，我们建议确定 HSV-1 感染期间参与 ESCRT+ EV、CD63+ EV 和 L 颗粒产生的 EV 生物发生途径。为此，我们将使用基因沉默方法来破坏选定的 EV 生物发生途径，或者我们将使用突变病毒来确定选定的病毒基因对 EV 生物发生的影响。在目标 3 中，我们建议确定 CD63+ EV、ESCRT+ EV 和 L 颗粒在受体细胞中触发的反应类型，并确定感染的后果。这些研究将提供有关 HSV-1 感染细胞释放的 EV 的生物发生、货物和功能的重要信息，这对于确定它们对 HSV-1 发病机制的影响至关重要。