Regulation and Manipulation of Innate Immunity During HIV Infection

HIV 感染期间先天免疫的调节和操纵

• 批准号: 10874020
• 负责人: Jarrod Sean Johnson
• 金额: $ 66.84万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-15 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10874020
• 关键词: APOCEC3G gene; Acute Myelocytic Leukemia; Address; Affect; Antiviral Response; Binding; Biological Assay; CBFB gene; CD4 Positive T Lymphocytes; CRISPR/Cas technology; Cell Maturation; Cells; ChIP-seq; Chromatin; Complex; Data; Dendritic Cells; Detection; Equilibrium; Family; Family member; Gene Expression; Gene Targeting; Genes; Genetic; Genetic Transcription; Genetic study; Goals; HIV; HIV Infections; HIV-1; Host Defense; Immune; Immune Targeting; Immune response; Immune signaling; Infection; Inflammation; Inflammatory; Inflammatory Response; Innate Immune Response; Innovative Therapy; Interferon Type II; Interferons; Knock-out; Knowledge; Life Cycle Stages; Macrophage; Macrophage Activation; Maps; Methods; Microbe; Modeling; Molecular; Myelogenous; Myeloid Cells; Natural Immunity; Pathogenesis; Pathway interactions; Post-Translational Modification Site; Predisposition; Process; Production; Protein Isoforms; RNA Interference; RUNX1 gene; Regulation; Ribonucleoproteins; Role; Signal Transduction; System; T-Cell Activation; T-Lymphocyte; Testing; Time; Transcriptional Activation; Transcriptional Regulation; Up-Regulation; Validation; Viral; Viral Genes; Viral reservoir; Virus; Virus Diseases; Virus Replication; Work; acute infection; adaptive immunity; cell type; cofactor; differential expression; experimental study; gene network; gene regulatory network; genome-wide; immune activation; improved; inhibitor; innate immune sensing; insight; knock-down; loss of function mutation; malignant breast neoplasm; monocyte; mutant; novel; pharmacologic; response; small hairpin RNA; tool; transcription factor; transcriptome sequencing; vif Gene Products

PROJECT SUMMARY Innate immune sensing of microbes leads to activation of signal transduction cascades that trigger multiple transcription factors to rewire gene expression for host defense. However, the mechanisms that regulate this sophisticated response are not completely understood. This proposal addresses a novel transcriptional mechanism that regulates the innate response to HIV-1. Our preliminary data define a gene regulatory network that maps the innate immune response in monocyte-derived dendritic cells, and highlight new transcriptional circuitry that controls interferon signaling, which is highly relevant for HIV infection. Type I and type III interferon are known to block acute infection of HIV. However, dysregulated interferon signaling is a hallmark of pathogenesis and can increase virus replication and spread. Based on our network predictions and experimental validations, our data indicate that Core-Binding Factor Subunit Beta (CBFb, encoded by the gene CBFB) is a cell-type specific regulator of interferon and inflammation. Loss-of-function mutations in CBFb have been associated with breast cancer and acute myeloid leukemia, but a role in regulating antiviral responses through interferon signaling has not been described. We have found that perturbation of CBFb by CRISPR-Cas9 or RNA interference leads to spontaneous induction of specific interferon stimulated genes and inflammatory factors in myeloid cells but not T cells. CBFb is known to be hijacked by HIV-1 Vif in T cells to reduce expression of APOBEC3 family restriction factors, but our work suggests that CBFb’s impact on HIV is more complex than previously appreciated. Interestingly, our data indicate that HIV-1 replicates more efficiently in CBFb-depleted myeloid cells in single-cycle infections, but the virus fails to spread efficiently in replication-competent assays (and in this case, effects are independent of Vif). The experiments outlined in this proposal will provide a greater mechanistic understanding of how the transcription cofactor CBFb regulates innate immune responses and influences HIV-1 replication. With this project we aim to: 1) Determine the mechanism by which CBFb limits IFN responses in myeloid cells, 2) Determine the direct and indirect targets of CBFb and their impact on HIV infection, and 3) Validate roles for CBFb during maturation of primary immune cells. By systematically testing mutant CBFb constructs in rescue experiments, testing the roles of CBFb binding partners (such as RUNX family members), and testing CBFb-dependent genes that are known to impact HIV replication, we expect to uncover the molecular mechanism of how CBFb suppresses innate immune signaling and how dysregulation of this function is exploited during virus infection. Our long-term goals are to understand how cellular parameters can be manipulated to tune innate responses to HIV-1 and optimize our cell’s antiviral defenses. With this project, we seek to gain mechanistic insight into the transcriptional regulation of innate immunity, knowledge that could lead to new treatments to target the virus reservoir and engage anti-HIV immune responses.

项目概要 微生物的先天免疫感应导致信号转导级联的激活，从而触发 多个转录因子重新连接基因表达以进行宿主防御。然而，调节机制 这种复杂的反应尚未完全被理解。该提案提出了一种新颖的转录 调节对 HIV-1 的先天反应的机制。我们的初步数据定义了基因调控网络 绘制了单核细胞衍生的树突状细胞的先天免疫反应，并突出了新的转录 控制干扰素信号传导的电路，与艾滋病毒感染高度相关。 I型和III型干扰素 已知可以阻止艾滋病毒的急性感染。然而，失调的干扰素信号传导是 发病机制并可以增加病毒的复制和传播。基于我们的网络预测和实验 验证，我们的数据表明核心结合因子亚基 Beta（CBFb，由基因 CBFB 编码）是 干扰素和炎症的细胞类型特异性调节剂。 CBFb 的功能丧失突变 与乳腺癌和急性髓性白血病相关，但通过调节抗病毒反应发挥作用 尚未描述干扰素信号传导。我们发现 CRISPR-Cas9 或 RNA 对 CBFb 的扰动 干扰会导致特定干扰素刺激基因和炎症因子的自发诱导 骨髓细胞，但不是 T 细胞。已知 CBFb 会被 T 细胞中的 HIV-1 Vif 劫持，从而减少 T 细胞中 APOBEC3家族限制因素，但我们的工作表明CBFb对HIV的影响比 以前赞赏。有趣的是，我们的数据表明 HIV-1 在 CBFb 耗尽的情况下复制效率更高 骨髓细胞处于单周期感染中，但病毒在复制能力测定中无法有效传播 （在这种情况下，效果与 Vif 无关）。本提案中概述的实验将提供更大的 对转录辅助因子 CBFb 如何调节先天免疫反应的机制理解 影响 HIV-1 复制。通过这个项目，我们的目标是： 1) 确定 CBFb 限制 IFN 的机制 骨髓细胞的反应，2) 确定 CBFb 的直接和间接靶点及其对 HIV 感染的影响， 3) 验证 CBFb 在初级免疫细胞成熟过程中的作用。通过系统地测试突变体 CBFb 在救援实验中构建，测试 CBFb 结合伴侣（例如 RUNX 家族成员）的作用， 并测试已知影响 HIV 复制的 CBFb 依赖性基因，我们希望揭示分子机制 CBFb 如何抑制先天免疫信号传导的机制以及如何利用该功能的失调 病毒感染期间。我们的长期目标是了解如何操纵细胞参数 调整对 HIV-1 的先天反应并优化细胞的抗病毒防御。通过这个项目，我们寻求获得 对先天免疫转录调控的机制洞察，这些知识可能会带来新的发现 针对病毒库并参与抗艾滋病毒免疫反应的治疗。