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.

项目总结 微生物的先天免疫感应导致信号转导通路的激活，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，监管的机制 人们并不完全理解这种复杂的反应。这项提案涉及一种新的转录 调节对HIV-1的先天反应的机制。我们的初步数据定义了一个基因调控网络 这映射了单核细胞来源的树突状细胞的先天免疫反应，并突出了新的转录 控制干扰素信号的电路，干扰素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，干扰素信号的失调是 发病机制和可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)是一种 干扰素和炎症的细胞型特异性调节因子。Cbfb的功能丧失突变已经被 与乳腺癌和急性髓系白血病有关，但在调节抗病毒反应方面发挥作用 干扰素信号还没有被描述。我们发现CRISPR-Cas9或RNA对Cbfb的扰动 干扰导致特异性干扰素刺激基因和炎症因子的自发诱导 髓系细胞，但不是T细胞。已知Cbfb被HIV-1 Vif劫持在T细胞中以减少其表达 APOBEC3家族限制因素，但我们的工作表明，Cbfb对HIV的影响比 之前很欣赏。有趣的是，我们的数据表明，在缺乏Cbfb的人群中，HIV-1的复制更有效 单周期感染中的髓系细胞，但病毒未能在复制能力强的检测中有效传播 (在这种情况下，效果与VIF无关)。这项提案中概述的实验将提供更大的 转录辅助因子Cbfb如何调节先天免疫反应和 影响HIV-1复制。通过这个项目，我们的目标是：1)确定Cbfb限制干扰素的机制 髓系细胞的反应，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在原代免疫细胞成熟过程中的作用。通过系统地检测突变的Cbfb 在救援实验中构建，测试Cbfb结合伙伴(如RUNX家族成员)的作用， 测试依赖于Cbfb的已知影响HIV复制的基因，我们希望能发现分子 Cbfb抑制先天免疫信号的机制及其功能失调的利用 在病毒感染期间。我们的长期目标是了解细胞参数是如何被操纵来 调整对HIV-1的先天反应，优化我们细胞的抗病毒防御。通过这个项目，我们寻求获得 对先天免疫转录调控的机械论洞察，知识可能导致新的 针对病毒库并进行抗艾滋病毒免疫反应的治疗。