Regulation of antiviral APOBEC3G and APOBEC3F by interferons

描述（由申请人提供）：APOBEC3G （A3G）和相关的APOBEC3F （A3F）对逆转录病毒和乙型肝炎病毒（HBV）具有广泛的抗病毒活性。然而，它们在病毒天然靶细胞（如CD4+ T淋巴细胞、巨噬细胞和原代肝细胞）中的调控作用尚未得到充分研究。我们的长期目标是阐明A3G和ASF在ifn介导的先天免疫中的作用。具体的假设是，某些人类细胞可能利用一种新的ifn介导的信号通路来上调A3G和ASF蛋白，以防御病毒。我们的假设基于以下观察结果。1)我们发现干扰素（IFN）以细胞类型依赖的方式上调A3G。IFN-a诱导巨噬细胞、原代肝细胞和肝细胞系的A3G。IFN-y也能诱导肝细胞中的A3G。这两种细胞因子都不能上调原发CD4+ T细胞中的A3G，而其他已知的ifn刺激基因（ISG），如PKR和IRF-1，在所有三种细胞类型中都被诱导。2)在巨噬细胞和肝细胞系中，IFN-a对A3G的诱导依赖于JAK激酶。虽然典型的IFN-a JAK/STAT信号通路需要STAT1和STAT2，但我们观察到IFN-a在HepSB肝细胞中诱导A3G和某些其他ISG依赖于STAT2，但不依赖于STAT1。本实验将重点确定IFN在巨噬细胞中介导A3G抗病毒活性的性质，并表征IFN在巨噬细胞和肝细胞中诱导A3G的特定分子信号通路。具体目的是：目的1)研究IFN-a诱导的A3G和A3F表达在巨噬细胞抗hiv -1活性中的作用。我们将研究A3G在IFN-a诱导下对巨噬细胞中HIV-1的抗病毒作用。实验将确定IFN诱导的A3G是否会在巨噬细胞中产生功能性抗病毒活性。目的2)进一步表征肝细胞和巨噬细胞中IFN-a诱导A3G和其他ISG的stat1独立通路。ifn - stat1独立的基因转录途径尚未被描述。这些实验将确定参与A3G和ASF转录激活的成分。虽然我们已经确定JAK激酶是诱导A3G所必需的，但我们将进一步表征其他STAT家族蛋白以及潜在的新型介质的参与。拟议的研究应提供深入了解A3G和ASF在自然免疫和对IFN治疗的反应中发挥的抗病毒作用。此外，这项工作可能会增强对ifn介导的一般信号通路的了解。

DESCRIPTION (provided by applicant): APOBEC3G (A3G) and related APOBEC3F (A3F) have broad anti-viral activity against retroviruses and hepatitis B virus (HBV). However, their regulation in viral natural target cells such CD4+ T lymphocytes, macrophages, and primary liver cells have not been well-studied. Our long-term goal is to elucidate the role of A3G and ASF in IFN-mediated innate immunity. The specific hypothesis is that certain human cells may utilize a novel IFN-mediated signaling pathway to upregulate the A3G and ASF proteins in the defense against viruses. We base the hypothesis on the following observations. 1) We showed that A3G was upregulated by interferons (IFN) in a cell-type dependent manner. IFN-a induced A3G in macrophages, primary hepatocytes and liver cell lines. IFN-y also induced A3G in liver cells. Neither cytokine could upregulate A3G in primary CD4+ T cells, while other known IFN-stimulated genes (ISG) such as PKR and IRF-1 were induced in all three cell-types. 2) In macrophages and liver cell lines, IFN-a induction of A3G was dependent upon JAK kinases. While the canonical IFN-a JAK/STAT signaling pathway requires both STAT1 and STAT2, we observed that induction of A3G and certain other ISG by IFN-a in HepSB liver cells was STAT2-dependent but STAT1-independent. The proposed experiments will focus on determining the nature of IFN-mediated A3G antiviral activity in macrophages and characterizing the specific molecular signaling pathway of IFN induction of A3G in macrophages and liver cells. The specific aims are: Aim 1) to study the role of IFN-a induced A3G and A3F expression in anti-HIV-1 activity of these proteins in macrophages. We will investigate the antiviral effect of A3G on HIV-1 in macrophages upon induction by IFN-a. The experiments will determine if IFN induction of A3G will result in functional antiviral activity in macrophages. Aim 2) to further characterize the STAT1-independent pathway of IFN-a induction of A3G and other ISG in liver cells and macrophages. IFN-a STAT1-independent pathways of gene transcription have not been described. These experiments will define components involved in the transcriptional activation of A3G and ASF. While we have determined that JAK kinases are required to induce A3G, we will further characterize the involvement of other STAT family proteins as well as potential novel mediators. The proposed research should provide insight into the antiviral role that A3G and ASF play both in natural immunity and in response to IFN treatment. Furthermore, the work would potentially enhance knowledge of general IFN-mediated signaling pathways.