Interferon Gene Expression in VSV-Infected Cells

病毒感染通常触发抗病毒基因的转录，包括1型干扰素（IFN）。新产生的IFN从细胞中释放出来，并激活信号转导级联，诱导许多干扰素应答基因的转录，其产物在未感染的邻近细胞中建立抗病毒状态。许多病毒已经进化到抵消这种宿主防御机制，允许病毒复制发生。本研究的总体重点是研究宿主诱导抗病毒IFN应答的能力与水泡性口炎病毒（VSV）阻断该系统诱导的能力之间的平衡。细胞毒性VSV基质（M）蛋白抑制宿主定向转录并限制IFN基因 在VSV感染的细胞中表达。本项目的目的是确定是否M蛋白是唯一负责抑制IFN基因表达的基础上，其抑制宿主转录的能力，或者是否有一个特定的机制，VSV调节IFN基因的表达。我们的初步数据表明，后者是正确的。我们已经证明，在VSV的IFN诱导型和IFN抑制型菌株感染的细胞中，IFN基因表达的上游激活因子NF-κ B的激活受到不同的调节。我们提出第二种病毒功能抑制NF-κ B活化。因此，这种病毒成分将发挥关键作用， 诱导IFN基因表达。为了验证这一假设，要解决的具体目标是：（1）通过克隆和确定每个病毒基因的序列来表征来自VSV的IFN抑制和IFN诱导株的VSV印第安纳州Hr基因。(2)为了鉴定负责NF-κ B活化的病毒组分，我们将研究用重组VSV M缺陷株感染的细胞中的NF-κ B活化，以及确定用编码病毒基因之一的表达载体转染的细胞中NF-κ B的活化是否受到抑制。(3)为了了解病毒调节NF-κ B激活的机制，我们将确定IkappaB是否被降解，并研究其在细胞中的表达。 图1显示了在VSV感染和转染的细胞中IkappaB激酶的磷酸化状态。在完成这个项目后，我们将获得新的见解病毒调节宿主干扰素反应。详细了解这种细胞对病毒感染的反应可能有助于开发新的抗病毒和抗癌疗法。

Viral infections commonly trigger the transcription of antiviral genes, including type 1 interferon (IFN). Newly produced IFNs are released from the cell, and activate a signal transduction cascade that induces the transcription of many interferon responsive genes whose products set up an antiviral state in uninfected neighboring cells. Many viruses have evolved to counteract this host defense mechanism, allowing virus replication to occur. The overall focus of this research is to investigate the balance between the ability of the host to induce the antiviral IFN response, and the ability of vesicular stomatitis virus (VSV) to block induction of this system. The cytotoxic VSV matrix (M) protein inhibits host-directed transcription and limits IFN gene expression in VSV-infected cells. The goal of this project is to determine if M protein is solely responsible for suppression of IFN gene expression based on its ability to inhibit host transcription, or if there is a specific mechanism by which VSV regulates IFN gene expression. Our preliminary data suggest that the later is true. We've demonstrated that NF-kappaB activation, an upstream activator of IFN gene expression, is regulated differently in cells infected with an IFN-inducing and IFN-suppressing strains of VSV. We propose that a second viral function inhibits NF-kappaB activation. This viral component would therefore play a crucial role in induction of IFN gene expression. To test this hypothesis, the specific aims to be addressed are: (1) To characterize the VSV Indiana Hr genes from an IFN-suppressing and IFN-inducing strain of VSV by cloning and determining the sequence of each viral gene. (2) To identify the viral component responsible for activation of NF-kappaB, we will investigate NF-kappaB activation in cells infected with recombinant M-defective strains of VSV, as well as determining if activation of NF-kappaB is suppressed in cells transfected with an expression vector encoding one of the viral genes. (3) To understand the mechanism by which the virus regulates NF-kappaB activation, we will determine if IkappaB has been degraded and investigate the phosphorylation status of the IkappaB Kinase in VSV-infected and transfected cells. Upon completion of this project, we will gain new insights into virus modulation of the host interferon response. Detailed knowledge of this cellular response to viral infection could be useful to the development of new antiviral and anticancer therapies.