Interferons (IFNs) are a large class of immunomodulatory proteins released by host cells. Type I IFNs attract much attention due to their protective role against viral infections, and they may also be used to treat cancer, inflammatory bowel diseases, and multiple sclerosis. Binding of type I IFNs to its receptor initiates a signaling cascade that induces the expression of hundreds of IFN-stimulated genes (ISGs), the majority of which are not fully characterized. A previous study from the Rice lab screened hundreds of ISGs for antiviral effects on diverse viruses. From this screen one ISG, ADAR1 (adenosine deaminases acting on RNA 1) - specifically the isoform ADAR1p150 - enhanced rather than inhibited replication of a group of clinically important positive- strand (+)RNA viruses; West Nile virus (WNV), chikungunya virus (CHIKV), Venezuelan equine encephalitis virus (VEEV), and Sindbis virus (SINV). ADAR1 catalyzes deamination of adenosine (A) to inosine (I) in double-stranded RNA (dsRNA), and also functions as a dsRNA binding protein independent of its editing activity. Our primary hypothesis is that ADAR1p150 is A-to-I editing a virus or host dsRNA, which leads to the enhancement of (+)RNA virus replication. The main goal of this proposal is to understand the mechanism by which ADAR1p150 enhances (+)RNA virus replication, and to further identify RNA targets of ADAR1p150 in the host and virus.
Aim 1 proposes to determine the mechanism by which ADAR1p150 enhances virus replication by mutating different domains of the ADAR1p150 protein - including its deaminase domain responsible for RNA editing - to determine the domains/activities of ADAR1p150 required to enhance (+)RNA virus replication. In addition to defining the critical ADAR1 activities required for enhanced virus replication, the affected virus lifecycle step(s) will be determined.
Aim 2 will further identify RNA sequences 'edited by' or 'bound to' ADAR1p150 in the host and virus by performing high-throughput RNA-sequencing in cells expressing WT or mutant ADAR1p150. Currently there is no specific treatment available for SINV, WNV, CHIKV, and VEEV. Moreover, ADAR1 is a pro-viral ISG for human immunodeficiency virus (HIV), and an antiviral ISG for hepatitis C virus (HCV), two global pandemics that cause numerous human deaths. Thus, efforts to understand the mechanism and targets of ADAR1 may lead to new therapeutics to combat these various viral infections. Importantly, ADAR1 deletion in mice causes rapid apoptosis, defective hematopoiesis, and a global up-regulation in IFN signaling, which lead to embryonic lethality. However, we still do not have a clear picture of ADAR1 mechanism of action and its specific RNA targets in the mammalian host. Therefore, results gained from examining ADAR1 mechanism and targets in uninfected and virally infected cells will serve as a valuable resource to understand ADAR's roles in host biology.
Here we propose to understand the mechanism by which ADAR, an essential RNA editing protein to the host, enhances the replication of a group of positive-strand (+) RNA viruses - including Sindbis virus, West Nile virus, chikungunya virus, and Venezuelan equine encephalitis. Currently there is no specific treatment available for this group of viruses. Therefore, understanding ADAR mechanism of action and the identification of functionally significant RNA targets may lead to new antiviral therapeutics, and will provide us with a better understanding of ADAR functions in host biology.
| Chung, Hachung; Calis, Jorg J A; Wu, Xianfang et al. (2018) Human ADAR1 Prevents Endogenous RNA from Triggering Translational Shutdown. Cell 172:811-824.e14 |
