An Analysis of Host Cell Interferon-beta Expression in Response to Ad5 Infection
Faculty Mentor(s) Name(s)
Kasey Karern
Abstract
When a cell is infected by a virus there are two opposing goals; the virus tries to replicate while the cell tries to prevent this. The virus attempts to reprogram particular cell types for viral growth and spread. The cell hosting the virus has evolved defensive mechanisms, which the virus must combat. We study adenovirus serotype 5 (Ad5). This double-stranded DNA virus models the mechanisms of other similar viruses, such as human papillomaviruses (HPV) and herpesviruses. One defense mechanism of the host cell involves Retinoic Acid-Inducible Gene I (RIG-I), which detects the presence of viral RNA in the cytoplasm and initiates a type 1 interferon immunoresponse. This starts with RIG-I stimulating a pathway where another cytosolic protein, Interferon Regulatory Factor 3 (IRF3), is phosphorylated. This phosphorylated protein, pIRF3, is a transcription factor activating genes to be transcribed, such as interferon beta (IFN-B). We used RT-qPCR, to quantify mRNA levels of RIG-I and IFN-B at various time points during an adenovirus infection. Our data support the biphasic expression of IFN-B seen in other works. To complement these studies on mRNA expression levels, we are also conducting western blot. Western blotting is an important extension of this project as it quantifies the amount of mRNA being translated into functional proteins. Several viruses will be used to determine which adenovirus proteins are important for controlling this immune response. Our focus will be on the viral E4 11k protein, which is known to interact with Ddx6, a cellular protein that can help RIG-I activation.
Start Date
27-3-2024 10:10 AM
End Date
27-3-2024 10:18 AM
Location
Arts and Sciences 2-70
An Analysis of Host Cell Interferon-beta Expression in Response to Ad5 Infection
Arts and Sciences 2-70
When a cell is infected by a virus there are two opposing goals; the virus tries to replicate while the cell tries to prevent this. The virus attempts to reprogram particular cell types for viral growth and spread. The cell hosting the virus has evolved defensive mechanisms, which the virus must combat. We study adenovirus serotype 5 (Ad5). This double-stranded DNA virus models the mechanisms of other similar viruses, such as human papillomaviruses (HPV) and herpesviruses. One defense mechanism of the host cell involves Retinoic Acid-Inducible Gene I (RIG-I), which detects the presence of viral RNA in the cytoplasm and initiates a type 1 interferon immunoresponse. This starts with RIG-I stimulating a pathway where another cytosolic protein, Interferon Regulatory Factor 3 (IRF3), is phosphorylated. This phosphorylated protein, pIRF3, is a transcription factor activating genes to be transcribed, such as interferon beta (IFN-B). We used RT-qPCR, to quantify mRNA levels of RIG-I and IFN-B at various time points during an adenovirus infection. Our data support the biphasic expression of IFN-B seen in other works. To complement these studies on mRNA expression levels, we are also conducting western blot. Western blotting is an important extension of this project as it quantifies the amount of mRNA being translated into functional proteins. Several viruses will be used to determine which adenovirus proteins are important for controlling this immune response. Our focus will be on the viral E4 11k protein, which is known to interact with Ddx6, a cellular protein that can help RIG-I activation.