Host Response to Viral Infection
Determining how viruses inactivate cellular signaling networks to both promote their replication and short-circuit the host response
We are actively engaged in understanding how the negative strand RNA virus vesicular stomatitis virus (VSV) takes over the protein synthesis apparatus. Our work has shown that soon after virus replication, the virus disassembles part of the protein synthesis machinery through the activation of a cellular protein called 4E-BP1 that disrupts a key initiator of protein synthesis initiation called eIF4G. The activation of 4E-BP1 comes about because VSV dominantly turns off an intracellular kinase called Akt soon after infection. By blocking the activity of this kinase, the virus causes the dephosphorylation of 4E-BP1 and brings a halt to host protein synthesis. Interestingly, while host protein synthesis is completely inhibited, virus protein synthesis continues, leaving viral proteins as the only proteins made inside the cell. We have discovered that this is due to the actions of the viral Matrix protein, and we are working to understand how the Matrix protein encourages this selective translation.
Determining the response of the circulating immune system to infection by hemorrhagic fever viruses such as Ebola, Marburg and Lassa fever viruses
A growing focus in the lab is the effort to understand how viruses induce pathogenesis. To do this we are collaborating with research laboratories at USAMRIID in Fort Detrick, MD to study the host response to Ebola and Marburg virus infection. Ebola and Marburg, both classified as hemorrhagic fever viruses, are an excellent examples of viruses that cause extreme symptoms. With fatality rates as high as 90%, it is clear that the immune system responds ineffectively to these viruses. It is becoming increasingly clear that part of the pathogenesis of these viruses is that the immune system overreacts to infection, hyperproducing cytokines and triggering coagulation deficiencies that lead to a poor outcome.
What remains poorly understood is how this unbalancing occurs. We are studying samples of circulating immune cells to understand this unbalanced response. Our approach is to analyze how these circulating immune cells respond to infection with hemorrhagic fever viruses, using microarray and deep sequencing. These approaches allow us to interrogate the entire genome to identify which genes are responding to infection and to determine whether they are increasing or decreasing. By comparing multiple samples over a time-course of infection, we can build a picture of the up- and downregulated genes that are responding virus infection. We can then analyze these genes to see which signaling pathways are being altered during infection, and we can identify proteins that serve as good markers of infection at early stages (when diagnosis is difficult).
This analysis also allows us to compare how organisms respond differently to the different viruses. Specifically we can ask how the immune systems respond to more virulent versus less virulent strains of these viruses. We can also analyze treatment studies to determine correlates of immunity and successful responses to infection.
Identifying host responses that allow infected cells to “win” the battle with an invading virus
We are interested in a rapidly deployable cell defense that we have named the antiviral granule or AVG. This is an antiviral program that is largely centered in the cytoplasm of cells, an area where most pathogenic human viruses replicate.
Our studies have revealed that when cells are infected with a defective vaccinia virus, numerous cell proteins form small, dense granules that surround the area where the virus is replicating. We have termed these granules “antiviral granules” and have shown that they must form correctly to limit virus replication. The identification of these granules suggests that they are a normal part of the cell’s response. We have also shown that it is possible to induce the formation of these granules in cells that were previously being taken over by an invading virus by adding specific small molecules to cells. This leads them to recognize and then throttle a previously active virus. This suggests that AVGs can be effectively used to limit virus replication under multiple conditions and that being able to trigger the AVG defense can help limit the spread of viral disease.
Identifying host responses that block virus infection
It has been widely recognized that viruses entering cells can trip “pathogen sensors” inside the cell that can limit or completely block virus infection. These sensors can take several forms, all of which are keyed to identify one or more unique aspects of viral infection (double stranded RNA, triphosphorylated RNA ends, DNA in the cytoplasm). Many of these sensors stimulate a transcriptional response inside infected cells that ultimately will stimulate the production of antiviral proteins and long-range signaling molecules that warn uninfected cells of danger.
We are interested in a cell defense that is complimentary to the transcription-dependent response. This response is largely centered in the cytoplasm of cells, an area where most pathogenic human viruses replicate. Our studies have revealed that when cells are infected with a defective vaccinia virus, numerous cell proteins form small, dense granules that surround the area where the virus is replicating. We have termed these granules “antiviral granules” and have shown that they must form correctly to limit virus replication. The identification of these granules suggests that they are a normal part of the cell’s response to virus infection, and raises several questions, including “how are these granules formed?” and “Is it possible to make these granules form during different types of virus infection?” These and other questions are under investigation in the laboratory.
Early Diagnostics for Viral Infection
Applying cutting-edge optics approaches to the detection of viruses.
We are actively engaged in developing label-free virus detection techniques. The ability to transcend the normal limitations of light microscopy to directly image virus particles without the need for electron microscopy has been difficult in the past. We are working with the Unlu laboratory in the Photonics center at Boston University to develop a method for directly visualizing and characterizing viruses. Our approach utilizes a controlled reflection of light to identify individual virions after they have been captured on a specially prepared surface. This allows the digital detection of virus particles, and images generated using this approach allow us to determine particle size and shape. This approach has multiple applications, including being able to rapidly characterize the morphology of viruses and virus particles present in complex solutions and providing a platform for identifying antibodies that capture intact virions. This approach can also serve as a powerful diagnostic, allowing multiplexed virus detection capable of identifying a single virus particle from blood, serum, or plasma.
This work has been highlighted in several recent publications, including Wired magazine, Forbes, USA Today , and Computerworld.