The Geballe Lab at Fred Hutchinson Cancer Research Center studies the mechanisms and evolution underlying the interactions between large DNA viruses and their host cells. Our primary efforts focus on understanding how herpesviruses overcome intrinsic cellular defense systems mediated by interferon-stimulated genes, such as protein kinase R (PKR) and MxB. The conflict between these host factors and the viral antagonists that counteract them leads to an evolutionary “arms race” in which both the host and virus undergo rapid adaptations. Comparisons of the evolutionary trajectories taken by the host and viral genes that have occurred in nature or that arise in laboratory-based experimental systems provides a powerful approach for elucidating mechanisms of viral adaptation and for understanding the risks and barriers to cross-species transmission of viruses.
Studies in an increasing number of systems demonstrate that epigenetic and translational regulatory events impact eukaryotic cellular and viral gene expression. Additional research projects in the Geballe lab aim to identify the cis-acting signals and trans-acting factors that underlie lytic and latent herpesvirus replication and to elucidate these epigenetic and posttranscriptional control mechanisms.
Herpesviruses are complex DNA viruses that infect a large proportion of humans throughout the world and cause a wide range of diseases. Human cytomegalovirus (HCMV) typically produces few if any symptoms in otherwise healthy individuals, but causes life-threatening infections in newborns, solid organ and hematopoietic stem cell transplant recipients, and other immunocompromised patients. Herpes simplex viruses cause neurological disease and skin infections that often recur throughout life. Kaposi’s sarcoma-associated herpesvirus (HSHV) causes several types of cancer that are very common in Africa but also occur in the U.S. In addition to their medical importance, herpesviruses provide useful model systems for dissecting mechanisms of regulating gene expression, host-virus interactions, and evolutionary principles.
Host cells respond to viral infections by activating antiviral pathways, including ones that are induced by interferons. For example, double-stranded RNA (dsRNA) that is produced during infection activates PKR and thereby shuts off protein synthesis. To ensure the continued protein synthesis that is necessary for replication, many viruses have evolved factors that inactivate PKR. In the case of HCMV, we discovered two genes that participate in maintaining translational capacity in cells despite the presence of dsRNA and PKR. We subsequently identified PKR antagonists encoded by several other primate and rodent CMV species. These viral genes bind directly to both dsRNA and to PKR, and both of these interactions are necessary for blocking the antiviral impact of PKR. Comparisons among these species have revealed surprising complexity and diversity in the mechanisms by which CMVs overcome PKR. Current efforts are underway to understand how these genes act, how they evolve, and how we can exploit these systems for development of a new vaccines for preventing CMV infections.
Another interferon-regulated gene with broad antiviral activity is MxB. Like PKR this gene is rapidly evolving, likely in response to viral antagonists. MxB restricts all subfamilies of human herpesviruses but its mechanism of action is not clear. Moreover, we do not know how the evolution of MxB has impacted the species-specificity of its function against herpesviruses. We are now comparing the activity of MxB from multiple primate species against human and nonhuman primate herpesviruses. In addition, we are performing experimental evolution followed by next generation sequencing to elucidate how MxB restricts herpesviruses.
After acute infections subside, herpesviruses remain in the host in a quiescent or latent state. Latency seems to be dependent to a large extent on epigenetic silencing of the viral genome, but the identity of epigenetic modifications critical for latency and how they function is unknown. To address this question, we are undertaking a comprehensive CRISPR screen of epigenetic readers, writers and erasers that are needed to maintain latency in a model of Kaposi’s sarcoma.
The regulation of viral gene expression involves controls that operate at many steps. One intriguing example is evident in the gpUL4 gene of human cytomegalovirus. Expression of gpUL4 is inhibited by an unusual translational mechanism in which repression depends on ribosomal stalling mediated by the nascent peptide product of a short upstream open reading frame. Current efforts seek to elucidate how the stalling event is regulated and what its consequence is for viral replication.