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Dewhurst Lab

URMC / Labs / Dewhurst Lab / Projects

 

Research Projects

Understanding the Interaction of HIV With Glial Cells

Combination antiretroviral therapies (cART) have transformed the health and life expectancy of persons living with HIV. However, despite the successes of cART, HIV is still capable of establishing persistent infection in long-lived T cells and in the central nervous system (CNS). HIV-1 infection of the CNS can cause neurological symptoms such as motor disturbances and cognitive impairment, referred to as HIV-associated neurocognitive disorders (HAND). Estimates of the prevalence of HAND are as high as 52%, even in the cART era. These pathologies likely arise due to a combination of cellular reservoirs of viral infection and persistent neuroinflammation. Microglia represent the primary reservoir for productive HIV-1 infection within the CNS, while astrocytes are also thought to be capable of harboring the virus in a quiescent/latent state. In collaborative studies with the Steve Goldman laboratory at UR, we are examining the interplay between HIV and glial cells (both microglia and astrocytes), using a combination of in vitro and in vivo model systems. The former include cell line models and embryonal stem cell (ESC)-derived in vitro culture systems, while the latter include a novel humanized mouse model for HIV infection, in which immunodeficient mice are stably engrafted with human microglia and astrocytes. Collectively, the goal of this work is to better understand the underlying mechanisms by which HIV causes synaptic degeneration and neurocognitive dysfunction during HAND.

Developing an Improved Live Attenuated Influenza Vaccine (LAIV)

Live Attenuated Influenza Vaccines (LAIVs) are one of the currently FDA approved vaccines to prevent seasonal Influenza infection. Mutations in the Influenza RNA dependent RNA polymerase (RdRp) have been previously identified as being responsible for the temperature sensitive (ts) and attenuated (att) phenotypes of LAIV, but the effects of these mutations on the enzymatic function and physical stability of the tripartite RdRp complex are unclear. Through a collaboration with the Baek Kim laboratory at Emory University, we have shown that purified polymerase complexes derived from the prototypical A/PR/8/34 strain of IAV (PR8), but containing the two sets of LAIV mutations (AA, Len), showed a significant decrease in transcriptional activity at higher temperatures (consistent with an in vitro ts phenotype). Moreover, the polymerase complexes containing the LAIV mutations were also structurally unstable at increasing temperatures. while the wild type complex was not. We are presently testing whether this structural and enzymatic instability may alter the balance between viral mRNA transcription and viral genomic replication in infected cells, and if this may contribute to the safety and immunogenicity of LAIV. Ultimately, this work is expected to shed light on the biochemical basis for the ts phenotype of LAIV, and offer insight into approaches to produce safer and more effective LAIVs in the future.

Elucidating the Host Cell Response to Infection by Viral Pathogens (With Emphasis on SARS-CoV-2 and Adeno-associated Virus, AAV)

We are interested in innate immune responses in respiratory cells infected by SARS-CoV-2. These collaborative studies take advantage of sophisticated gene network models developed by Juilee Thakar, as well as primary human respiratory cell culture models developed by Tom Mariani, Chris Anderson and colleagues. We are collectively working to develop and refine gene network models, to identify key genes that may regulate differential responses to viral infection, and to determine how age and sex influence those responses.

In a separate project, we are examining the innate immune response elicited by adeno-associated virus (AAV) vectors. This is important because AAV is widely used in gene therapy applications, but has also been linked to an overwhelming innate immune response in some subjects, when delivered at very high doses. This project is a collaboration with the Casey Maguire laboratory at Massachusetts General Hospital, and seeks to: (1) understand whether anti-AAV antibodies may contribute to the high levels of pro-inflammatory cytokines that can be elicited by AAV capsids, and (2) whether the innate immune response to conventional AAV is different to that of extracellular vesicle enveloped AAV. (Added Note: Casey is an alumnus of the IMV PhD program at UR).

Finally, we are also collaborating with Brandon Harvey, the Chief of the Molecular Mechanisms of Cellular Stress and Inflammation Unit at the National Institute on Drug Abuse (NIDA) to determine whether SARS-CoV-2 infection of target cells results in Endoplasmic Reticulum (ER) "Exodosis" (or the release of ER protein contents from the cell). This has been observed in other viral infections, including SARS-CoV-1, and may have important consequences (e.g., it may contribute to COVID-19 related comorbidities such as thrombosis). (Added Note: Brandon is a former graduate student at UR).