By Rachel M. Miller, Graduate Student (Smith)
& Rachel A. Knoener, Postdoctoral Scientist(Smith, Sherer)
The onset of the COVID-19 pandemic has been a call to action for many within the scientific community. Long-time collaborators, Lloyd Smith, professor of chemistry, and Nathan Sherer, associate professor of molecular virology and oncology with the McArdle Laboratory for Cancer Research and Institute for Molecular Virology, set out, along with their students, to contribute to the global understanding of SARS-CoV-2 by adapting the Smith group’s Hybridization Purification of RNA-protein complexes followed by Mass Spectrometry (HyPR-MS) technology to the study of SARS-CoV-2.
Rachel Knoener, a joint postdoc between the labs, developed HyPR-MS as a graduate student with Smith. HyPR-MS enables the identification of host proteins that interact with specific RNAs. These protein interactors are likely to have important roles in viral replication, making them potential drug targets. The Smith and Sherer groups will emulate the strategy devised by Nevan Krogan, professor of cellular and molecular pharmacology at the University of California San Francisco (https://www.nytimes.com/2020/03/17/science/corona
virus-treatment.html) to identify FDA approved drugs which may disrupt viral RNA and host protein interactions, providing promising candidates for a safe and readily available SARS-CoV-2 treatment.
This work required the participation of a collaborator capable of safely growing SARS-CoV-2, and infecting human cells. Yoshihiro Kawaoka, professor of pathobiological sciences, and virologist Peter Halfmann began researching SARS-CoV-2 early in the pandemic and were willing to generate and provide infected cell cultures to the Smith and Sherer groups.
Initial HyPR-MS experiments used Vero cells (African green monkey) infected with SARS-CoV-2, while the groups investigated which human lung cell lines were susceptible to infection. The pilot experiments demonstrated efficient capture of the SARS-CoV-2 RNAs and provided preliminary candidates for protein interactors. Particularly promising is an RNA-activated antiviral enzyme called OAS3 that interacts with the viral genomic RNA and a protein called SNX27 which interacts with the RNA that encodes the viral spike protein and has been implicated in regulation of receptor molecules for other viruses.
The team has identified a human lung cell line, Calu-3 2B4, which can be reliably infected with SARS-CoV-2. HyPR-MS. Analysis using these human cells is just beginning, but the team is excited about what they will find. Once a list of human proteins that interact with the SARS-CoV-2 viral RNAs is finalized, it will be made available to other research groups, and the team will begin biological studies investigating the importance of the RNA-binding host proteins in the viral replication process.
Members of the Smith Lab are pleased to have the opportunity to contribute to the growing body of knowledge on SARS-CoV-2, and for the opportunity to work with such wonderful collaborators. They are thankful to the Sherer and Kawaoka laboratories for making this exciting work possible.