RAPID / SARS-CoV-2 host cell interactions: quantitative investigations via Scanning Helium-ion Microscopy

The SARS-COV-2 virus, the cause of the COVID19 pandemic, attacks the human body by inhalation, attachment on and entry into a target cell, followed by virus multiplication and, in the most severe cases, disruption of the immune system. This research will reveal images of viruses, while they are attacking the outside of a cell, similar to an observer flying a helicopter on a battlefield. At the same time, it will be possible to zoom-in so powerfully that the very moment of the virus entry into the cell will be captured. The understanding of this cell-infecting stage depends on documenting it multiple times and in a practical way. It is critical to have the ability to visualize the entry without the limitations of traditional microscopy techniques. Spikes on the surface of the virus play a critical role in the cell insertion step. The visualization task requires specialized tools able to identify the virus itself (approximately 1/1000 the diameter of a human hair), the spike (1/10 of the virus), and tiny features of the cell membrane (comparable to the dimension of the spike). This project employs a new microscope ideally suited to this problem, a "Helium-ion Microscope" (HeIM). The COVID19 pandemic, and its possible successors, represent an acute emergency. New discoveries and new methods of investigation provide a significant social benefit. This project will establish HeIM as a technique critical to examine the virus-cell interaction for possible future needs. Involvement of students and young scientists in the research, together with its introduction to investigators of viral diseases, provide a mechanism for this new technology to be learned and spread to the broader virology community.

The project will study the SARS-COV-2 virus-cell interaction by using the new imaging modality of Helium-ion Microscopy (HeIM) which allows identification of morphological details superior to other established techniques. In particular, the study will determine the viral density on the cell surface, as it is known that the virus preferentially targets specific cell types, like the Type II Pneumocytes of the lungs. The study will quantify the number of cell-entry and cell-exit events that occur as a function of viral density on the cell surface. In so doing, these measurements will: i) quantify viral cell-entry events, differentiating between "endocytosis" and "membrane fusion" and ii) quantify viral cell-exit events, differentiating between "free-virion" and "extracellular vesicle" release. Such data provides a basis for COVID19 cell infection modeling and for designing models of therapeutic options. The project will also focus on HeIM use for such studies of optimizing sample preparation, determining procedures for imaging clinical samples, developing bi-directional correlative microscopy with fluorescent-labelled samples which better define the virus-cell biochemistry.

This RAPID award is made by the Molecular Biophysics Program in the Division of Molecular and Cellular Biosciences.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.