The main goal of this project is to understand how viruses assemble and transport their genetic material (genome) into cells. When cells are infected by viruses, the cellular machinery is used to produce viral building blocks (proteins) that assemble with the genome into mature viruses that can be thought of as little machines ready to infect other cells. For microviruses, which have genomes made up of single-stranded DNA, the machines contain energy and instructions to form a tubular DNA delivery device to transport DNA from the virus to a host cells. The tube only forms when the virus is on the surface of the cell to be infected, and it falls apart once the DNA has been delivered. Deciphering the mechanisms underlying construction and function of this kind of biologically-based tubular delivery device will have applications to the fields of nanotechnology and protein engineering. The project will also have an educational impact by reaching students enrolled in a laboratory course taught by the principal investigator. Part of the proposed research will be conducted in a virology laboratory course in which students will conduct hypothesis-driven research. The educational paradigm for this course has already been proven successful, yielding two class-generated publications with undergraduates as co-authors.
The production of infectious viruses requires a complex array of macromolecular interactions. The tail-less, icosahedral bacterial virus, phiX174, transiently produces a tube by oligomerization of a DNA pilot protein H, through which DNA is delivered from the viral capsid to the host cell. After DNA delivery, the tube disassembles. This proposal seeks to address several important questions about viral assembly and infection. A combination of genetic and structural approaches will explore the structure-function relationships in the H protein to understand how the chemical make-up of this pilot protein enables it to transport DNA from virus to host, what factors contribute to making the tube the right length, and what other proteins interact with H to promote proper function. Other experiments will explore how the H protein interacts with internal scaffolding proteins within the viral capsid to promote assembly of the mature virus, and these studies will be extended by reconstituting the early viral assembly steps in vitro. Because other viruses are likely to operate the same way as phiX174, the results of this research should provide generalized insights on the protein-protein interactions and protein-DNA interactions important for assembly and infection by tail-less viruses.
