News and Publications

Assembly and Uncoating of Icosahedral Viruses

A. Schneemann, D. Marshall, N. Krishna, V. Reddy, F. Dong, J. Johnson

Coat proteins of nonenveloped, icosahedral animal viruses have multiple functions during the course of viral infection, including assembly of the viral capsid, specific encapsidation of the viral genome, binding to a cellular receptor, and uncoating. In some viruses, a single type of protein is sufficient to carry out these functions; we are interested in the chemical and structural determinants that endow a polypeptide chain with such versatility. Our investigations focus on a structurally and genetically well-characterized model system, the T = 3 nodaviruses.

Nodaviruses are assembled from 180 copies of a single coat protein and 2 strands of messenger sense RNA. To acquire infectivity, assembled particles must undergo a maturation step in which subunits of the coat protein autocatalytically cleave into 2 polypeptides. We are investigating the precise pathway of viral assembly and the mechanism of the associated cleavage reaction. Our studies are guided by the high-resolution structures of several nodaviruses that have been determined in the laboratory of J.E. Johnson, Department of Molecular Biology. Analysis of the atomic structures of these viruses has enabled us to detect regions in the coat protein that appear to be critical in regulating viral assembly, stability, and maturation. Using genetic, biochemical, and biophysical approaches, we have shown that the N-terminal part of the coat protein plays a role in regulating the shape of viral particles, whereas the C-terminal portion controls specific encapsidation of the viral genome. Currently, we are elucidating the mechanism by which the coat protein selects the 2 genomic RNAs for encapsidation into a single virion.

A second area of research focuses on viral uncoating. Uncoating and delivery of viral genomes into the cytosol of susceptible cells remain poorly understood events. The viral nucleic acid must cross a cellular membrane at some point during entry, but the mechanistic details of this step remain unknown. The nodaviruses contain a preformed helical bundle just below the surface of the particle that may form a channel within the endosomal membrane after receptor-mediated endocytosis of the virion. This channel might be used for translocation of genomic RNA across the membrane into the cytosol. We have obtained preliminary evidence that supports this model.

The third project in our laboratory is determining and isolating the viral receptor protein. We have generated monoclonal antibodies that inhibit nodavirus infection of cultured cells; these antibodies are being used to isolate the receptor protein. Availability of the viral receptor will enable us to study the initial interactions of the virus particle with the cell surface and help us elucidate the mechanism of viral uncoating in detail.

PUBLICATIONS

Dong, X.F., Natarajan, P., Tihova, M., Johnson, J.E., Schneemann, A. Particle polymorphism caused by deletion of a peptide molecular switch in a quasi-equivalent virus. J. Virol. 7:6024, 1998.