The alphaviruses and flaviviruses are two groups of RNA containing animal viruses whose members cause hundreds of millions of cases of human illness each year. Symptoms of diseases caused by these viruses include encephalitis, arthritis, fever including hemorrhagic fever and yellow fever, and rash. Our long term goal is to understand the molecular biology of replication of these viruses and their evolution in nature, which is essential for the ultimate control of these disease agents. In the upcoming project period two areas of research are proposed. 1) The interactions that occur between alphavirus glycoproteins and nucleocapsid and between nucleocapsid protein and viral RNA during virus assembly will be characterized. During virus assembly the glycoproteins interact specifically with the nucleocapsid during budding of the virus through the plasma membrane of the infected cell, and the amino acid residues responsible for this interaction will be characterized. Approaches include site specific mutagenesis and characterization of recombinant viruses in which the nucleocapsid of one virus must interact with the envelope glycoproteins of another virus. There is also an encapsidation signal in the genomic RNA which is required for formation of nucleocapsids capsid protein. We propose a series of experiments to define the encapsidation signals of Aura and Ross River viruses. 2) The cellular receptors used by alphaviruses to enter cells will be further characterized. Alphaviruses can infect and replicate in arthropods, birds, and mammals, and the receptors used determine in part the pathology of the illness caused by the virus. The alphaviruses appear to be able to use more than one receptor and the receptor for Sindbis virus appears to be different in birds and mammals. Approaches include use of immune reagents, expression experiments, and protein isolation. Changes in the viral glycoproteins can alter the constellation of receptors that can be utilized by the virus to enter cells and lead to changes in host range, tissue tropism, and virulence, and the effects of such changes in virus glycoproteins will be further characterized.
