The long-term goal of this project is to determine the influence of nucleotide binding and protein phosphorylation on the regulation of pathogenesis and define specific mechanisms mediating RNA replication. A host encoded, autophosphorylating, dsRNA dependent protein kinase (p68) has been identified whose activity is regulated in host plants by virus or viroid infection. The protein has biochemical and immunological similarities to proteins implicated in the replication of poliovirus and in the regulation of protein synthesis. Specific aims of this proposal are directed at determining the role of p68 in infected and healthy tissues and at studying the mechanisms by which its activity is regulated. In order to define the function and regulation of this protein, it is being purified and biochemically characterized to determine the gene sequence. Antiserum to the protein is being raised and will be used in localization studies, screening of cDNA libraries and to probe the activity of the protein. Further studies on the purified protein will include characterization of the nucleotide binding and phosphorylation stoichometries, partial sequencing, phosphoamino acid analysis and tryptic peptide mapping in the presence and absence of dsRNA and use of various nucleic acid species to define the mechanism by which dsRNA regulates the activity of p68. If time permits the gene and mRNA sequences will be obtained, yielding information on the total amino acid sequence of the protein and allowing further characterization of regulatory events at the genetic level. Viruses and viroids effectively interact with and regulate host proteins during replication and pathogenesis. The identification and characterization of host proteins directly involved in pathogenesis is important to determine fundamental mechanisms regulating host-pathogen interactions. These studies will provide a foundation to determine how pathogens regulate host proteins, what effect on disease development the interaction has, and provide insight into RNA regulatory mechanisms.***//

Project Start
Project End
Budget Start
1989-02-01
Budget End
1992-07-31
Support Year
Fiscal Year
1988
Total Cost
$180,000
Indirect Cost
Name
University of Wyoming
Department
Type
DUNS #
City
Laramie
State
WY
Country
United States
Zip Code
82071