The cardio-and aphthovirus genera of picornaviruses are distinguished among viruses by the presence of long, polypyrimidine tracts within the 5' non-coding regions of their genomes. In cardioviruses, like EMCV and Mengo, the tracts contain virtually pure cytidine sequences (C115UCUC3UC10 and C44UC10, respectively), or """"""""Poly(C)."""""""" Genetic manipulation of cDNAs has clearly shown the specific length of poly(C) is a critical determinant of Mengo pathogenicity. Wild-type cardioviruses are highly virulent and infect many species of animals, including primates, rodents and pigs. The murine LD50 is between 1-100 pfu when administered i.c. In contrast, engineered deletion of the Mengo poly(C) has produced viral strains (e.g. vMC[0]) with LD50s of 10[6]-10[9] pfu. This attenuation is accompanied by a high degree of genetic stability in tissue culture and animals. Inoculated recipients protectively seroconvert with long-lived immunity. This has allowed the poly(C) phenomenon to be genetically harnessed for the effective delivery of other heterologous epitopes engineered within the Mengo cDNAs. Live, attenuated chimeras that carry and express 1000 extra nucleotides as protein-coding sequences have been tested. HIV, SIV, and malaria determinants are excellent, potent immunogens in mice and monkeys when delivered in this manner. The mechanism of poly(C) attenuation remains enigmatic. It is proposed that wild-type viruses rely on their long poly(C)s to bind and inactivate sentinel cellular enzymes, such as dsRNA-activated protein kinase (PKR) in a manner analogous to adenovirus VAI RNAs. The short-tract viruses, unable to lure or trap PKR with the same efficiency, seem unable to avoid a consequent antiviral response by the cells, and in essence, vaccinate the host instead of killing it. In support of this hypothesis are data with PKR knockout mice that lack this essential gene, and thus are susceptible to wild-type like killing by the normally attenuated short poly(C) viruses.
The specific aims of this proposal are: (1) to test the novel """"""""mousetrap"""""""" hypothesis which predicts that wild-type cardioviruses are pathogenic because their long poly(C) tracts enable binding or inactivation of sentinel cellular enzymes and consequent avoidance of an antiviral state; (2) to examine the genetic differences between EMCV and Mengo near the poly(C) and identify all local sequences which may contribute to the attenuation phenomenon; (3) to document the genetic stability and viral persistence of short-tract Mengo in mice using forced-passage techniques and revertant analysis; (4) to examine the heterologous carrying capacity of novel Mengo constructions and chimeras, with the intent of exploitation as live, attenuated vaccine-vector delivery systems.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI030566-09
Application #
6163872
Study Section
Experimental Virology Study Section (EVR)
Program Officer
Meegan, James M
Project Start
1991-01-01
Project End
2002-02-28
Budget Start
2000-03-01
Budget End
2002-02-28
Support Year
9
Fiscal Year
2000
Total Cost
$197,237
Indirect Cost
Name
University of Wisconsin Madison
Department
Veterinary Sciences
Type
Schools of Veterinary Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Osorio, J E; Grossberg, S E; Palmenberg, A C (2000) Characterization of genetically engineered mengoviruses in mice. Viral Immunol 13:27-35
Martin, L R; Neal, Z C; McBride, M S et al. (2000) Mengovirus and encephalomyocarditis virus poly(C) tract lengths can affect virus growth in murine cell culture. J Virol 74:3074-81
Backues, K A; Hill, M; Palmenberg, A C et al. (1999) Genetically engineered Mengo virus vaccination of multiple captive wildlife species. J Wildl Dis 35:384-7
Martin, L R; Duke, G M; Osorio, J E et al. (1996) Mutational analysis of the mengovirus poly(C) tract and surrounding heteropolymeric sequences. J Virol 70:2027-31
Duque, H; Palmenberg, A C (1996) Epitope mapping of monoclonal antibodies raised to recombinant Mengo 3D polymerase. Virus Genes 13:159-68
Osorio, J E; Hubbard, G B; Soike, K F et al. (1996) Protection of non-murine mammals against encephalomyocarditis virus using a genetically engineered Mengo virus. Vaccine 14:155-61
Osorio, J E; Martin, L R; Palmenberg, A C (1996) The immunogenic and pathogenic potential of short poly(C) tract Mengo viruses. Virology 223:344-50
Martin, L R; Palmenberg, A C (1996) Tandem mengovirus 5' pseudoknots are linked to viral RNA synthesis, not poly(C)-mediated virulence. J Virol 70:8182-6
Hahn, H; Palmenberg, A C (1995) Encephalomyocarditis viruses with short poly(C) tracts are more virulent than their mengovirus counterparts. J Virol 69:2697-9
Palmenberg, A C; Osorio, J E (1994) Cardioviral poly(C) tracts and viral pathogenesis. Arch Virol Suppl 9:67-77

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