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Mutational analysis of the mengovirus poly(C) tract and surrounding heteropolymeric sequences
L R Martin1, G M Duke, J E Osorio
1Institute for Molecular Virology, University of Wisconsin, Madison 53706, USA. LRM@AHABS.wisc.edu
Abstract:
Previously, we described three mengovirus mutants derived from cDNA plasmids, containing shortened poly(C) tracts (C8, C12, and C13UC10), that exhibited strong attenuation for virulence in mice yet grew like wild-type virus in HeLa cells. Thirteen additional mutants hav now been constructed and characterized. Five of these differ only in poly(C) length, including one with a precise deletion of the tract. The other mutants bear deletions into the regions juxtaposing poly(C). Studies with HeLa cells confirm the essential dispensability of mengovirus's poly(C) tract but reveal a subtle, measurable correlation between poly(C) length and plaque diameter. Virulence studies with mice also revealed a strong correlation between poly(C) length and virulence. For the poly(C)-flanking mutations, the 15 bases directly 5' of the tract proved dispensable for virus viability, whereas the 20 to 30 bases 3' of poly(C) were critical for growth, thus implicating this region in the basal replication of the virus.
Insights
Mengovirus poly(C) tract length impacts virulence and plaque size in mice. Deletions near the poly(C) tract show specific regions critical for viral replication.
Area of Science:
- Virology
- Molecular Biology
- Genetics
Background:
- Mengovirus mutants with shortened poly(C) tracts showed reduced virulence in mice but normal growth in HeLa cells.
- The poly(C) tract's role in viral replication and virulence was previously suggested but not fully elucidated.
Purpose of the Study:
- To further investigate the function of the mengovirus poly(C) tract and its flanking regions.
- To characterize additional mengovirus mutants with varying poly(C) tract lengths and flanking deletions.
Main Methods:
- Construction and characterization of thirteen new mengovirus mutants.
- Analysis of poly(C) tract length, flanking deletions, plaque diameter, and viral growth in HeLa cells.
- Assessment of viral virulence in mouse models.
Main Results:
- The poly(C) tract is dispensable for mengovirus viability but influences plaque diameter and virulence.
- A correlation exists between poly(C) tract length and both plaque size and mouse virulence.
- Deletions within 15 bases upstream of the poly(C) tract did not affect viability.
- 20-30 bases downstream of the poly(C) tract are critical for viral growth, suggesting a role in basal replication.
Conclusions:
- The poly(C) tract length is a key determinant of mengovirus virulence and replication efficiency.
- Specific sequences flanking the poly(C) tract, particularly downstream, are essential for mengovirus viability and replication.