Comparison of Moloney murine leukemia virus mutation rate with the fidelity of its reverse transcriptase in vitro

The role of Moloney murine leukemia virus (MoMLV) reverse transcriptase (RT) in the generation of base substitution mutations during retroviral replication was analyzed. To that effect, the in vitro fidelity of the MoMLV RT was compared to the rate of base substitution mutations occurring during the...

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Bibliographic Details
Published in:The Journal of biological chemistry 1992-12, Vol.267 (34), p.24681-24688
Main Authors: VARELA-ECHAVARRIA, A, GARVEY, N, PRESTON, B. D, DOUGHERTY, J. P
Format: Article
Language:English
Subjects:
3T3 Cells
Animals
Base Sequence
Biological and medical sciences
Blotting, Southern
Codon - genetics
DNA, Viral - genetics
DNA, Viral - isolation & purification
Fundamental and applied biological sciences. Psychology
Genetic Vectors
Kinetics
Mice
Molecular and cellular biology
Molecular genetics
Molecular Sequence Data
Moloney murine leukemia virus - enzymology
Moloney murine leukemia virus - genetics
Moloney murine leukemia virus - physiology
Mutation
Plasmids
Proviruses - enzymology
Proviruses - genetics
Proviruses - physiology
Replication
Restriction Mapping
RNA-Directed DNA Polymerase - metabolism
Transfection
Virus Replication
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Summary:The role of Moloney murine leukemia virus (MoMLV) reverse transcriptase (RT) in the generation of base substitution mutations during retroviral replication was analyzed. To that effect, the in vitro fidelity of the MoMLV RT was compared to the rate of base substitution mutations occurring during the replication of an MoMLV-based retroviral vector. Using the vector in an amber reversion assay, the base substitution mutation rate at a single locus was found to be 2 x 10(-6)/base pair in one cycle of vector virus replication. Analysis of the fidelity of the purified RT using the same template sequence revealed that, of the two mispairs (A.C and T.G) that would lead to reversion of the amber codon during replication, A.C occurs at a rate of 4.0 x 10(-6), and T.G occurs at a rate of 0.7 x 10(-4). While the rate of formation of A.C is very similar to the vector mutation rate, the rate of formation of T.G is more than 30 times higher. This discrepancy in rates suggests that there are other elements in the infected cells that contribute to the fidelity of viral replication.
ISSN:0021-9258
1083-351X
DOI:10.1016/S0021-9258(18)35818-6