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Myristylation site in Pr65gag is essential for virus particle formation by Moloney murine leukemia virus
Abstract:
It was previously reported that the gag proteins of mammalian type C retroviruses are modified by the addition of myristate to the N-terminal glycine residue. We have performed oligonucleotide-directed mutagenesis to change this glycine codon in the Moloney murine leukemia virus genome to an alanine codon and also to specifically delete the glycine codon. Upon transfection into mammalian cells, these mutant genomes direct the synthesis of gag proteins, but these proteins are not myristylated. The mutants do not form virus particles or any recognizable virus-specific structures visible in thin sections with the electron microscope. Further, the mutant gag proteins appear to remain in the cytosol, whereas the wild type is found principally in particulate fractions of the cell. The results are consistent with the theory that myristate is required for the association of the gag protein with the plasma membrane and that this association is necessary for virus assembly.
Insights
Myristoylation of Moloney murine leukemia virus gag proteins is essential for virus assembly. Modifying or deleting the N-terminal glycine prevents myristoylation, blocking virus particle formation and membrane association.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Mammalian type C retroviruses exhibit N-terminal myristoylation of gag proteins.
- This modification is thought to be crucial for viral replication and assembly.
Purpose of the Study:
- To investigate the role of N-terminal myristoylation in Moloney murine leukemia virus (MMLV) gag protein function.
- To determine if myristoylation is essential for gag protein membrane association and virus assembly.
Main Methods:
- Oligonucleotide-directed mutagenesis was used to alter the N-terminal glycine codon of the MMLV gag gene.
- Mutant MMLV genomes were transfected into mammalian cells.
- Myristoylation status, gag protein localization, and virus particle formation were analyzed.
Main Results:
- Mutant gag proteins, lacking N-terminal myristoylation, were synthesized but not modified with myristate.
- These myristoylation-deficient mutants failed to form virus particles or recognizable virus structures.
- Mutant gag proteins remained in the cytosol, unlike wild-type gag proteins found in particulate fractions.
Conclusions:
- N-terminal myristoylation of MMLV gag proteins is indispensable for virus assembly.
- Myristoylation is required for the association of gag proteins with the plasma membrane, a critical step for virus formation.