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Characterization of phi 6 mutants that are temperature sensitive in the morphogenetic protein P12
Insights
The bacteriophage phi 6 P12 protein is essential for viral membrane envelopment. Mutants lacking functional P12 form unenveloped nucleocapsids, indicating its critical role in viral assembly.
Area of Science:
- Virology
- Molecular Biology
- Bacteriophage Research
Background:
- The bacteriophage phi 6 P12 protein is crucial for the envelopment of the nucleocapsid with the viral membrane.
- Gene 12, encoding P12, is located on the smallest chromosome (C) of the phi 6 virion.
Purpose of the Study:
- To investigate the function of the P12 morphogenetic protein in bacteriophage phi 6 assembly.
- To characterize temperature-sensitive (ts) mutants affecting P12 function and their impact on viral morphogenesis.
Main Methods:
- Isoelectric focusing was used to characterize phi 6 proteins.
- Screening of phi 6 ts mutants assigned to chromosome C identified mutations affecting P12.
- Morphological analysis of nucleocapsids formed at restrictive and permissive temperatures.
Main Results:
- Three independently isolated P12 ts mutants were identified, exhibiting similar phenotypes to gene 12 nonsense mutants.
- At restrictive temperatures, these mutants produced only unenveloped nucleocapsids.
- Shifting labeled cells from restrictive to permissive temperatures resulted in normal phage formation, but hydrophobic membrane proteins remained unlabeled, suggesting P12 is required during their synthesis.
Conclusions:
- Functional P12 protein is necessary for the proper assembly and envelopment of bacteriophage phi 6 virions.
- The P12 protein's role is critical during the synthesis or integration of hydrophobic membrane proteins into the viral structure.
Abstract:
P12 is a morphogenetic protein necessary for the envelopment of the bacteriophage phi 6 nucleocapsid with the viral membrane. Gene 12 is located along with three other genes on the smallest chromosome of the virion. ts mutants in P12 were obtained by first characterizing the isoelectric focusing behavior of phi 6 proteins and then screening ts mutants of phi 6 that had previously been assigned to chromosome C for changes in the behavior of P12. In this manner, three independently isolated mutants were identified and were found to have morphogenetic consequences at restrictive temperatures similar to gene 12 nonsense mutants in nonsuppressor cells in that only unenveloped nucleocapsids were formed. When infected cells were labeled at restrictive temperature, 27 degrees, and then shifted to 21 degrees, normal phage particles were formed; however, the hydrophobic membrane proteins in the particles were not labeled, indicating that functional P12 must be present at the time of synthesis of the membrane proteins for them to assemble into virions or that the defective P12 leads the membrane proteins into a nonfunctional pathway.