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Selective inhibition of translation of the mRNA coding for measles virus membrane protein at elevated temperatures
Abstract:
The elevation of culture temperatures of C6 cells that were persistently infected with the Lec strain of the subacute sclerosing panencephalitis (SSPE) virus (C6/SSPE) resulted in immediate selective inhibition of membrane (M) protein synthesis. This phenomenon was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of total cytoplasmic lysates and immunoprecipitation with monoclonal antibody against the M protein in short-time labeling experiments. The synthesis of various viral mRNAs in the presence of actinomycin D decreased gradually at similar rates after a shift to 39 degrees C. No specific disappearance of the mRNA coding for the M protein was observed when viral RNAs isolated from the infected cells were compared before and after a shift up by Northern blot analysis. Results of pulse-chase experiments did not show any significant difference in M protein stability between 35 and 39 degrees C. This rapid block of M protein synthesis was observed not only in Vero cells that were lytically infected with plaque-purified clones from the Lec strain, clones isolated from C6/SSPE cells and the standard Edmonston strain of measles virus but also in CV1, MA160, and HeLa cells that were lytically infected with the Edmonston strain. Poly(A)+ RNAs that were extracted from C6/SSPE cells before and after a shift to 39 degrees C produced detectable phospho, nucleocapsid, and M proteins in cell-free translation systems at 32 degrees C. Even higher incubation temperatures did not demonstrate the selective depression of M protein synthesis described above in vitro. All these data indicate that M protein synthesis of measles virus is selectively suppressed at elevated temperatures because of an inability of the translation apparatus to interact with the M protein-encoded mRNA.
Insights
Elevated temperatures selectively inhibit measles virus membrane (M) protein synthesis in infected cells. This occurs due to a translation issue, not mRNA degradation or protein instability.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Subacute sclerosing panencephalitis (SSPE) is a persistent measles virus infection.
- Measles virus replication involves the synthesis of various viral proteins, including the membrane (M) protein.
Purpose of the Study:
- To investigate the mechanism behind the selective inhibition of measles virus M protein synthesis at elevated temperatures.
Main Methods:
- Utilized C6/SSPE cells persistently infected with the SSPE virus.
- Employed sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoprecipitation to analyze protein synthesis.
- Conducted Northern blot analysis to assess viral mRNA levels.
- Performed pulse-chase experiments to evaluate protein stability.
- Used cell-free translation systems to study mRNA-directed protein synthesis.
Main Results:
- Elevated culture temperatures (39°C) selectively inhibited M protein synthesis in C6/SSPE cells.
- Viral mRNA levels and M protein stability remained largely unaffected by temperature shifts.
- Cell-free translation experiments showed that M protein synthesis was not inhibited in vitro at elevated temperatures.
- The inhibition was observed across various cell types and measles virus strains.
Conclusions:
- Selective suppression of measles virus M protein synthesis at elevated temperatures is due to a translational defect.
- The translation apparatus is unable to efficiently interact with M protein-encoded mRNA at higher temperatures.
- This finding provides insight into the regulation of viral protein synthesis during persistent infections.