Stimulation of Sendai virus C' protein synthesis by cycloheximide.
The polycistronic Sendai virus P/C mRNA is translated into five proteins (P, C', C, Y1 and Y2) from distinct start sites in virus-infected cells. The translation mechanism(s) of these proteins from two overlapping open reading frames in the P/C mRNA are poorly understood [Gupta, Ono and Xu (1996) Biochemistry 35, 1223-1231]. While investigating the initiation mechanism of C' from an ACG start site, we found that C' synthesis was resistant to inhibitors of peptide chain elongation such as cycloheximide (CHX) and anisomycin, but not to pactamycin (an inhibitor of chain initiation) or puromycin (a peptide chain terminator). Moreover, low levels (less than 30 microg/ml) of CHX significantly stimulated C' synthesis. Whereas C' synthesis was stimulated, synthesis of the P and C proteins, which are translated from the same mRNA, decreased by more than 95%. Stimulation of C' synthesis by CHX is not related to its initiation at an ACG codon. Mutation of ACG to alternative start sites had no effect on the CHX-stimulated C' synthesis. Similarly, C' synthesis was preferentially stimulated when Sendai virus-infected cells were exposed to hypotonic growth medium. These results suggest that the P/C mRNA may exist in at least two reversible conformations: whereas one conformation allows synthesis of the P and C proteins, the alternative conformation allows synthesis of the C' protein. It might be that low concentrations of CHX somehow increase the alternative conformation, which increases C' synthesis. The C' protein synthesis is reminiscent of the synthesis of stress-related proteins. Perhaps Sendai virus has evolved a novel mechanism to express both non-stress-related and stress-related proteins from the same mRNA.
Gupta
K C
KC
Department of Immunology/Microbiology, Rush Medical College, Chicago, IL 60612, U.S.A.
Ono
E
E
D000818:Animals; D000841:Anisomycin / Q000494:pharmacology; D015152:Blotting, Northern; D019556:COS Cells; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D010142:Pactamycin / Q000494:pharmacology; D010458:Peptidyl Transferases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011233:Precipitin Tests; D014176:Protein Biosynthesis / Q000187:drug effects / Q000235:genetics; D011691:Puromycin / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D014162:Transfection / Q000235:genetics; D014764:Viral Proteins / Q000378:metabolism*
D003470:Culture Media; D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D014764:Viral Proteins; C051221:nonstructural C protein, Sendai virus; D010142:Pactamycin; D011691:Puromycin; D000841:Anisomycin; D003513:Cycloheximide; D010458:Peptidyl Transferases