The nucleic acid-binding zinc finger protein of potato virus M is translated by internal initiation as well as by ribosomal frameshifting involving a shifty stop codon and a novel mechanism of P-site slippage.
The genes for the capsid protein CP and the nucleic acid-binding 12K protein (pr12) of potato virus M (PVM) constitute the 3' terminal gene cluster of the PVM RNA genome. Both proteins are presumably translated from a single subgenomic RNA. We have identified two translational strategies operating in pr12 gene expression. Internal initiation at the first and the second AUG codon of the pr12 coding sequence results in the synthesis of the 12K protein. In addition the protein is produced as a CP/12K transframe protein by ribosomal frameshifting. For these studies parts of the CP and pr12 coding sequences including the putative frameshift region were introduced into an internal position of the beta-glucuronidase gene. Mutational analyses in conjunction with in vitro translation experiments identified a homopolymeric string of four adenosine nucleotides which together with a 3' flanking UGA stop codon were required for efficient frameshifting. The signal AAAAUGA is the first frameshift signal with a shifty stop codon to be analyzed in the eukaryotic system. Substitution of the four consecutive adenosine nucleotides by UUUU increased the efficiency of frameshifting, while substitution by GGGG or CCCC dramatically reduced the synthesis of the transframe protein. Also, UAA and UAG could replace the opal stop codon without effect on the frameshifting event, but mutation of UGA to the sense codon UGG inhibited transframe protein formation. These findings suggest that the mechanism of ribosomal frameshifting at the PVM signal is different from the one described by the 'simultaneous slippage' model in that only the string of four adenosine nucleotides represents the slippery sequence involved in a -1 P-site slippage.
Gramstat
A
A
Max-Planck-Institut für Züchtungsforschung, Köln, Germany.
Prüfer
D
D
Rohde
W
W
D001483:Base Sequence; D002213:Capsid / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics*; D003062:Codon; D004268:DNA-Binding Proteins; D016368:Frameshift Mutation; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009029:Mosaic Viruses / Q000737:chemistry* / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D009696:Nucleic Acids / Q000378:metabolism*; D030821:Plants, Genetically Modified; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D011198:Solanum tuberosum / Q000821:virology*; D014764:Viral Proteins / Q000235:genetics*; D016335:Zinc Fingers / Q000235:genetics*
D002352:Carrier Proteins; D003062:Codon; D004268:DNA-Binding Proteins; D009696:Nucleic Acids; D012367:RNA, Viral; D014764:Viral Proteins; C089926:pr12 protein, Potato virus M