A cis-acting element known to block 3' mRNA degradation enhances expression of polyA-minus mRNA in wild-type yeast cells and phenocopies a ski mutant.
mRNA lacking a 3' polyA tail is not translated efficiently in wild-type eukaryotic cells, but is translated efficiently in yeast ski mutants. This enhanced expression could be due to altered translational specificity. However, as the SKI genes are required for 3' mRNA degradation, it could be a consequence of inhibition of 3' mRNA decay. Therefore, we asked if inhibition of 3' decay of a polyA-minus mRNA in cis would allow its efficient expression in wild-type cells. Capped in vitro reporter transcripts were prepared with or without a 3' cis-acting element known to inhibit 3' degradation (oligoG) and electroporated into yeast cells. The addition of oligoG to a polyA-minus mRNA enhanced expression 30-fold in wild-type cells. This level of expression was the same as that for an oligoG-minus, polyA-minus transcript in a ski mutant. The addition of oligoG did not significantly enhance the expression of polyA-minus mRNA in a ski mutant. The oligoG-dependent increase in expression was due to an increase in initial rate of translation and an increase in the functional half-life of the mRNA, similar to the effects observed in a ski mutant. The enhanced expression of the oligoG-containing RNA did not require Pab1p. We conclude that the enhanced translation of polyA-minus RNA in a ski mutant is due to inhibition of 3' mRNA degradation. Furthermore, a polyA-minus mRNA is expressed in wild-type cells when terminated in an element known to inhibit 3' decay in cis.
Brown
J T
JT
Department of Molecular Genetics and Microbiology and The Institute for Cellular and Molecular Biology, The University of Texas at Austin, 78712-1095, USA.
Johnson
A W
AW
D020413:3' Untranslated Regions / Q000378:metabolism*; D000225:Adenine; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D006147:Guanine; D007700:Kinetics; D016296:Mutagenesis; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D009843:Oligoribonucleotides / Q000378:metabolism; D011061:Poly A / Q000378:metabolism*; D039102:Poly(A)-Binding Proteins; D016601:RNA-Binding Proteins / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins
D020413:3' Untranslated Regions; D005656:Fungal Proteins; D009687:Nuclear Proteins; D009843:Oligoribonucleotides; D039102:Poly(A)-Binding Proteins; D016601:RNA-Binding Proteins; C406671:SKI3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D011061:Poly A; D006147:Guanine; D000225:Adenine