Browsing by keyword "*RNA, Messenger"
Now showing items 1-7 of 7
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A single amino acid substitution in yeast eIF-5A results in mRNA stabilizationMost factors known to function in mRNA turnover are not essential for cell viability. To identify essential factors, approximately 4000 temperature-sensitive yeast strains were screened for an increase in the level of the unstable CYH2 pre-mRNA. At the non-permissive temperature, five mutants exhibited decreased decay rates of the CYH2 pre-mRNA and mRNA, and the STE2, URA5 and PAB1 mRNAs. Of these, the mutant ts1159 had the most extensive phenotype. Expression of the TIF51A gene (encoding eIF-5A) complemented the temperature-sensitive growth and mRNA decay phenotypes of ts1159. The tif51A allele was rescued from these cells and shown to encode a serine to proline change within a predicted alpha-helical segment of the protein. ts1159 also exhibited an approximately 30% decrease in protein synthesis at the restrictive temperature. Measurement of amino acid incorporation in wild-type cells incubated with increasing amounts of cycloheximide demonstrated that a decrease in protein synthesis of this magnitude could not account for the full extent of the mRNA decay defects observed in ts1159. Interestingly, the ts1159 cells accumulated uncapped mRNAs at the non-permissive temperature. These results suggest that eIF-5A plays a role in mRNA turnover, perhaps acting downstream of decapping.
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Differential mRNA translation and meiotic progression require Cdc2-mediated CPEB destructionTranslational activation of several dormant mRNAs in vertebrate oocytes is mediated by cytoplasmic polyadenylation, a process controlled by the cytoplasmic polyadenylation element (CPE) and its binding protein CPEB. The translation of CPE-containing mRNAs does not occur en masse at any one time, but instead is temporally regulated. We show here that in Xenopus, partial destruction of CPEB controls the temporal translation of CPE-containing mRNAs. While some mRNAs, such as the one encoding Mos, are polyadenylated at prophase I, the polyadenylation of cyclin B1 mRNA requires the partial destruction of CPEB that occurs at metaphase I. CPEB destruction is mediated by a PEST box and Cdc2-catalyzed phosphorylation, and is essential for meiotic progression to metaphase II. CPEB destruction is also necessary for mitosis in the early embryo. These data indicate that a change in the CPEB:CPE ratio is necessary to activate mRNAs at metaphase I and drive the cells' entry into metaphase II.
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Histone messenger RNA from HeLa cells: evidence for modified 5' terminiThe distribution of [3H]methyl radioactivity in cytoplasmic histone mRNA, isolated during the DNA synthetic (S) phase of the HeLa S3 cell cycle, has been investigated. Evidence is presented that approximately 30% of the radioactivity is in m7GpppXmpYp oligonucleotides, where Xm represents 2'-O-methylated adenosine and guanosine with a molar ratio of 4:1, respectively. The remainder of the radioactivity is present as m7GpppCmpYmpZp oligonucleotides, where Xm is again 2'-O-methylated adenosine and guanosine (4:1) and where ym represents 2'-O-methylated adenosine, guanosine, cytidine, and uridine with ratios of 2:1:1:1, respectively. While 48.6% of the [3H]methyl radioactivity was present as N6-methyladenosine in poly(adenylic acid)-terminated mRNA from S-phase cells, no evidence for N6-methyladenosine was found in histone mRNA. It thus appears that histone mRNA which lacks 3'-terminal poly(adenylic acid) sequences and functions on cytoplasmic polyribosomes during a limited protion of the cell cycle is capped but lacks internal-modified nucleosides.