Turnover rate of yeast PGK mRNA can be changed by specific alterations in its trailer structure

Vreken, P.; Veen, Renske M. van der; Regt, V.C.H.F. de; Maat, A.L. de; Planta, Rudi J.; Raué, Hendrik A.

doi:10.1016/0300-9084(91)90053-4

Cited by 21 publications

(33 citation statements)

References 37 publications

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“…Results of Vreken and Raue (48) also suggest a role for a 5'-3' exoribonuclease. Vreken et al (49) found that insertion of a G18 tract into the 3' UTR of yeast PGKI mRNA increases its half-life by a factor of 2 without affecting its translation. Further studies of Vreken and Raue (48) and Vreken et al (47) demonstrated that the same insertion causes the accumulation of high levels of a short fragment of the mRNA extending from the G18 sequence to the mRNA cleavage-polyadenylation site.…”

Section: Discussionmentioning

confidence: 99%

Yeast cells lacking 5'-->3' exoribonuclease 1 contain mRNA species that are poly(A) deficient and partially lack the 5' cap structure.

Hsu¹,

Stevens²

1993

Mol. Cell. Biol.

365

319

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Analysis of the slowed turnover rates of several specific mRNA species and the higher cellular levels of some of these mRNAs in Saccharomyces cerevisiae lacking 5'--3' exoribonuclease 1 (xrnl cells) has led to the finding that these yeast contain higher amounts of essentially full-length mRNAs that do not bind to oligo(dT)-cellulose. On the other hand, the length of mRNA poly(A) chains found after pulse-labeling of cells lacking the exoribonuclease, the cellular rate of synthesis of oligo(dT)-bound mRNA, and the initial rate of its deadenylation appeared quite similar to the same measurements in wild-type yeast cells. Examination of the 5' cap structure status of the poly(A)-deficient mRNAs by comparative analysis of the m7G content of poly(A)-and poly(A)+ RNA fractions of wild-type and xrnl ceUs suggested that the xrnl poly(A)-mRNA fraction is low in cap structure content. Further analysis of the 5' termini by measurements of the rate of 5'-3' exoribonuclease 1 hydrolysis of specific full-length mRNA species showed that approximately 50%o of the xrnl poly(A)-deficient mRNA species lack the cap structure. Primer extension analysis of the 5' terminus of ribosomal protein 51A (RPSIA) mRNA showed that about 30% of the poly(A)-deficient molecules of the xrnl cells are slightly shorter at the 5' end. The finding of some accumulation of poly(A)-deficient mRNA species partially lacking the cap structure together with the reduction of the rate of mRNA turnover in cells lacking the enzyme suggest a possible role for 5'--3' exoribonuclease 1 in the mRNA turnover process.5'-3' exoribonuclease 1 from Saccharomyces cerevisiae has been highly purified and characterized (40,44). In recent studies of gene cloning and sequencing (21,22), the enzyme was designated XRN1. The enzyme was found to be a 160-kDa protein acting as a processive exoribonuclease and producing 5'-mononucleotides by a 5'--3' direction of hydrolysis (40, 44). The mode of hydrolysis by XRN1 was studied in detail by using poly(A), rRNA, and oligo(A) as substrates. Capped mRNA was quite resistant to hydrolysis (39). Poly(I) was not hydrolyzed by the enzyme (44). As a step toward determining the metabolic role of XRN1, the yeast gene designated XRN1 was cloned and disrupted to determine the effect on yeast cell growth (22 protein's role in mRNA metabolism. The purification and characterization of a yeast mRNA-decapping enzyme yielding as products m7GDP and mRNA chains with 5'-phosphate termini led us to suggest that the two enzymes might be involved in mRNA turnover (41,42).In this report, we describe further analysis of mRNA species found in an XRN1 gene disruptant of S. cerevisiae. Special emphasis was placed on analyzing the structure of the 5' termini of the mRNAs to determine the status of the 5' cap structure. Essentially full-length mRNA species that are both poly(A) deficient and low in cap structure content are found in the xrnl yeast cells. The possibility that such mRNA molecules are intermediates in mRNA turnover is discussed. MATERIALS AND METHODSYea...

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Section: Discussionmentioning

confidence: 99%

Yeast cells lacking 5'-->3' exoribonuclease 1 contain mRNA species that are poly(A) deficient and partially lack the 5' cap structure.

Hsu¹,

Stevens²

1993

Mol. Cell. Biol.

365

319

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“…Well-established tools for adjusting translation efficiency in S. cerevisiae are rare, even though fundamental research is focusing on this issue (358,380). The translation efficiency of heterologous genes depends on codon usage in yeast.…”

Section: Changing Protein Cellular Levelsmentioning

confidence: 99%

Progress in Metabolic Engineering of Saccharomyces cerevisiae

Nevoigt

2008

Microbiol Mol Biol Rev

526

333

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SUMMARY The traditional use of the yeast Saccharomyces cerevisiae in alcoholic fermentation has, over time, resulted in substantial accumulated knowledge concerning genetics, physiology, and biochemistry as well as genetic engineering and fermentation technologies. S. cerevisiae has become a platform organism for developing metabolic engineering strategies, methods, and tools. The current review discusses the relevance of several engineering strategies, such as rational and inverse metabolic engineering, evolutionary engineering, and global transcription machinery engineering, in yeast strain improvement. It also summarizes existing tools for fine-tuning and regulating enzyme activities and thus metabolic pathways. Recent examples of yeast metabolic engineering for food, beverage, and industrial biotechnology (bioethanol and bulk and fine chemicals) follow. S. cerevisiae currently enjoys increasing popularity as a production organism in industrial (“white”) biotechnology due to its inherent tolerance of low pH values and high ethanol and inhibitor concentrations and its ability to grow anaerobically. Attention is paid to utilizing lignocellulosic biomass as a potential substrate.

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confidence: 99%

“…Moreover, the genetic and molecular biological advantages of yeast cells compared with mammalian cells should considerably facilitate the unraveling of the mechanism(s) of mRNA degradation (37). Studies on turnover of specific yeast mRNAs, however, have so far provided only limited information, mainly concerning cis-acting (de)stabilizing elements (19,43,62 In a previous study from our laboratory (62), we showed that the half-life of the yeast phosphoglycerate kinase (PGK1) mRNA can be increased about twofold by insertion of an 18-nucleotide (nt)-long poly(G) stretch into the 3'-UTR. In this report, we present experiments on the mechanism of degradation of PGK1 mRNA evolving from the observation that this insertion causes the accumulation of a 3-terminal fragment of the PGK1 transcript, while insertion of a similar poly(G) tract into the 5'-UTR results in the accumulation of a 5'-terminal fragment.…”

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confidence: 99%

“…1. A detailed description of the construction of dINpG and YEpR5-pG, which carry a PGKI gene containing an 18-nt-long poly(G) insertion in the 5'-UTR (dINpG) or the 3'-UTR (YEpR5-pG), has been published elsewhere (59,60,62 (62). In accordance with in vivo studies on the degradation of other, prokaryotic as well as eukaryotic mRNAs, we did not detect any products shorter than the intact transcript, indicating that as a rule, intermediates formed during mRNA degradation are extremely short-lived.…”

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The rate-limiting step in yeast PGK1 mRNA degradation is an endonucleolytic cleavage in the 3'-terminal part of the coding region.

Vreken¹,

Raué²

1992

Mol. Cell. Biol.

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Insertion of an 18-nucleotide-long poly(G) tract into the 3'-terminal untranslated region of yeast phosphoglycerate kinase (PGKI) mRNA increases its chemical half-life by about a factor of 2 (P. Vreken, R. Van der Veen, V. C. H. F. de Regt, A. L. de Maat, R. J. Planta, and H. A. Raue, Biochimie 73:729-737, 1991). In this report, we show that this insertion also causes the accumulation of a degradation intermediate extending from the poly(G) sequence down to the transcription termination site. Reverse transcription and Si nuclease mapping experiments demonstrated that this intermediate is the product of shorter-lived primary fragments resulting from endonucleolytic cleavage immediately downstream from the U residue of either of two 5'-GGUG-3' sequences present between positions 1100 and 1200 close to the 3' terminus (position 1251) of the coding sequence. Similar endonucleolytic cleavages appear to initiate degradation of wild-type PGKI mRNA. Insertion of a poly(G) tract just upstream from the AUG start codon resulted in the accumulation of a 5'-terminal degradation intermediate extending from the insertion to the 1100-1200 region. RNase H degradation in the presence of oligo(dT) demonstrated that the wild-type and mutant PGKI mRNAs are deadenylated prior to endonucleolytic cleavage and that the half-life of the poly(A) tail is three-to sixfold lower than that of the remainder of the mRNA. Thus, the endonucleolytic cleavage constitutes the rate-limiting step in degradation of both wild-type and mutant PGKI transcripts, and the resulting fragments are degraded by a 5'-*3' exonuclease, which appears to be severely retarded by a poly(G) sequence.Over the past decade, it has become evident that the pattern of gene expression in a cell is determined not only by factors controlling transcription but to a large extent also by posttranscriptional events, including the rate of degradation of the individual mRNAs. In both prokaryotic and eukaryotic cells, different mRNAs show sometimes considerable differences in stability that are of fundamental importance in establishing their relative cellular levels. Furthermore, there are numerous examples of adjustments in the half-life of particular mRNAs in response to specific stimuli that play a major role in the adaptation of the cells to changing conditions (for recent reviews, see references 1, 4, 9, 12, and 48).To gain more insight into the manner in which turnover of their transcripts contributes to the control of expression of particular genes, considerable effort is being put into the characterization of structural features within mRNA governing the rate of decay, as well as the trans-acting factors involved. As a result, a number of structural determinants of prokaryotic as well as eukaryotic mRNA stability have now been identified. Among the eukaryotic stability determinants so far characterized are the AU-rich sequences responsible for the rapid turnover of a group of mRNAs encoding lymphokines, interferons, and cellular growth factors (13,46,49,50), the stem-loop struc...

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Turnover rate of yeast PGK mRNA can be changed by specific alterations in its trailer structure

Cited by 21 publications

References 37 publications

Yeast cells lacking 5'-->3' exoribonuclease 1 contain mRNA species that are poly(A) deficient and partially lack the 5' cap structure.

Yeast cells lacking 5'-->3' exoribonuclease 1 contain mRNA species that are poly(A) deficient and partially lack the 5' cap structure.

Progress in Metabolic Engineering of Saccharomyces cerevisiae

The rate-limiting step in yeast PGK1 mRNA degradation is an endonucleolytic cleavage in the 3'-terminal part of the coding region.

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