Maintenance and Regulation of mRNA Stability of the Saccharomyces cerevisiae OLE1 Gene Requires Multiple Elements within the Transcript That Act through Translation-independent Mechanisms

Vemula, Muralikrishna; Kandasamy, Pitchaimani; Oh, Chan-Seok; Chellappa, Ramesh; González, Caleb; Martin, Charles E.

doi:10.1074/jbc.m308812200

Cited by 22 publications

(18 citation statements)

References 36 publications

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Section: Discussionmentioning

confidence: 99%

“…For example, the S. cerevisiae OLE1-encoded ⌬-9 fatty acid desaturase (63,108) and the mammalian FAS-encoded fatty acid synthase (109) are also regulated by this mechanism. The stability of these mRNAs is regulated by nutrient availability (63,108,109). The OLE1 transcript is rapidly degraded upon fatty acid supplementation (63,108), whereas the stability of the FAS transcript increases upon glucose supplementation (109).…”

Section: Discussionmentioning

confidence: 99%

“…The stability of these mRNAs is regulated by nutrient availability (63,108,109). The OLE1 transcript is rapidly degraded upon fatty acid supplementation (63,108), whereas the stability of the FAS transcript increases upon glucose supplementation (109).…”

Section: Discussionmentioning

confidence: 99%

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Regulation of Phospholipid Synthesis in the Yeast cki1Δ eki1Δ Mutant Defective in the Kennedy Pathway

Choi

Sreenivas

Han

et al. 2004

Journal of Biological Chemistry

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In the yeast Saccharomyces cerevisiae, the most abundant phospholipid phosphatidylcholine is synthesized by the complementary CDP-diacylglycerol and Kennedy pathways. Using a cki1⌬ eki1⌬ mutant defective in choline kinase and ethanolamine kinase, we examined the consequences of a block in the Kennedy pathway on the regulation of phosphatidylcholine synthesis by the CDP-diacylglycerol pathway. The cki1⌬ eki1⌬ mutant exhibited increases in the synthesis of phosphatidylserine, phosphatidylethanolamine, and phosphatidylcholine via the CDP-diacylglycerol pathway. The increase in phospholipid synthesis correlated with increased activity levels of the CDPdiacylglycerol pathway enzymes phosphatidylserine synthase, phosphatidylserine decarboxylase, phosphatidylethanolamine methyltransferase, and phospholipid methyltransferase. However, other enzyme activities, including phosphatidylinositol synthase and phosphatidate phosphatase, were not affected in the cki1⌬ eki1⌬ mutant. For phosphatidylserine synthase, the enzyme catalyzing the committed step in the pathway, activity was regulated by increases in the levels of mRNA and protein. Decay analysis of CHO1 mRNA indicated that a dramatic increase in transcript stability was a major component responsible for the elevated level of phosphatidylserine synthase. These results revealed a novel mechanism that controls phospholipid synthesis in yeast.PC 1 is the most abundant phospholipid in the membranes of eukaryotic cells (1-4). It serves as a structural component of the membrane and as a source of bioactive lipids (e.g. lyso-PC, PA, DAG) (1-5). The importance of PC is underscored by the fact that alterations in its metabolism are linked to apoptosis (6 -9) and oncogenic transformation (10 -12). In the model eukaryote Saccharomyces cerevisiae, PC is synthesized by complementary pathways that are common to those found in mammalian cells 2 ( Fig. 1) (1,4,(13)(14)(15)(16)(17). In the CDP-DAG pathway, PC is synthesized from CDP-DAG via the reactions catalyzed by PS synthase (18 -20), PS decarboxylase (21-23), PE methyltransferase (24, 25), and phospholipid methyltransferase (24,26). In the CDP-choline branch of the Kennedy pathway, PC is synthesized from choline via the reactions catalyzed by choline kinase (27), phosphocholine cytidylyltransferase (28), and choline phosphotransferase (29,30). Analyses of mutations in S. cerevisiae (4, 31, 32), as well as in mammalian cells (33,34) indicate that the physiological role(s) of PC synthesized by the two pathways are different.The contribution of the CDP-DAG and Kennedy pathways for PC synthesis in wild type S. cerevisiae is dependent on the exogenous supply of choline (35). When grown in the presence of choline, yeast primarily synthesizes PC via the Kennedy pathway (35). On the other hand, when cells are grown in the absence of choline, PC is primarily synthesized via the CDP-DAG pathway (35). Yet, even under this growth condition, the Kennedy pathway still contributes to the synthesis of PC (36 -41). The choline required for the Kenne...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Regulation of Phospholipid Synthesis in the Yeast cki1Δ eki1Δ Mutant Defective in the Kennedy Pathway

Choi

Sreenivas

Han

et al. 2004

Journal of Biological Chemistry

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“…2). We have shown previously that in wild type cells the half-life of OLE1 mRNA expressed under the control of the GAL1 promoter is similar to that of the native transcript, yielding a basal half-life of 10 -12 min and a regulated half-life of Ͻ2 min (10,15).…”

Section: Mga2p But Not Spt23pmentioning

confidence: 99%

Regulation of Unsaturated Fatty Acid Biosynthesis in Saccharomyces

Kandasamy

Vemula

et al. 2004

Journal of Biological Chemistry

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The Saccharomyces cerevisiae OLE1 gene encodes a membrane-bound ⌬9 fatty-acid desaturase, whose expression is regulated through transcriptional and mRNA stability controls. In wild type cells grown on fatty acid-free medium, OLE1 mRNA has a half-life of 10 ؎ 1.5 min (basal stability) that becomes highly unstable when cells are exposed to unsaturated fatty acids (regulated stability). Activation of OLE1 transcription is dependent on N-terminal fragments of two membrane proteins, Mga2p and Spt23p, that are proteolytically released from the membrane by a ubiquitin-mediated mechanism. Surprisingly, disruption of the MGA2 gene also reduces the half-life of the OLE1 transcript and abolishes fatty acid regulated instability. Disruption of its cognate, SPT23, has no effect on the half-life of the mRNA. Mga2p appears to have two distinct functions with respect to the OLE1 mRNA stability: a stabilizing effect in cells grown in fatty acid-free medium and a destabilizing function in cells that are exposed to unsaturated fatty acids. These functions are independent of OLE1 transcription and can confer basal and regulated stability on OLE1 mRNAs that are produced under the control of the unrelated GAL1 promoter. Expression of soluble, N-terminal fragments of Mga2p stabilize the transcript but do not confer fatty acid-regulated instability on the mRNA suggesting that the stabilizing functions of Mga2p do not require membrane processing and that modifications to the protein introduced during proteolysis may play a role in the destabilizing effect. An analysis of mutants that are defective in mRNA degradation indicate that the Mga2p-requiring control mechanism that regulates the fatty acid-mediated instability of the OLE1 transcript acts by activating exosomal 3 3 5-exonuclease degradation activity.Unsaturated fatty acids are formed in the yeast Saccharomyces cerevisiae by Ole1p, a membrane-bound ⌬9 fatty-acid desaturase that converts long chain saturated fatty acyl-CoA substrates into monounsaturated species (1, 2). Expression of OLE11 is regulated by nutrient fatty acids by both transcriptional and mRNA stability controls. Recently, the expression of Ole1p was found to be dependent on two homologous ER membrane proteins, Mga2p and Spt23p (3). These are cleaved from the membrane by a novel ubiquitin-mediated proteolytic mechanism that releases soluble N-terminal polypeptides (4). Although neither protein appears to have a DNA binding domain, the N-terminal fragments specifically activate OLE1 transcription presumably by influencing chromatin structure or by associating with DNA binding proteins that are bound to elements on the OLE1 promoter (5, 6). Disruption of either SPT23 or MGA2 does not affect the growth rate or production of fatty acids under normal laboratory growth conditions (7). Disruption of both genes, however, blocks expression of Ole1p and results in a synthetic auxotrophy that can be overcome by adding unsaturated fatty acids to the medium or expressing OLE1 under a different promoter (4). Evidence for the direc...

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“…Stabilizing elements reside in the ORF of the yeast OLE1 mRNA (42), and destabilizing elements map to the ORFs of mammalian PAI-2 and MnSOD mRNAs (14,40). The CRDs in PAI-2 and MnSOD mRNAs do not require ribosome transit and function when inserted into a 3Ј UTR.…”

Section: Discussionmentioning

confidence: 99%

Drosophila Maternal Hsp83 mRNA Destabilization Is Directed by Multiple SMAUG Recognition Elements in the Open Reading Frame

Semotok

Luo

Cooperstock

et al. 2008

Molecular and Cellular Biology

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SMAUG (SMG) is an RNA-binding protein that functions as a key component of a transcript degradationpathway that eliminates maternal mRNAs in the bulk cytoplasm of activated Drosophila melanogaster eggs. We previously showed that SMG destabilizes maternal Hsp83 mRNA by recruiting the CCR4-NOT deadenylase to trigger decay; however, the cis-acting elements through which this was accomplished were unknown. Here we show that Hsp83 transcript degradation is regulated by a major element, the Hsp83 mRNA instability element (HIE), which maps to a 615-nucleotide region of the open reading frame (ORF). The HIE is sufficient for association of a transgenic mRNA with SMG protein as well as for SMG-dependent destabilization. Although the Hsp83 mRNA is translated in the early embryo, we show that translation of the mRNA is not necessary for destabilization; indeed, the HIE functions even when located in an mRNA's 3 untranslated region. The Hsp83 mRNA contains eight predicted SMG recognition elements (SREs); all map to the ORF, and six reside within the HIE. Mutation of a single amino acid residue that is essential for SMG's interaction with SREs stabilizes endogenous Hsp83 transcripts. Furthermore, simultaneous mutation of all eight predicted SREs also results in transcript stabilization. A plausible model is that the multiple, widely distributed SREs in the ORF enable some SMG molecules to remain bound to the mRNA despite ribosome transit through any individual SRE. Thus, SMG can recruit the CCR4-NOT deadenylase to trigger Hsp83 mRNA degradation despite the fact that it is being translated.

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Maintenance and Regulation of mRNA Stability of the Saccharomyces cerevisiae OLE1 Gene Requires Multiple Elements within the Transcript That Act through Translation-independent Mechanisms

Cited by 22 publications

References 36 publications

Regulation of Phospholipid Synthesis in the Yeast cki1Δ eki1Δ Mutant Defective in the Kennedy Pathway

Regulation of Phospholipid Synthesis in the Yeast cki1Δ eki1Δ Mutant Defective in the Kennedy Pathway

Regulation of Unsaturated Fatty Acid Biosynthesis in Saccharomyces

Drosophila Maternal Hsp83 mRNA Destabilization Is Directed by Multiple SMAUG Recognition Elements in the Open Reading Frame

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