Translational Regulation of Prostaglandin Endoperoxide H Synthase-1 mRNA in Megakaryocytic MEG-01 Cells

Duquette, Maryse.; Laneuville, Odette

doi:10.1074/jbc.m207007200

Cited by 13 publications

(9 citation statements)

References 24 publications

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“…Our research group had previously documented regulation at the translational step of PGHS-1 enzyme synthesis within a context of MEG-01 cell differentiation following treatment with TPA [7,8]. We noted a 2 to 3-day delay between an early rise in PGHS-1 mRNA and an increase in protein steady state levels [7,8]. Our data are in agreement with other studies, which also demonstrate a delay between mRNA transcription and increase in the levels of PGHS-1 protein [9][10][11].…”

Section: Introductionsupporting

confidence: 83%

“…Previous studies from our laboratory demonstrate that translation of PGHS-1 mRNA is delayed after induction of gene expression [8]. Furthermore, this translational regulation is influenced by the presence of both the 5'UTR and the first two exons of PGHS-1 and nucleolin protein that binds to the 5'end of PGHS-1 mRNA [14].…”

Section: Resultsmentioning

confidence: 97%

“…The molecular mechanisms governing the regulation of PGHS-1 gene expression and contributing to an increase in protein levels are poorly understood. Our research group had previously documented regulation at the translational step of PGHS-1 enzyme synthesis within a context of MEG-01 cell differentiation following treatment with TPA [7,8]. We noted a 2 to 3-day delay between an early rise in PGHS-1 mRNA and an increase in protein steady state levels [7,8].…”

Section: Introductionmentioning

confidence: 84%

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Characterization of proteins associating with 5’ terminus of PGHS-1 mRNA

Bunimov

Laneuville

2010

Cellular and Molecular Biology Letters

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Abbreviations used: ARRE-2 -antigen receptor response element-2; ATCC -American Type Culture Collection; DRBP76 -double-stranded RNA-binding protein-76; dsRNAdouble stranded RNA; FBS -fetal bovine serum; GAPDH -glyceraldehyde 3-phosphate dehydrogenase; IL-2 -interleukin-2; IP -immunoprecipitation; IRES -internal ribosome entry site; LC-MS -liquid chromatography mass-spectrometry; Luc -luciferase; MKP-1 -mitogen-activated protein kinase phosphatase 1; MNEI -monocyte/neutrophil elastase inhibitor; mRNP -messenger ribonucleoprotein; NCL -nucleolin; NF45 -nuclear factor 45; NF90 -nuclear factor 90; NFAT -nuclear factor of activated T-cells; ORF -open reading frame; PG -prostaglandins; PGHS -prostaglandin endoperoxide H synthase; PMA -phorbol 12-myristate 13-acetate; SB1 or serpin B1 -serine protease inhibitor; TPA -12-O-tetradecanoylphorbol -13-acetate; Tx -thromboxanes; UTR -untranslated region Immunoprecipitation experiments using MEG-01 protein extracts validated mass spectrometry data and confirmed binding of nucleolin, serpin B1, NF45 and NF90. The RNA fraction was extracted from immunoprecipitated mRNP complexes and association of RNA binding proteins, serpin B1, NF45 and NF90, to PGHS-1 mRNA target sequence was confirmed by RT-PCR. Together these data suggest that serpin B1, NF45 and NF90 associate with PGHS-1 mRNA and can potentially participate in the formation a single or a number of PGHS-1 ribonucleoprotein complexes, through nucleolin that possibly serves as a docking base for other protein complex members.

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Section: Introductionsupporting

confidence: 83%

Section: Resultsmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 84%

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Characterization of proteins associating with 5’ terminus of PGHS-1 mRNA

Bunimov

Laneuville

2010

Cellular and Molecular Biology Letters

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“…At the 3Ј-untranslated region of COX-2 mRNA are multiple AUUUA elements that target mRNAs for rapid exonuclease cleavage (21,26). COX-1 mRNA lacks these 3Ј-untranslated region AUrich elements, and COX-1 mRNA is stable in megakaryocytes and vascular endothelial cells (27)(28)(29). We have recently reported that the C-terminal 19-aa of COX-2 causes the enzyme to undergo proteasomal degradation via the endoplasmic reticulum-associated degradation (ERAD) pathway (18).…”

mentioning

confidence: 99%

Two Distinct Pathways for Cyclooxygenase-2 Protein Degradation

Mbonye

Yuan

Harris

et al. 2008

Journal of Biological Chemistry

103

130

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Cyclooxygenases (COX-1 and COX-2) are N-glycosylated, endoplasmic reticulum-resident, integral membrane proteins that catalyze the committed step in prostanoid synthesis. COX-1 is constitutively expressed in many types of cells, whereas COX-2 is usually expressed inducibly and transiently. The control of COX-2 protein expression occurs at several levels, and overexpression of COX-2 is associated with pathologies such as colon cancer. Here we have investigated COX-2 protein degradation and demonstrate that it can occur through two independent pathways. One pathway is initiated by post-translational N-glycosylation at Asn-594. The N-glycosyl group is then processed, and the protein is translocated to the cytoplasm, where it undergoes proteasomal degradation. We provide evidence from site-directed mutagenesis that a 27-amino acid instability motif (27-IM) regulates posttranslational N-glycosylation of Asn-594. This motif begins with Glu-586 8 residues upstream of the N-glycosylation site and ends with Lys-612 near the C terminus at Leu-618. Key elements of the 27-IM include a helix involving residues Glu-586 to Ser-596 with Asn-594 near the end of this helix and residues Leu-610 and Leu-611, which are located in an apparently unstructured downstream region of the 27-IM. The last 16 residues of the 27-IM, including Leu-610 and Leu-611, appear to promote N-glycosylation of Asn-594 perhaps by causing this residue to become exposed to appropriate glycosyl transferases. A second pathway for COX-2 protein degradation is initiated by substrate-dependent suicide inactivation. Suicide-inactivated protein is then degraded. The biochemical steps have not been resolved, but substrate-dependent degradation is not inhibited by proteasome inhibitors or inhibitors of lysosomal proteases. The pathway involving the 27-IM occurs at a constant rate, whereas degradation through the substrate-dependent process is coupled to the rate of substrate turnover.

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“…The major translational control element of COX-2, the first 60 nucleotides of the 3Ј-UTR, is highly conserved across species, is AU-rich, and contains multiple repeats of the regulatory sequence AUUUA (55)(56)(57). The sequence of the 3Ј-UTR of the COX-1 transcripts is highly divergent from that of COX-2, suggesting a distinct function in the regulation of expression at the post-transcriptional and/or translational levels (10,58). The 3Ј-UTR of the G. fruticosa COX-A is more similar to COX-2, being AU-rich (73.8%) and containing many repeats.…”

Section: Evolutionary Comparison Of Cox Protein Sequences-herementioning

confidence: 99%

On the Evolutionary Origin of Cyclooxygenase (COX) Isozymes

Järving

Kurg

et al. 2004

Journal of Biological Chemistry

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In vertebrates, COX-1 and COX-2, two cyclooxygenase isozymes with different physiological functions and gene regulation, catalyze identical reactions in prostaglandin synthesis. It is still not understood why there are multiple forms of COX enzyme in the same cell type and when the evolutionary duplication of the COX gene occurred. Here we report the structure of two genes encoding for COX isozymes in the coral Gersemia fruticosa, the first non-vertebrate organism from which a cyclooxygenase was characterized. Both genes are about 20 kb in size and consist of nine exons. Intron/exon boundaries are well conserved between coral and mammalian COX genes. mRNAs of the previously reported G. fruticosa COX-A (GenBank TM accession number AY004222) and the novel COX-B share 94% sequence identity in the coding regions and less than 30% in the 5-and 3-untranslated region. Transcripts of both COX genes are detectable in coral cells, although the transcriptional level of COX-A is 2 orders of magnitude higher than COX-B. Expression of both coral genes in mammalian cells gave functional proteins with similar catalytic properties. By data base analyses we also detected and constructed different pairs of COX genes from the primitive chordates, Ciona savignyi and Ciona intestinalis. These two gene pairs encode proteins with 50% intra-species and only 70% cross-species sequence identity. Our results suggest that invertebrate COX gene pairs do not correspond to vertebrate COX-1 and COX-2 and are consistent with duplication of the COX gene having occurred independently in corals, ascidians, and vertebrates. It is evident that due to the importance and complexity of its regulatory role, COX has multiple isoforms in all organisms known to express it, and the genes encoding for the isozymes may to be regulated differently.Prostaglandins are important signaling molecules that are involved in inflammation, ovulation, modulation of immune responses, and mitogenesis. The key enzyme in prostaglandin biosynthesis is prostaglandin-endoperoxide G/H synthase (EC 1.14.99.1), commonly known also as cyclooxygenase (COX) 1 (1, 2). Vertebrates from fish to human beings have two different COX isozymes, COX-1 and COX-2 (3-6). COX-1 is constitutively expressed in most mammalian tissues, and expression levels of this enzyme do not vary greatly in adult animals. COX-2, although absent in most cells, can be rapidly induced in many cell types by inflammatory cytokines, growth factors, and tumor promoters (2, 7). Mammalian COX isozymes are encoded by separate single copy genes that map to distinct chromosomes (1, 8). The human gene for COX-1 is ϳ22 kb in length with 11 exons and is transcribed as 2.8-and 5.2-kb mRNA (9, 10). The gene for COX-2 is about 8.3 kb long with 10 exons, and it is transcribed as 2.8-and 4.6-kb mRNA variants (11-15). The gene structures of COX-1 and COX-2 demonstrate remarkable conservation of exon/intron junctions (16 -19). The main differences between the COX-1 and COX-2 genes consist of different intron lengths and one missing int...

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Translational Regulation of Prostaglandin Endoperoxide H Synthase-1 mRNA in Megakaryocytic MEG-01 Cells

Cited by 13 publications

References 24 publications

Characterization of proteins associating with 5’ terminus of PGHS-1 mRNA

Characterization of proteins associating with 5’ terminus of PGHS-1 mRNA

Two Distinct Pathways for Cyclooxygenase-2 Protein Degradation

On the Evolutionary Origin of Cyclooxygenase (COX) Isozymes

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