Post-transcriptional control of cyclooxygenase-2 gene expression. The role of the 3'-untranslated region.
The cyclooxygenase (COX)-2 enzyme is responsible for increased prostaglandin formation in inflammatory states and is the major target of nonsteroidal anti-inflammatory drugs. Normally COX-2 expression is tightly regulated, however, constitutive overexpression plays a key role in colon carcinogenesis. To understand the mechanisms controlling COX-2 expression, we examined the ability of the 3'-untranslated region of the COX-2 mRNA to regulate post-transcriptional events. When fused to a reporter gene, the 3'-untranslated region mediated rapid mRNA decay (t(1/2) = 30 min), which was comparable to endogenous COX-2 mRNA turnover in serum-induced fibroblasts treated with actinomycin D or dexamethasone. Deletion analysis demonstrated that a conserved 116-nucleotide AU-rich sequence element (ARE) mediated mRNA degradation. In transiently transfected cells, this region inhibited protein synthesis approximately 3-fold. However, this inhibition did not occur through changes in mRNA stability since mRNA half-life and steady-state mRNA levels were unchanged. RNA mobility shift assays demonstrated a complex of cytoplasmic proteins that bound specifically to the ARE, and UV cross-linking studies identified proteins ranging from 90 to 35 kDa. Fractionation of the cytosol showed differential association of ARE-binding proteins to polysomes and S130 fractions. We propose that these factors influence expression at a post-transcriptional step and, if dysregulated, may increase COX-2 protein as detected in colon cancer.
Dixon
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DA
Department of Oncological Sciences, Eccles Program in Human Molecular Biology and Genetics, and the Huntsman Cancer Institute, University of Utah, Salt Lake City, Utah 84112, USA. dan.dixon@hci.utah.edu
Kaplan
C D
CD
McIntyre
T M
TM
Zimmerman
G A
GA
Prescott
S M
SM
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D051546:Cyclooxygenase 2; D004591:Electrophoresis, Polyacrylamide Gel; D015971:Gene Expression Regulation, Enzymologic; D006367:HeLa Cells; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis*; D008565:Membrane Proteins; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis*; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000378:metabolism
D007527:Isoenzymes; D008565:Membrane Proteins; D010446:Peptide Fragments; D012333:RNA, Messenger; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases