The AU-rich element (ARE) is an important instability determinant for a large number of early-response-gene mRNAs. AREs also mediate the stabilization of certain pro-inflammatory mRNAs, such as tumour necrosis factor (TNF)-α and cyclo-oxygenase-2 (COX-2), in response to inflammatory stimuli. To understand how AREs control mRNA stability, it is necessary to identify trans-acting factors. We have purified a new ARE-binding protein and identified it as CArG box-binding factor-A (CBF-A). The amino acid sequence of CBF-A is highly similar to that of the ARE-binding protein AUF1. Recombinant CBF-A bound the COX-2 and TNF-α AREs, but not a non-specific control RNA. In contrast, in an electrophoretic-mobility-shift assay (EMSA) of crude RAW 264.7 macrophage-like cell extracts, an antiserum that recognizes both AUF1 and CBF-A failed to supershift complexes formed on the TNF-α ARE, but did supershift a complex specific for the COX-2 ARE. CBF-A exists as two isoforms, p37 and p42, that differ by a 47-amino-acid insertion close to the C-terminus. By expressing epitope-tagged isoforms of CBF-A it was shown that the p42 isoform binds the COX-2 ARE in EMSA of crude cell extracts. In a HeLa-cell tetracycline-regulated reporter system, overexpression of the p42 CBF-A isoform resulted in stabilization of a COX-2 ARE reporter mRNA. Epitope-tagged p42 CBF-A expressed in HeLa cells co-immunoprecipitated with endogenous COX-2 mRNA, but not glyceraldehyde-3-phosphate dehydrogenase mRNA, as shown by reverse-transcription PCR. The similarity between CBF-A and AUF1 suggests that CBF-A could be re-named AUF2.
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September 2002
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Research Article|
September 15 2002
Identification of a novel AU-rich-element-binding protein which is related to AUF1
Jonathan L.E. DEAN;
Jonathan L.E. DEAN
1
Kennedy Institute of Rheumatology Division, Faculty of Medicine, Imperial College of Science, Technology and Medicine, 1 Aspenlea Rd, Hammersmith, London W6 8LH, U.K.
1To whom correspondence should be addressed (e-mail jonathan.dean@ic.ac.uk).
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Gareth SULLY;
Gareth SULLY
2
Kennedy Institute of Rheumatology Division, Faculty of Medicine, Imperial College of Science, Technology and Medicine, 1 Aspenlea Rd, Hammersmith, London W6 8LH, U.K.
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Robin WAIT;
Robin WAIT
2
Kennedy Institute of Rheumatology Division, Faculty of Medicine, Imperial College of Science, Technology and Medicine, 1 Aspenlea Rd, Hammersmith, London W6 8LH, U.K.
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Lesley RAWLINSON;
Lesley RAWLINSON
Kennedy Institute of Rheumatology Division, Faculty of Medicine, Imperial College of Science, Technology and Medicine, 1 Aspenlea Rd, Hammersmith, London W6 8LH, U.K.
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Andrew R. CLARK;
Andrew R. CLARK
Kennedy Institute of Rheumatology Division, Faculty of Medicine, Imperial College of Science, Technology and Medicine, 1 Aspenlea Rd, Hammersmith, London W6 8LH, U.K.
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Jeremy SAKLATVALA
Jeremy SAKLATVALA
Kennedy Institute of Rheumatology Division, Faculty of Medicine, Imperial College of Science, Technology and Medicine, 1 Aspenlea Rd, Hammersmith, London W6 8LH, U.K.
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Publisher: Portland Press Ltd
Received:
March 12 2002
Revision Received:
June 20 2002
Accepted:
June 27 2002
Accepted Manuscript online:
June 27 2002
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2002
2002
Biochem J (2002) 366 (3): 709–719.
Article history
Received:
March 12 2002
Revision Received:
June 20 2002
Accepted:
June 27 2002
Accepted Manuscript online:
June 27 2002
Citation
Jonathan L.E. DEAN, Gareth SULLY, Robin WAIT, Lesley RAWLINSON, Andrew R. CLARK, Jeremy SAKLATVALA; Identification of a novel AU-rich-element-binding protein which is related to AUF1. Biochem J 15 September 2002; 366 (3): 709–719. doi: https://doi.org/10.1042/bj20020402
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