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Molecular and Cellular Biology, March 2000, p. 1846-1854, Vol. 20, No. 5
Department of Cell Biology, Lerner Research
Institute, Cleveland Clinic Foundation, Cleveland, Ohio
44195,1 and W. M. Keck Biomedical
Mass Spectrometry Laboratory, Department of Microbiology, University of
Virginia, Charlottesville, Virginia 229082
Received 21 October 1999/Returned for modification 27 November
1999/Accepted 1 December 1999
The C-to-U editing of apolipoprotein B (apo-B) mRNA is catalyzed by
a multiprotein complex that recognizes an 11-nucleotide mooring
sequence downstream of the editing site. The catalytic subunit of the
editing enzyme, apobec-1, has cytidine deaminase activity but requires
additional unidentified proteins to edit apo-B mRNA. We purified a
65-kDa protein that functionally complements apobec-1 and obtained
peptide sequence information which was used in molecular cloning
experiments. The apobec-1 complementation factor (ACF) cDNA encodes a
novel 64.3-kDa protein that contains three nonidentical RNA recognition
motifs. ACF and apobec-1 comprise the minimal protein requirements for
apo-B mRNA editing in vitro. By UV cross-linking and
immunoprecipitation, we show that ACF binds to apo-B mRNA in vitro and
in vivo. Cross-linking of ACF is not competed by RNAs with mutations in
the mooring sequence. Coimmunoprecipitation experiments identified an
ACF-apobec-1 complex in transfected cells. Immunodepletion of ACF from
rat liver extracts abolished editing activity. The immunoprecipitated
complexes contained a functional holoenzyme. Our results support a
model of the editing enzyme in which ACF binds to the mooring sequence
in apo-B mRNA and docks apobec-1 to deaminate its target cytidine. The
fact that ACF is widely expressed in human tissues that lack apobec-1 and apo-B mRNA suggests that ACF may be involved in other RNA editing
or RNA processing events.
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Molecular Cloning of Apobec-1 Complementation
Factor, a Novel RNA-Binding Protein Involved in the Editing of
Apolipoprotein B mRNA

*
Corresponding author. Mailing address: Department of
Cell Biology, Lerner Research Institute, Cleveland Clinic Foundation, 9500 Euclid Ave. #NC-10, Cleveland, OH 44195. Phone: (216) 445-9758. Fax: (216) 444-9404. E-mail: driscod{at}ccf.org.
Present address: Department of Cell Biology, Lerner Research
Institute, Cleveland Clinic Foundation, Cleveland, OH 44195.
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