Molecular and Cellular Biology, October 1998, p. 6014-6022, Vol. 18, No. 10
0270-7306/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Seattle Biomedical Research Institute,
Seattle, Washington, 98109-1651,1 and
Department
of Pathobiology,
Received 6 April 1998/Returned for modification 3 June
1998/Accepted 19 June 1998
RNA editing in Trypanosoma brucei mitochondria produces
mature mRNAs by a series of enzyme-catalyzed reactions that
specifically insert or delete uridylates in association with a
macromolecular complex. Using a mitochondrial fraction enriched for in
vitro RNA editing activity, we produced several monoclonal antibodies that are specific for a 21-kDa guide RNA (gRNA) binding protein initially identified by UV cross-linking. Immunofluorescence studies localize the protein to the mitochondrion, with a preference for the
kinetoplast. The antibodies cause a supershift of previously identified
gRNA-specific ribonucleoprotein complexes and immunoprecipitate in
vitro RNA editing activities that insert and delete uridylates. The
immunoprecipitated material also contains gRNA-specific
endoribonuclease, terminal uridylyltransferase, and RNA ligase
activities as well as gRNA and both edited and unedited mRNA. The
immunoprecipitate contains numerous proteins, of which the 21-kDa
protein, a 90-kDa protein, and novel 55- and 16-kDa proteins can be UV
cross-linked to gRNA. These studies indicate that the 21-kDa protein
associates with the ribonucleoprotein complex (or complexes)
that catalyze RNA editing.
*
Corresponding author. Mailing address: Seattle
Biomedical Research Institute, 4 Nickerson St., Seattle, WA
98109-1651. Phone: (206) 284-8846, ext. 316. Fax: (206) 284-0313. E-mail: kstuart{at}u.washington.edu.
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