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A minor fraction of basic fibroblast growth factor mRNA is deaminated in Xenopus stage VI and matured oocytes

Published online by Cambridge University Press:  01 January 1999

LISA SACCOMANNO
Affiliation:
Department of Biochemistry and Howard Hughes Medical Institute, 50 North Medical Drive, University of Utah, Salt Lake City, Utah 84132, USA Present address: Department of Cell and Molecular Biology, Northwestern University Medical School, 303 East Chicago, Chicago, Illinois 60611, USA.
BRENDA L. BASS
Affiliation:
Department of Biochemistry and Howard Hughes Medical Institute, 50 North Medical Drive, University of Utah, Salt Lake City, Utah 84132, USA
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Abstract

Adenosine deaminases that act on RNA (ADARs) convert adenosine to inosine in double-stranded regions of RNA. ADAR activity is in the nucleus in Xenopus laevis stage VI oocytes, and released into the cytoplasm at oocyte maturation. We previously demonstrated that a cytoplasmic double-stranded RNA (dsRNA) binding factor(s), cyto-dsRBP, protects microinjected dsRNA from the ADAR released at maturation. Here we describe experiments to determine whether an endogenous dsRNA, the duplex formed between sense and antisense transcripts of basic fibroblast growth factor (bFGF), is protected in a similar manner. Consistent with the presence of cyto-dsRBP, we observed that the majority of bFGF RNA was not deaminated, before or after maturation. However, a minor fraction of the bFGF RNA was deaminated whether the RNA was isolated from stage VI oocytes or matured oocytes. Since ADAR activity is in the nucleus in stage VI oocytes, our results suggest that a fraction of the bFGF RNAs are hybridized in the nucleus and are ADAR substrates. Adenosine deaminations result in A-to-G changes in cDNAs, so we quantified the fraction of modified molecules using restriction-enzyme assays of RT-PCR products. Caveats due to recombination during RT-PCR are discussed.

Type
Research Article
Copyright
© 1999 RNA Society

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