APOBEC-1, THE CATALYTIC SUBUNIT OF THE MAMMALIAN APOLIPOPROTEIN-B MESSENGER-RNA EDITING ENZYME, IS A NOVEL RNA-BINDING PROTEIN

Apolipoprotein B (apoB) mRNA editing is mediated by an enzyme complex which includes the catalytic subunit, apobec-1. Recombinant GST/APOBEC -1 binds with high specificity to a rat apoB RNA template as demonstra ted by UV cross-linking and electrophoretic mobility shift assay (ERIS A), ApoB RNA binding was competed by poly(U), poly(A,U), and tRNA, but not by poly(A) or other homopolymeric ribonucleotides, UV cross-linki ng of GST/APOBEC-1 to an apoB RNA template was uninfluenced by the bin ding of proteins of similar to 60 and similar to 44 kDa, present in S1 00 extracts prepared from different sources, The binding of these prot eins was similarly uninfluenced by the simultaneous binding of GST/APO -BEC-1. Moreover, the inclusion of heterologous S100 extracts in the R NA binding reactions completely abrogated the competitive displacement of GST/APOBEC-1 by tRNA. EMSA revealed the onset of RNA binding withi n 1-2 min, and its specificity was confirmed by a supershift with anti -GST/APOBEC-1 antisera. The structural specificity for apoB RNA bindin g, as inferred from EMSA, appears to be distinct from apoB RNA editing since wild-type chicken apoB RNA,which is not editable, and several m utant chicken apoB RNAs containing clustered mutations within the mini mal apoB RNA editing cassette, bound with efficiency similar to the ra t apoB RNA template. In conclusion, while the data suggest that apobec -1 binds AU-rich templates, the importance of this observation in the context of mammalian apoB mRNA editing remains unknown.