TY - JOUR
T1 - apobec-1, the catalytic subunit of the mammalian apolipoprotein B mRNA editing enzyme, is a novel RNA-binding protein
AU - Anant, S.
AU - MacGinnitie, A. J.
AU - Davidson, N. O.
PY - 1995/6/16
Y1 - 1995/6/16
N2 - 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 demonstrated by UV cross-linking and electrophoretic mobility shift assay (EMSA). ApoB RNA binding was competed by poly(U), poly(A,U), and tRNA, but not by poly(A) or other homopolymeric ribonucleotides. UV cross-linking of GST/APOBEC-1 to an apoB RNA template was uninfluenced by the binding of proteins of ~60 and ~44 kDa, present in S100 extracts prepared from different sources. The binding of these proteins was similarly uninfluenced by the simultaneous binding of GST/APOBEC-1. Moreover, the inclusion of heterologous S100 extracts in the RNA binding reactions completely abrogated the competitive displacement of GST/APOBEC-1 by tRNA. EMSA revealed the onset of RNA binding within 1-2 min, and its specificity was confirmed by a supershift with anti- GST/APOBEC-1 antisera. The structural specificity for apoB RNA binding, as inferred from EMSA, appears to be distinct from apoB RNA editing since wild- type chicken apoB RNA, which is not editable, and several mutant chicken apoB RNAs containing clustered mutations within the minimal apoB RNA editing cassette, bound with efficiency similar to the rat 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.
AB - 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 demonstrated by UV cross-linking and electrophoretic mobility shift assay (EMSA). ApoB RNA binding was competed by poly(U), poly(A,U), and tRNA, but not by poly(A) or other homopolymeric ribonucleotides. UV cross-linking of GST/APOBEC-1 to an apoB RNA template was uninfluenced by the binding of proteins of ~60 and ~44 kDa, present in S100 extracts prepared from different sources. The binding of these proteins was similarly uninfluenced by the simultaneous binding of GST/APOBEC-1. Moreover, the inclusion of heterologous S100 extracts in the RNA binding reactions completely abrogated the competitive displacement of GST/APOBEC-1 by tRNA. EMSA revealed the onset of RNA binding within 1-2 min, and its specificity was confirmed by a supershift with anti- GST/APOBEC-1 antisera. The structural specificity for apoB RNA binding, as inferred from EMSA, appears to be distinct from apoB RNA editing since wild- type chicken apoB RNA, which is not editable, and several mutant chicken apoB RNAs containing clustered mutations within the minimal apoB RNA editing cassette, bound with efficiency similar to the rat 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.
UR - http://www.scopus.com/inward/record.url?scp=0029004350&partnerID=8YFLogxK
U2 - 10.1074/jbc.270.24.14762
DO - 10.1074/jbc.270.24.14762
M3 - Article
C2 - 7782342
AN - SCOPUS:0029004350
SN - 0021-9258
VL - 270
SP - 14762
EP - 14767
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 24
ER -