Specific 3' sequences flanking a minimal apolipoprotein B (apoB) mRNA editing 'cassette' are critical for efficient editing in vitro.
Apolipoprotein B (apoB) mRNA in mammalian intestine undergoes direct conversion of cytidine to uridine at nucleotide 6666, generating a UAA stop codon which defines the carboxyl-terminus of apoB48. We have identified three distinct sequence elements which are required for efficient apoB RNA editing in vitro and have defined a 20 nucleotide 'editing cassette' which will support efficient editing in the context of a variety of AT-rich apoB mRNA backgrounds. An important question remaining to be addressed is whether this cassette can support editing in a GC-rich background characteristic of other mRNAs. We demonstrate that the context into which the editing cassette is inserted may determine the efficiency of editing site utilization. When an editing cassette is placed in the context of human albumin mRNA sequence, editing is reduced 6-fold relative to a construct of similar length in which the cassette is surrounded by apoB mRNA sequence. Chimeric RNA substrates may be efficiently edited, however, when albumin sequence resides only 5' of the editing cassette. The data suggest that the proximal of an AT-rich, apoB-like, mRNA sequence is only required 3' of the editing cassette. These results should prove useful in evaluating the potential for editing on mRNAs where similar cassettes can be found.
Backus
J W
JW
Department of Biochemistry, University of Rochester, NY 14642.
Smith
H C
HC
D000071479:APOBEC-1 Deaminase; D001055:Apolipoproteins B / Q000235:genetics*; D001483:Base Sequence; D002678:Chimera; D003564:Cytidine Deaminase / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D017393:RNA Editing; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000235:genetics; D000098830:AICDA (Activation-Induced Cytidine Deaminase)
D001055:Apolipoproteins B; D012333:RNA, Messenger; D000098830:AICDA (Activation-Induced Cytidine Deaminase); D000071479:APOBEC-1 Deaminase; C000605089:APOBEC1 protein, human; D003564:Cytidine Deaminase