Poster abstracts

Poster number 9 submitted by John Cook

Non-catalytic embryonic dADAR isoform is evolutionarily-conserved in Drosophila but not translated into protein during embryogenesis

John Cook (Zoology, Cell and Molecular and Structural Biology Program, Miami University), Lea Chhiba (Zoology, Cell and Molecular and Structural Biology Program, Miami University), Dana Doctor (Zoology, Cell and Molecular and Structural Biology Program, Miami University), Jack Vaughn (Zoology, Cell and Molecular and Structural Biology Program, Miami University)

Abstract:
Adenosine Deaminases Acting on RNA (ADARs) function to deaminate (edit) some adenosines to inosines in selected pre-mRNAs. The single-copy ADAR gene in Drosophila melanogaster (dADAR) produces two major mRNA transcript classes, termed full-length and truncated. The catalytic full-length class has been functionally well characterized as being most active in the post-pupal brain. In contrast, virtually nothing is known about the function of the non-catalytic truncated “embryonic” isoform, which in D. melanogaster has been fully sequenced and shown to contain a complete open reading frame, a stop codon in intron 6, and a poly(A)-tail. Here, we show via 3’-RACE that this mRNA transcript class is present in every Drosophila species studied, extending back to those diverging from D. melanogaster 40 million years ago. In every species studied, the sequenced transcripts terminate in intron 6, have a stop codon, and a poly(A)-tail. Utilization of Westerns employing a polyclonal anti-dADAR antibody produced using full-length protein shows that the full-length isoform class in D. melanogaster encodes a protein with the expected molecular weight at every developmental stage. Quantification of these results shows that this isoform is most abundant between 0 and 4 h of embryo development, then gradually declines during embryogenesis. Unexpectedly, the truncated isoform does not appear to encode a protein during any embryonic stage of development in D. melanogaster. This was a surprise, since this mRNA isoform class is generated in every species studied, and is highly abundant throughout embryogenesis, suggesting that it has a conserved function. These observations suggest that the truncated dADAR mRNA transcript class may function in some unknown way at the RNA level, perhaps playing a role in regulation of full-length isoform editase function, an isoform class which has virtually no catalytic activity during embryogenesis despite its demonstrated existence.

Keywords: RNA editing, dADAR mRNA isoforms, regulation of editase function