Novel Exon of Mammalian ADAR2 Extends Open Reading Frame

Abstract

BackgroundThe post-transcriptional processing of pre-mRNAs by RNA editing contributes significantly to the complexity of the mammalian transcriptome. RNA editing by site-selective A-to-I modification also regulates protein function through recoding of genomically specified sequences. The adenosine deaminase ADAR2 is the main enzyme responsible for recoding editing and loss of ADAR2 function in mice leads to a phenotype of epilepsy and premature death. Although A-to-I RNA editing is known to be subject to developmental and cell-type specific regulation, there is little knowledge regarding the mechanisms that regulate RNA editing in vivo. Therefore, the characterization of ADAR expression and identification of alternative ADAR variants is an important prerequisite for understanding the mechanisms for regulation of RNA editing and the causes for deregulation in disease.Methodology/Principal FindingsHere we present evidence for a new ADAR2 splice variant that extends the open reading frame of ADAR2 by 49 amino acids through the utilization of an exon located 18 kilobases upstream of the previously annotated first coding exon and driven by a candidate alternative promoter. Interestingly, the 49 amino acid extension harbors a sequence motif that is closely related to the R-domain of ADAR3 where it has been shown to function as a basic, single-stranded RNA binding domain. Quantitative expression analysis shows that expression of the novel ADAR2 splice variant is tissue specific being highest in the cerebellum.Conclusions/SignificanceThe strong sequence conservation of the ADAR2 R-domain between human, mouse and rat ADAR2 genes suggests a conserved function for this isoform of the RNA editing enzyme.

Adenosine Deaminases Acting on RNA, RNA Editing, and Interferon Action

“…While human and mouse ADAR2 genes have generally similar exon/intron organizations (Slavov and Gardiner 2002), the complexity of their exon structures has not been appreciated until recently. Both the human and mouse ADAR2 genes are now known to possess 15 exons (Slavov and Gardiner 2002;Maas and Gommans 2009) as summarized in Fig. 3.…”

“…Exons 0-9 represent the ORF that encodes ADAR2. Similar to ADAR1, various ADAR2 transcript isoforms exist as a result of multiple alternative splicing events (Gerber and others 1997;Lai and others 1997;Mittaz and others 1997;Slavov and Gardiner 2002;Kawahara and others 2005;Maas and Gommans 2009). …”

“…Comparison of human and mouse genomic sequences with the 5 0 -terminal sequences of ADAR2 cDNAs identifies 3 exons, denoted exon 1a, 0, and 1 for human and exon 1b, 0, and 1 for mouse, respectively (Slavov and Gardiner 2002;Maas and Gommans 2009). Each of these exons contains a potential AUG codon for initiation of ADAR2 translation.…”

“…In addition, the AUG-28 codon in human exon 1a (or mouse exon 1b) does not occur in the À3/þ4 context of a consensus Kozak sequence (A/GCCatgG) (Kozak 1989), raising the possibility that these transcripts are poorly translated. Recently, an additional exon designated as exon 0 positioned between exon À1 and exon 1 has been characterized for both human and mouse ADAR2 (Maas and Gommans 2009). Inclusion of exon 0 with initiation at AUG-49 results in the addition of 49 amino acids N-terminal of the previously identified initiation methionine (AUG1) in exon 1; the added sequence exhibits a high similarity to the Arg-rich domain present in the N-terminal region of ADAR3.…”

Host Response to Polyomavirus Infection Is Modulated by RNA Adenosine Deaminase ADAR1 but Not by ADAR2

“…Two size forms of ADAR1 protein are expressed by a mechanism involving alternative adar1 promoter usage and alternative exon 1 splicing: a short (p110) nuclear form that is constitutively made and a long (p150) form that is interferon inducible and present in both the cytoplasm and nucleus (7,8,36,37). The mouse adar2 gene maps to chromosome 10 C1, is constitutively expressed, and is not known to be regulated by IFN or infection (10,16,29). The ADARs possess multiple copies of the canonical double-stranded RNA (dsRNA) binding motif first discovered in the PKR kinase (30), with three copies present in both ADAR1 p110 and p150 and two copies in ADAR2 (33,46).…”