RNA editing of non-coding RNA and its role in gene regulation

Highlights

• Primate specific inverted Alu repeats frequently form double stranded structures in transcripts.

• Up to 100 million adenosines can be deaminated to inosine in the human transcriptome.

• Most A-to-I editing sites in human are located to non-coding Alu RNA.

• In concert Alu inverted repeats and RNA editing modify gene expression.

Abstract

It has for a long time been known that repetitive elements, particularly Alu sequences in human, are edited by the adenosine deaminases acting on RNA, ADAR, family. The functional interpretation of these events has been even more difficult than that of editing events in coding sequences, but today there is an emerging understanding of their downstream effects. A surprisingly large fraction of the human transcriptome contains inverted Alu repeats, often forming long double stranded structures in RNA transcripts, typically occurring in introns and UTRs of protein coding genes. Alu repeats are also common in other primates, and similar inverted repeats can frequently be found in non-primates, although the latter are less prone to duplex formation. In human, as many as 700,000 Alu elements have been identified as substrates for RNA editing, of which many are edited at several sites. In fact, recent advancements in transcriptome sequencing techniques and bioinformatics have revealed that the human editome comprises at least a hundred million adenosine to inosine (A-to-I) editing sites in Alu sequences. Although substantial additional efforts are required in order to map the editome, already present knowledge provides an excellent starting point for studying cis-regulation of editing. In this review, we will focus on editing of long stem loop structures in the human transcriptome and how it can effect gene expression.

Introduction

Up to 75% of the human genome is known to be transcribed but only a couple of percent of the total amount of RNA is protein coding [1]. Lately, much attention has been given to the increasing number of long non-coding RNAs (lncRNAs) with proven functionality. In the search for functional non-coding RNA, potential functionality within introns and UTRs is often overlooked. However, non-coding RNA elements can also be embedded within pre-mRNAs and structural elements in introns can influence RNA processing of exons. The primate specific Alu repeats, a repetitive small interspersed element (SINE), are abundant in the human transcriptome, particularly within gene rich regions [2], [3], and can act as cis-inducers for RNA modifications. In all multi-cellular organisms, double stranded RNA (dsRNA) molecules are subjected to RNA modifications by adenosine deamination. This was first discovered when long completely base paired RNA-duplexes, injected into frog oocytes, were observed to become unwinded to single stranded molecules [4]. This was later shown to be due to modifications of adenosine to inosine (A-to-I) catalyzed by the family of adenosine deamination enzymes, i.e., the ADAR family [5]. The mammalian genome has two active members of this family, ADAR1 and ADAR2, see Refs. [6], [7], [8]. ADAR1 is both nuclear and cytoplasmic; ADAR2 is in contrast mainly present in the nucleus [9], [10], [11], [12]. It is known that most editing events occur in the nucleus and, since inosine is structurally similar to guanosine, in cellular processes such as splicing and translation, inosine is typically recognized as guanosine [13]. Even though in principle any dsRNA could be subjected to editing, the only known substrates for ADAR enzymes are RNA polymerase II transcripts.

No specific sequence has been found that characterize editing sites of any of the ADAR enzymes; however, in the neighboring position downstream of the edited A, there is an overrepresentation of G, while G is underrepresented in the upstream neighboring position [14], [15]. ADAR1 has lower site selectivity than ADAR2, but their sets of target sites are overlapping [16]. It appears that, in general, the number of edited sites in a duplex depends on its length in a superlinear fashion. Short stems interrupted by bulges and internal loops are often edited only at one specific site, while longer stem loop structures often are hyper-edited at the majority of the adenosines in a double stranded region (reviewed in Ref. [17]). For a handful of brain specific genes, involved in neurotransmission, A-to-I editing of coding sequences has been found to be of high functional importance (reviewed in Ref. [18]). The vast majority of all editing sites reside, however, in non-coding regions, particularly in inverted Alu repeats [19], [20], [21], [22]. In this review, we will focus on how these and other highly structured non-coding RNA sequences regulate both expression and processing of encoded genes in cis by attracting the A-to-I RNA editing enzyme ADAR. We also discuss the role of ADAR in circRNA biogenesis.