Long non-coding RNA Airn (antisense to Igf2r RNA non-coding) was identified in the heart in 1997 in analysis of developmental process that induces parental-specific expression or repression of autosomal and X-chromosome-line genes 133,134. Airn is an imprinted gene transcribed specifically from paternal chromosome, which suppresses its counterparts, Igf2r gene (encoding the receptor for insulin-like growth factor type-2) from the maternal allele after embryonic implantation. On human and mouse chromosome, Airn is located in antisense orientation to Igf2r which is maternally transcribed. Igf2r locus harbors three maternally expressed protein-coding genes Igf2r, Slc22a2, and Slc22a3 133,134 and a paternally expressed gene, which is lncRNA Airn. It is now well known that transcription of Airn from intron-2 of Igf2r genes suppress the expression of Igf2r itself and the nearby protein coding genes Slc22a2, Slc22a3. However, it is not confirmed in which mechanism these genes are silenced even though it was clearly described that expression of Airn is necessary to silence the imprinted Igf2r cluster 133-137. Airn transcript underogoes alternative splicing which generates to six isoforms. Nonetheless, a non-spliced version of the transcript can be generated. Airn contains polyadenylated tail and 5′-cap as mRNAs 135. A further study demonstrates that spliced Airn isoforms dwell mainly in the nucleus but is capable of cytoplasmic shuttling. Oppositely, the mature unspliced version exists only in nucleus and is highly unstable 134.
According to ENSEMBL database, it is described that Airn is located on chromosome 17: 12,741,311-12,860,122 (+), it gives rise to 118 kb unspliced lncRNA and the following 6 spliced isoforms: Airn-001 (ENSMUST00000079529; 1,113-nt), Airn-002 (ENSMUST00000160932; 842-nt), Airn-003 (ENSMUST00000162826; 521-nt), Airn-004 (ENSMUST00000159791; 485-nt), Airn-005 (ENSMUST00000159731; 435-nt), and Airn-006 (ENSMUST00000161890; 1,375-nt) (Figure 7). The level of expression and abundance of Airn was first analysed by using RNase protection assays (RPA) to determine the steady-state levels of Igf2r and Airn in adult mouse tissues and cell lines expressing different levels of these two genes. These study demonstrated that Airn-001 expression is equally abundant in the heart and E11.5 placenta 140. It was described previously that Airn-001 isoform has most abundant level of expression and investigated for genome imprinting of nearby genes 133-136.A study reported that unspliced Airn has a half-life between 1.6 h and 2.1 h that it is approximately 10-fold less stable than the Igf2r mRNA. A comparative study revealed that Airn-001 is also three-fold less stable than the nuclear-localized Xist ncRNA in mouse embryonic fibroblasts (MEF). Recent study described that ratio of stability between Igf2r and Airn resembles to that of Xist/Tsix sense/antisense ncRNAs 133.
It is well known that paternally expressed lncRNA Airn transcribed in the antisense orientation to its host gene Igf2r 137. A recent study by using inserted termination signals at various distance from the transcription initiation site of Airn in ES cells suggests that Airn transcription, rather than the transcript, controls the silencing of Igf2r. Interestingly, for the Airn transcription-dependent silencing of Igf2r a reversible and continuous transcription of Airn is required 138.
Airn itself has been shown to silence the non-overlapping genes via recruitment of chromatin modifiers in a sequence-specific fashion to the non-overlapping gene promoters 138.
More importantly, both Airn transcript and the act of its transcription regulate the transcription of overlapping and non-overlapping genes (Figure 8). Regrettably, the understanding of lncRNA transcription-dependent silencing still has shortcoming. In addition, the mechanisms by which DNMTs are attracted to the Igf2r promoter are still elusive 137. An outstanding study was conducted to determine the role of Airn in silencing. In this study, Airn was truncated to 4% of its length by inserting a 1.2-kb polyadenylation cassette from the b-globin gene at the downstream border of the 3.7-kb ICE (Imprintnig control element). Such modification was designed to truncate Airn without disrupting the function of the 3.7-kb ICE region, as assessed by its ability to attract a maternal-specific methylation imprint and to show paternal-specific expression of the Airn promoter (139). The investigation has resulted in viable heterozygous or homozygous mice with the paternally inherited allele of Airn showing 15% reduced body weight in comparison to wild-type littermates at birth 134. Conversely, the phenotype caused by targeted disruption of Igf2r is demonstrated with an increased serum and tissue level of IGF-II, overgrowth, and a number of developmental defects (Organomegaly Kinky tail, postaxial polydactyly, heart abnormalities, dyspnea, and perinatal death, which are affected by genetic background 140.
A recent study described human homolog of Airn in the human IGF2R intron-2 CpG Island that has promoter activity. Human AIRN is also expressed from human IGF2R intron-2 CpG Island 141.
Nevertheless, experimental situations have some distinct features such as in humans, adult tissues lack IGF2R imprinted expression, but it is exists in found in fetal tissues and Wilms’ tumours 141. Mouse Igf2r imprinted expression is controlled by Airn whose promoter lies in an intronic maternally-methylated CpG Island. The human IGF2R gene conveys a homologous intronic maternally-methylated CpG island of unknown function 141.
In contrast with the majority of lncRNAs, most of the imprinted lncRNAs are relatively highly conserved in the functional and primary sequence level. Thus, imprinting lncRNA becoming one of the most tractable systems to investigate in development and disease 141, 142. Importantly, lncRNAs from well-investigated imprinted clusters offer insights into their mechanism of action in regulating target genes, and these mechanisms have formed the underlying basis for uncovering the mode of action of lncRNAs in several other biological contexts.