Non-coding RNAs (ncRNA) are RNA molecules that do not code for a protein and therefore are not translated into proteins. While showing a wide range of functionality, most of the ncRNAs have regulatory or housekeeping roles. Epigenetically functional ncRNAs include microRNAs (miRNA), long non-coding RNAs (IncRNA) and circular RNAs (circRNA). In addition to their functional diversity, ncRNAs can also be classified according to their size as short ncRNAs and long ncRNAs (Zaratiegui, Irvine, and Martienssen 2007). ncRNAs with maximum length of 200 nucleotides (nt) are considered short ncRNAs while ncRNAs longer than 200 nt are classified as long ncRNAs.
The most extensively studied group of short ncRNAs are miRNAs. miRNAs are single stranded ~20 nt long regulatory RNAs which mainly function as post-tran- scriptional regulators of gene expression.
miRNAs are transcribed by RNA polymerase II as a hairpin loop structure, called the pri-miRNA. Following transcription, pri-miRNAs are capped and poly- adenilated (Cai, Hagedorn, and Cullen 2004). Pri-miRNAs undergo a nuclear processing where the pri-miRNA is cleaved by microprocessor complex, comprised of DiGeorge syndrome critical region 8 (DGCR8) and Drosha proteins (Gregoryi, Chendrimada, and Shiekhattar 2006; Conrad et al. 2014). The cleavage product is called a pre-miRNA. Nuclear processing is followed by the export of pre-miRNA to cytoplasm by Exportin. Pre-miRNA is cleaved by an enzyme called dicer to yield a double stranded miRNA complex. Following the cleavage by dicer, one strand of this miRNA duplex is loaded into the RNA-induced silencing complex (RISC) to interact with its target (Kim and Kim 2012; Park et al. 2011). Gene silencing by miRNAs has two modes depending on the miRNA target complementarity. Perfect or near perfect complementarity between the miRNA and its target mRNA induces the cleavage and degradation of target mRNA. In case of a non-perfect complementarity, the target mRNA is silenced through inhibition of translation (Lim et al. 2005).
Circular RNAs are formed through a process called backsplicing during which splice acceptor and splice donor sites of a pre-mRNA are covalently joined to produce a circular transcript (Figure 15.4) (Barrett, Wang, and Salzman 2015).
The lack of a 5' or 3' end makes these RNAs more stable due to their resistance to exonuclease-mediated degradation. One study has shown that their half-life is at least 2.5 times longer than the half-life of their linear RNAs (Enuka et al. 2016).
Several circRNAs show tissue-specific expression, indicating a tissue-dependent function role for ncRNAs (Salzman et al. 2013). This is also supported by the discrepancy between circRNA and corresponding mRNA levels (Salzman et al. 2012).
One of first circRNAs to be characterized is the mouse circRNA: Sry. Findings indicate a very interesting function for this circRNA. Evidently, Sry represses miR- 138 activity by binding 16 miR-138 molecules and acting as a miRNA sponge (Hansen et al. 2013). ciRS-7 is another circRNA w'hich functions as a miRNA sponge. A recent study has shown that ciRS-7 can bind more than 70 molecules of miR-7 (Memczak et al. 2013).
Transcriptional or post-transcriptional regulation has also been suggested as a possible function for circRNAs. It has been shown that some circRNAs contain intron and are localized to the nucleus. These circRNAs can also interact with U1 small nuclear ribonucleoprotein to promote transcription (You et al. 2015).
Long Non-Coding RNAs
ncRNAs longer than 200 nt are classified as long ncRNAs. In a fashion similar to pre- miRNAs IncRNAs are also transcribed by RNA polymerase II and undergo 5' capping and polyadenylation. The human genome encodes for approximately 16,000 lcnRNAs
which give rise to almost 30,000 different transcripts. Despite being considered new additions to the field of RNA biology, the function of some IncRNAs has been known for almost 30 years. One of the IncRNAs whose function has been identified in the early 1990s is IncRNA Xist. Xist plays a central role in X-chromosome inactivation (Brown et al. 1992). Recent studies have demonstrated several new functions for IncRNAs including antiviral response in addition to regulation of development and differentiation (Fatica and Bozzoni 2014; Fortes and Morris 2016). A common mechanism for exerting such functions for IncRNAs is to act as post transcriptional regulators by altering mRNA/protein stability and translation (Yoon, Abdelmohsen, and Gorospe 2013).
Certain IncRNAs interact with chromatin modification proteins to regulate gene expression depending on the type modifying protein complex (Morlando et al. 2014; Marchese and Huarte 2014). In terms of regulation of epigenetic mechanisms IncRNAs can also interact with methyltransferases to induce transcriptional repression (Schmitz et al. 2010).
Another way of gene regulation by IncRNAs is achieved via interaction with transcription factors. One of the best studied examples of this, is the binding of GAS5 IncRNA to glucocorticoid receptor (GR). This binding impairs GR’s interaction with glucocorticoid response elements (GRE) and suppresses the expression of GRE- containing genes (Kino et al. 2010).