Small non‐coding RNA and colorectal cancer

Abstract Colorectal cancer (CRC) is the third most common malignance. Although great efforts have been made to understand the pathogenesis of CRC, the underlying mechanisms are still unclear. It is now clear that more than 90% of the total genome is actively transcribed, but lack of protein‐coding potential. The massive amount of RNA can be classified as housekeeping RNAs (such as ribosomal RNAs, transfer RNAs) and regulatory RNAs (such as microRNAs [miRNAs], PIWI‐interacting RNA [piRNAs], tRNA‐derived stress‐induced RNA, tRNA‐derived small RNA [tRFs] and long non‐coding RNAs [lncRNAs]). Small non‐coding RNAs are a group of ncRNAs with the length no more than 200 nt and they have been found to exert important regulatory functions under many pathological conditions. In this review, we summarize the biogenesis and functions of regulatory sncRNAs, such as miRNAs, piRNA and tRFs, and highlight their involvements in cancers, particularly in CRC.


| INTRODUC TI ON
Colorectal cancer (CRC) is the third most common malignance and is one leading cause of cancer-related deaths in the world. 1 Despite great efforts that have been made to understand the pathogenesis of CRC, the underlying mechanisms are still largely unknown.
Increasing evidence has demonstrated that CRC is a heterogeneous disease, and its pathogenesis is involved with the activation of oncogenes and inactivation of tumour-suppressive genes, which are mostly resulting from genetic mutations and epigenetic alterations, the latter including DNA methylation, histone modification and noncoding RNAs (ncRNAs). [2][3][4][5][6][7] Large-scale genome sequencing has indicated that the human genome encodes approximately 20 000 protein-coding transcripts, which account for only around 2% of the genome, while more than 90% of the total genome is actively transcribed, but lack of protein-coding potential, referred to as ncRNAs. 8,9 For several decades, ncRNAs were considered as 'evolutionary junk'. However, more and more evidence has shown that a part of ncRNAs are functional RNA molecules. 10 These functional RNA transcripts are composed by housekeeping ncRNAs, including highly abundant ribosomal RNAs (rRNAs) and transfer RNAs (tRNAs), as well as regulatory ncRNAs, such as microRNAs (miRNAs), PIWI-interacting RNA (piRNAs), tRNA-derived small RNA (tRFs), small nucleolar RNA (snoRNAs), siR-NAs, long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs).
In addition, according to the length of ncRNAs, they are divided into two subclasses: small or short non-coding RNAs (sncRNAs,  nt) and lncRNAs (>200 nt).
Long non-coding RNAs are a subgroup of non-coding RNAs, with more than 200 nucleotides in length and no protein coding potential.
Long non-coding RNAs have tissue-specific expression and exert regulatory functions in many biological and pathological processes.
They can function as both tumour suppressors and promoters in CRC development. 11 For example, H19 is an imprinted oncofoetal ncRNA, but it is hypomethylated and thus up-regulated in CRC.
H19 can promote the development of CRC via generating miRNA or by serving as ceRNA. 12 Down-regulation of lncRNA MEG3 can promote colorectal adenocarcinoma cell proliferation and inhibit the apoptosis by up-regulating TGF-β1 and its downstream sphingosine kinase 1. 13  can regulate the proliferation and apoptosis of CRC cells as miRNA sponge. 15 Circ-ZNF609 promotes migration of CRC by inhibiting Gli1 expression via microRNA-150. 16 Recent studies have revealed that regulatory sncRNAs (miR-NAs, piRNAs, tRFs and snoRNAs) can also function as important regulators in gene expression, and play crucial roles in many physiological and pathological processes. And the abnormal expression of these sncRNAs is involved in many human diseases, including cancers. 17 In this review, we provide an overview of the representative classes of sncRNAs, including miRNAs, piRNAs and tRFs, and summarize their involvements in CRC, focusing on their roles in the initiation and development of CRC, as well as their biomarker potential (Table 1).

| MICRORNA S
MicroRNAs are a class of sncRNAs containing approximately 18-25 nucleotides, highly conserved and present in eukaryotic cells. In general, the majority of miRNA-coding genes are located in intergenic or intragenic regions. The generation of mature miR-NAs is a multi-step process that starts in the nucleus and ends in the cytoplasm. Canonically, most miRNAs are transcribed as large mono-or polycistronic primary miRNA precursors (pri-miRNAs) in the nucleus by RNA polymerase II (Pol. II), while other pri-miRNAs are transcribed by RNA Pol. III. All the pri-miRNAs contain a 5′ cap and a poly A tail at the 3′ untranslated region (UTR). Then, pri-miR-NAs are processed to generate miRNA precursors (pre-miRNAs) by RNase III Drosha in complex with DGCR8, a RNA-binding protein functioning as a ruler to measure cleavage point. The pre-miRNAs are subsequently recognized and exported to the cytoplasm by Exportin-5 (Exp5). In the cytosol, the pre-miRNAs are cleaved by the RNase III endonuclease Dicer to yield a miRNA duplex, which contains a leading strand or miR and a passenger strand or miR*. The tissues and blood derived from CRC patients, of which, miR-23a, miR-193a-3p and miR-338-5p were significantly up-regulated in CRC tissues, and the levels of these miRNAs are positively related between CRC tissue and blood. 35 Meanwhile, an exosomal miRNA profile was used to highlight the most abundant miRNAs in the blood of 88 patients with CRC. 36 As well as circulating miRNAs, some stool-based miRNAs were also observed to be dysregulated and they also can be used as a biomarker for CRC. For example, lung cancer cell proliferation and the mechanism is associated with its binding to 3′ UTR of mTOR messenger RNA (mRNA). 97 piR-32051, piR-39894 and piR-43607 were found up-regulated in kidney cancer tissues. 98 PIWI-interacting RNA DQ594040 is one down-regulated piRNA in bladder cancer, and its overexpression can inhibit bladder cancer cell proliferation, colony formation and promote cell apoptosis through up-regulating TNFSF4 protein. 99 As for CRC, piR-651 was found to be up-regulated in tumour tissues, which is associated with metastasis state. piR59056, piR-54878 and piR-62701 are also highly expressed in CRC, and are associated with recurrence-free survival. 91 piR-823 contributes to colorectal tumorgenesis by enhancing the transcriptional activity of HSF1. 100 109 The production of tRF-5s and tRF-3s has reported to be dependent on Dicer. 109 However, the specific process of generating tRF-5s and tRF-3s is still not clear.
Meanwhile, the biogenesis mechanism of tRF-1s is also unknown.
Recently, tRFs have been found to play an important role in many biological and pathological processes, and they are associated with many diseases, such as virus infection, metabolic disorder, neurodegenerative diseases and cancers. 17,109,110 To date, the regulatory mechanisms of tRFs mainly involve several strategies as follows.

| CON CLUS ION
Although great achievements have been made in understanding the pathogenesis of CRC, the underlying mechanisms are still needed F I G U R E 2 The types of tsRNAs are classified by size and sequence location in the tRNA structure. Based on the length and cleavage sites of tRNAs, small non-coding RNA derived from tRNAs can be classified into two major groups: tRNA-derived stress-induced RNA (tiRNAs), with the length of 28-36 nt, and tRNA-derived fragment (tRFs), about 14-30 nt length for further investigations. For decades, many studies have provided a lot of evidence that sncRNAs are important regulatory molecules and play a crucial role in cancer development and progression. So, it is believed that sncRNAs are involved in the pathogenesis of CRC.
Notably, many miRNAs have been demonstrated to be dysregulated in CRC, and they may exert important functions in the development of CRC. Furthermore, they have been found to act as potential diagnostic biomarkers for CRC. In addition, some piRNAs and tRFs have also been proven to be implicated in the development of CRC.
However, studies on them are still at a very early stage, and many questions still need to be addressed. In conclusion, considering sn-cRNAs as important regulators in CRC, we need to shed light on many aspects of their biogenesis and functions in CRC.

ACK N OWLED G EM ENTS
This work is supported by the fundament of People's Hospital of Taizhou.

CO N FLI C T O F I NTE R E S T
The authors declare no conflict of interest.