**1. Introduction**

Cancer is an extremely complex disorder characterized genetically, epigenetically, and histologically by highly heterogeneous proliferative cellular subpopulations, including cancer

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

stem cells (CSCs) and progenies. These cells harbor chromosomal abnormalities, alterations of suppressor genes (TSG) and oncogenes, and aberrant transcriptomic profiles generated by genetic and epigenetic alterations [1, 2]. These cancer cells are in close relationship with a tumor microenvironment (TME), composed of immune and nonimmune stromal cells and modified extracellular matrix. Reciprocal interactions between tumor cells and TME are pivotal in cancer progression, allowing remodeling of TME and reprogramming of cancer cells that develop adaptive strategies to adjust their phenotype to unfavorable environmental conditions. Recently, CSCs were implicated in a new paradigm accounting for tumor heterogeneity [3]. CSCs have the property of self-renewal, lack senescence, maintain an undifferentiated state, and proliferate rapidly. These properties are controlled by epigenetic mechanisms that induce changes in gene expression profiling of tumor cells. Opposite to aging cells that increase genomic and chromosomal instability during adulthood, nonaging immortal cells, such as germline, somatic, and cancer stem cells, harbor a genomic instability triggered by unrepaired mutations with either no or only limited number of genomic alterations [4]. Epigenetic abnormalities are early events in cancer progression, resulting from various environmental injuries, and associate heterogeneity of DNA methylation, posttranscriptional modifications of histones, and deregulation of noncoding RNAs (ncRNAs). Global DNA hypomethylation results in chromosomal instability, overexpression of oncogenes, and reactivation of transposable elements (TEs) [5]. Localized (genes promoters) or wide (>1 Mb) DNA hypermethylation initiates repression of TSGs and modification of epigenetic marks through histone alterations, resulting in occurrence of an aberrantly stemlike state of CSCs. These alterations of the genomic methylation during carcinogenesis allow reprogramming of atypical proliferative cells into highly malignant cells characterized by unlimited proliferation, epithelial-mesenchymal transition (EMT), invasion, and prometastatic properties [6].

functions of small ncRNAs are insured by Argonaute (AGO) protein family, which is a very well-conserved master component of RNA silencing complexes in all organisms [17]. The least well-investigated small ncRNAs are the piRNAs, which were first identified in 2006 in mouse and rat germ cells as ncRNAs interacting with PIWI proteins, a subclass of the Argonaute proteins [18–21]. PiRNAs actually constitute the largest and most diverse class of ncRNAs [16]. PiRNAs and PIWI proteins were initially implicated in epigenetic regulation of germline cells and their overexpressions have been more recently observed in various cancers through

Part 1: The PIWI-piRNA Pathway Is an Immune-Like Surveillance Process That Controls...

http://dx.doi.org/10.5772/intechopen.79974

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This review will provide an overview of the PIWI-piRNA pathway, focusing mainly on origin, biochemical properties, biogenesis, functions, and mechanisms of action in germline and somatic tissues. Furthermore, we will discuss emerging implications of piRNAs in carcinogenesis and highlight their potential clinical utilities as diagnostic/prognostic biomarkers and

RNA interference (RNAi) is a widely conserved small-RNA-mediated gene-silencing mechanism involved in crucial homeostatic events of most eukaryotes [22–24]. Small regulatory RNAs of 20–32 nt, such as endogenous siRNAs, miRNAs, and piRNAs, modulate transcriptional and posttranscriptional repression through complementary RNA or DNA recognition by interacting with well-conserved proteins of 95 kDa belonging to the Argonaute family that cleave their targets [25, 27]. AGO proteins include a PAZ domain [P-element-induced wimpy testis (PIWI)-AGO-Zwille] located in the N terminal region, which binds small ncRNAs and a PIWI domain in the C-terminal region that functions as double-strand-specific RNA endonuclease [28]. Based on sequence homology and functional domains in different species, the AGO family of proteins is divided into three subfamilies: AGO proteins (homologous to *Arabidopsis thaliana* AGO1), PIWI proteins (homologous to *Drosophila melanogaster* Piwi), and WAGO (worm-specific Argonaute clade). The number of AGO family proteins varies considerably between species. Eight different proteins were present in humans, whereas 27 proteins were identified in *C. elegans* and only one protein was observed in fission yeast [29]. AGO proteins interact functionally with siRNAs and miRNAs, which are small single-stranded RNAs of 20–22 nt in length processed in a Dicer-dependent manner from double-stranded precursors, to induce posttranscriptional gene silencing in the cytoplasm [30–32]. Conversely, PIWI proteins are implicated in biogenesis of piRNAs and in their main function through transcriptionally and posttranscriptionally repressing TEs in the nucleus and the cytoplasm [33]. PIWI proteins and piRNAs edify ribonucleoproteins named PiRNA-induced silencing complexes (pi-RISCs). Pi-RISCs specificity is determined by piRNA sequence, whereas Argonaute PIWI protein mediates its effector function. PiRNAs associate with PIWI proteins and guide piRISCs to recognize complementary targets and achieve RNA silencing at transcriptional and posttranscriptional levels (**Figure 1**). Cytoplasmic PIWI-piRNA complexes silence their targets posttranscriptionally *via* piRNA-directed cleavage and the "ping-pong" amplification cycle, whereas nuclear PIWI proteins and piRNAs silence gene transcriptionally through epigenetic changes, including DNA methylation, implementation of H3K9me3

repressive marks, interactions with Mael and HP1 proteins, and repression of Pol II.

aberrant DNA methylation.

**2. The PIWI-piRNA pathway**

therapeutic tools.

Until recently, RNAs were considered as epigenetic regulators and mediators of gene expression, functioning as intermediates of translation in the flow of genetic information from DNA to proteins [7]. Large-scale genomic technologies have provided an astonishing insight into human genome and transcriptome. Next-generation sequencing techniques combined with bioinformatics have revealed that more than 50% of mammalian genomes were composed of TEs and that more than 98% of the human genome was actively transcribed [8]. However, only 1.1% of the genome encodes proteins, and a majority of genes are noncoding RNAs (ncRNAs) [9]. NcRNAs play pivotal roles in developmental and homeostatic processes, and their alterations are implicated in the pathogenesis of many diseases, by modulating expression of numerous genes at epigenetic, transcriptional, and posttranscriptional levels [10]. Most importantly, ncRNAs are frequently deregulated in cancer and have crucial roles in tumor initiation, progression, and metastatic spread. NcRNAs are classified into housekeeper ncRNAs (rRNAs, tRNAs, and snoRNAs) and regulatory ncRNAs. Regulatory ncRNAs are divided into several subfamilies, depending on their size, biogenesis, and biological functions. Small ncRNAs are composed of transcripts shorter than 200 nucleotides (nt), whereas long noncoding RNAs (lncRNAs) comprise transcripts longer than 200 nt [11]. Small ncRNAs also differ by their precursor structure and their mechanisms of biogenesis. They comprise microRNAs (miRNAs), small interfering RNAs (siRNAs), and PIWI-interacting RNA (piRNAs) [12–15].

MiRNAs and siRNAs are generated from double-stranded precursors, whereas piRNAs are processed from long single-stranded precursors. The endoribonuclease Dicer is pivotal in the maturation of miRNAs and siRNAs, but not in the piRNAs processing [16]. Regulatory functions of small ncRNAs are insured by Argonaute (AGO) protein family, which is a very well-conserved master component of RNA silencing complexes in all organisms [17]. The least well-investigated small ncRNAs are the piRNAs, which were first identified in 2006 in mouse and rat germ cells as ncRNAs interacting with PIWI proteins, a subclass of the Argonaute proteins [18–21]. PiRNAs actually constitute the largest and most diverse class of ncRNAs [16]. PiRNAs and PIWI proteins were initially implicated in epigenetic regulation of germline cells and their overexpressions have been more recently observed in various cancers through aberrant DNA methylation.

This review will provide an overview of the PIWI-piRNA pathway, focusing mainly on origin, biochemical properties, biogenesis, functions, and mechanisms of action in germline and somatic tissues. Furthermore, we will discuss emerging implications of piRNAs in carcinogenesis and highlight their potential clinical utilities as diagnostic/prognostic biomarkers and therapeutic tools.
