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Review
, 886, 51-77

The piRNA Pathway Guards the Germline Genome Against Transposable Elements

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Review

The piRNA Pathway Guards the Germline Genome Against Transposable Elements

Katalin Fejes Tóth et al. Adv Exp Med Biol.

Abstract

Transposable elements (TEs) have the capacity to replicate and insert into new genomic locations. This contributs significantly to evolution of genomes, but can also result in DNA breaks and illegitimate recombination, and therefore poses a significant threat to genomic integrity. Excess damage to the germ cell genome results in sterility. A specific RNA silencing pathway, termed the piRNA pathway operates in germ cells of animals to control TE activity. At the core of the piRNA pathway is a ribonucleoprotein complex consisting of a small RNA, called piRNA, and a protein from the PIWI subfamily of Argonaute nucleases. The piRNA pathway relies on the specificity provided by the piRNA sequence to recognize complementary TE targets, while effector functions are provided by the PIWI protein. PIWI-piRNA complexes silence TEs both at the transcriptional level - by attracting repressive chromatin modifications to genomic targets - and at the posttranscriptional level - by cleaving TE transcripts in the cytoplasm. Impairment of the piRNA pathway leads to overexpression of TEs, significantly compromised genome structure and, invariably, germ cell death and sterility.The piRNA pathway is best understood in the fruit fly, Drosophila melanogaster, and in mouse. This Chapter gives an overview of current knowledge on piRNA biogenesis, and mechanistic details of both transcriptional and posttranscriptional TE silencing by the piRNA pathway. It further focuses on the importance of post-translational modifications and subcellular localization of the piRNA machinery. Finally, it provides a brief description of analogous pathways in other systems.

Keywords: Argonautes; DNA methylation; Germ granules; H3K9me3; Heterochromatin; Intramitochondrial cement; Nuage; Pi-bodies; Ping-Pong cycle; Piwi proteins; Pole plasm; Posttranscriptional silencing; Small RNA; TE; Transcriptional silencing; Transposable element; Transposon; Tudor domain; piRNA.

Figures

Fig. 4.1
Fig. 4.1
The domain structure of PlWI-clade argonaute proteins, (a) The domains of PIWI proteins are organized in a similar fashion to other Argonaute proteins, comprised of the N-terminal region, PAZ (Piwi-Argonaute-Zwille), mid and PIWI domains. The N-terminal region consists of a notional domain that is characterized by arginine rich motifs that are targeted for methylation and in the case of PIWI in flies and MIWI2 in mouse contains the NLS signal, (b) The organization of protein domains in space shows how the mid-domain anchors the piRNA at its 5′ end and the PAZ domain holds the 3′ end of the piRNA. PIWI is the largest domain and its catalytic site responsible for ‘slicer’ activity is positioned to cleave the backbone of annealed target RNA exactly 10 nt relative to the 5′ end of the piRNA
Fig. 4.2
Fig. 4.2
The structure of piRNA clusters in Drosophila ovaries and mouse spermatogenic cells, (a) Clusters in Drosophila are divided into two groups based on their structure and they show differential expression in the two cell types of the ovary. Unidirectional clusters are expressed in follicular cells, which are the somatic support cells of the ovary. These clusters produce piRNAs from only one strand of their genomic locus, and are loaded into PIWI. Bidirectional clusters are expressed in germline cells of the ovary, and piRNAs from these clusters are preferentially loaded into AUB and PIWI. (b) Mouse spermatogenic cells contain unidirectional as well as divergent clusters. Divergent clusters are transcribed from a central promoter into both directions. Putative transcription start sites are shown with red arrows. piRNA density profiles derived from the + and − strands are indicated in grey and blue as they would appear in small RNA-seq data
Fig. 4.3
Fig. 4.3
Biogenesis of piRNAs in flies begins in the nucleus and initiates the ping-ping cycle in the cytoplasm. (a) Precursor transcripts originating from piRNA clusters that contain anti-sense TE sequences are exported from the nucleus and processed into smaller fragments, possibly by the mitochondrial-associated endonuclease, Zucchini. (b) AUB binds precursor piRNA fragments with a preference for fragments that contain a 5′ terminal uracil (1U). Mature primary piRNAs are generated when an unknown 3′ ≥ 5′ exonuclease trims the precursor piRNA fragment bound to AUB and HEN1 catalyzes the 2-O methylation of the piRNA 3′ end. (c) When mRNA of active transposons is exported from the nucleus, AUB piRNA anneals to complementary sequences within the mRNA and cleaves the transcript. (d) The cleaved transcript is loaded into AGO3, followed by trimming and 2′ O-methylation of its 3′ end, to generate mature secondary piRNA. (e) AGO3 loaded with the secondary piRNA is believed to target and cleave newly generated precursor transcripts that are loaded into AUB to initiate a new round of Ping-Pong
Fig. 4.4
Fig. 4.4
The nuclear function of the piRNA pathway in flies. PIWI loaded with cluster-derived primary piRNAs enters the nucleus and scans nascent transcripts for complementarity to its piRNA. If a transposon RNA transcript is encountered, PIWI is believed to recruit chromatin factors such as histone methyl-transferases (HMT) to deposit repressive H3K9-methyl marks. The modifications induced by PIWI binding create sites for the assembly of additional factors that repress transcription of transposon loci
Fig. 4.5
Fig. 4.5
Many protein factors make up nuage granules in Drosophila nurse cells and some accumulate at the pole plasm of the developing oocyte. (a) Nuage granules surround the nucleus of nurse cells and consist of many factors involved in the piRNA pathway. Nuage also shows close proximity to mitochondria, which contain piRNA pathway components (ZUC, GASZ) in their outer membrane. Nuage integrity depends on nuclear piRNA biogenesis factors (such as RHI, CUFF, UAP56) and a number of nuage components of unknown function. Many nuage components contain Tudor domains, which are receptors for symmetrically di-methylated arginines (sDMA). These Tudor proteins are believed to form a scaffold for the factors involved in the pathway. AUB and AGO3 contain sDMA motifs at their N-terminal regions and can interact with Tudor proteins. The enzyme responsible for methylation, PRMT5, consisting of Capsuléen (CSUL) and Valois (VLS), is also present in nuage and associates with Tudor. Some nuclear proteins, such as PIWI and MAEL are believed to transiently visit the nuage and this is required for their function (Klenov et al. 2011; Saito et al. 2009b). Known interactions are indicated. (b) At later stages of oogenesis, nurse cells deposit their cytoplasmic contents, including piRNA pathway components, into the oocyte. Some components, including sDMA-modified AUB, accumulate at the posterior end of oocytes to form the pole plasm. The material that accumulates at the pole plasm becomes the cytoplasmic material of pole cells, which give rise to primordial germ cells of the embryo

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