DOCSLIB.ORG

Structural Basis for Pirna-Targeting

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Structural Basis for Pirna-Targeting bioRxiv preprint doi: https://doi.org/10.1101/2020.12.07.413112; this version posted December 7, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC 4.0 International license. Structural basis for piRNA-targeting Todd A. Anzelon1†, Saikat Chowdhury1,2†, Siobhan M. Hughes1†, Yao Xiao1, Gabriel C. Lander1, Ian J. MacRae1* †These authors contributed equally to this work. *Correspondence to: [email protected] (I.J.M.) 1Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA 2Current address: Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, NY 11794, USA 1 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.07.413112; this version posted December 7, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC 4.0 International license. Summary Piwi proteins use PIWI-interacting RNAs (piRNAs) to identify and silence the transposable elements (TEs) pervasively found in animal genomes. The Piwi targeting mechanism is proposed to be similar to targeting by Argonaute proteins, which employ microRNA (miRNA) guides to repress cellular mRNAs, but has not been characterized in detail. We present cryo-EM structures of a Piwi-piRNA complex with and without target RNAs and analysis of target recognition. Resembling Argonaute, Piwi identifies targets using the piRNA seed-region. However, Piwi creates a much weaker seed so that prolonged target association requires further piRNA-target pairing. Beyond the seed, Piwi creates wide central cleft wide for unencumbered piRNA-target pairing, enabling long-lived Piwi-piRNA-target interactions that are tolerant of mismatches. Piwi ensures targeting fidelity by blocking propagation of the piRNA-target duplex in the absence of faithful seed pairing, and by requiring extended piRNA-target pairing to reach an endonucleolytically active conformation. This mechanism allows Piwi to minimize off-targeting cellular mRNAs and adapt piRNA sequences to evolving genomic threats. Introduction PIWI-interacting RNAs (piRNAs) are the largest class of small (21-35 nt.) regulatory RNAs in animals, with tens of thousands to millions of unique piRNA species expressed in an individual organism (Wang, Zhang et al. 2019). piRNAs have diverse functions in animal biology, the most deeply conserved of which is preventing mobilization of transposable elements (TEs) (Ozata, Gainetdinov et al. 2019). piRNAs are ubiquitous throughout animal germlines, as failure to prevent TE-induced genome instability leads to partial or complete sterility (Rubin, Kidwell et al. 1982, Lin and Spradling 1997, Vagin, Sigova et al. 2006, Aravin, Sachidanandam et al. 2007, Brennecke, Aravin et al. 2007, Houwing, Kamminga et al. 2007, Kuramochi-Miyagawa, Watanabe et al. 2008, Khurana, Wang et al. 2011, Reuter, Berninger et al. 2011). Similarly, piRNA-associated factors are pervasive features of pluripotent stem cells in animals capable of extensive regeneration (Reddien, Oviedo et al. 2005, Rinkevich, Rosner et al. 2010, Zhu, Pao et al. 2012, Juliano, Reich et al. 2014). A wide variety of additional piRNA functions have been described, including acting as agents for passing epigenetic information from mother to embryo in fruit flies (Brennecke, Malone et al. 2008), determining gender in silkworms (Kiuchi, Koga et al. 2014), neutralizing viruses in mosquitos (Miesen, Joosten et al. 2016), and establishing long-term memories in sea slugs (Rajasethupathy, Antonov et al. 2012). However, functions for the majority of piRNA molecules found in most animals, including humans, are not known. On the molecular level, piRNAs function through association with PIWI proteins (Aravin, Gaidatzis et al. 2006, Girard, Sachidanandam et al. 2006, Grivna, Beyret et al. 2006, Lau, Seto et al. 2006, Vagin, Sigova et al. 2006). PIWIs use piRNAs as guides to identify complementary sequences in RNAs targeted for silencing via a variety of mechanisms. PIWI proteins within the nucleus can direct histone/DNA methylation enzymes to silence genetic loci expressing transcripts recognized as piRNA targets (Aravin, Sachidanandam et al. 2008, Kuramochi- Miyagawa, Watanabe et al. 2008, Sienski, Donertas et al. 2012, Le Thomas, Rogers et al. 2013), while in the cytoplasm PIWI proteins have been proposed to recruit deadenylases to degrade messenger RNAs (mRNA) targeted by piRNAs (Rouget, Papin et al. 2010, Gou, Dai et al. 2014). Most PIWI proteins also use an intrinsic 2 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.07.413112; this version posted December 7, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC 4.0 International license. endonuclease activity to cleave target RNAs (Gunawardane, Saito et al. 2007, Reuter, Berninger et al. 2011), which can subsequently be further fragmented to generate new piRNAs. Such cleavage events lead to piRNA amplification and expansion of the cellular piRNA pool to incorporate additional sequences within target RNA molecules (Brennecke, Aravin et al. 2007, Gunawardane, Saito et al. 2007, Han, Wang et al. 2015, Homolka, Pandey et al. 2015, Mohn, Handler et al. 2015, Gainetdinov, Colpan et al. 2018). Therefore, piRNA-targeting, the mechanism by which the PIWI-piRNA complex identifies targets, shapes the cellular piRNA repertoire and profoundly impacts gene silencing. Currently, small RNA targeting mechanisms are best understood for mammalian Argonaute2 (Ago2). Ago2 uses microRNAs (miRNAs), a second major class of animal small RNAs, to identify mRNA targets to be repressed as a normal part of health and development (Bartel 2018). Ago2 facilitates target recognition by pre- ordering the guide (g) nucleotides g2-g7 (referred to as the canonical “seed” region) of the miRNA, into a helical conformation and thereby lowers the entropic cost of target binding (Filipowicz 2005, Parker, Roe et al. 2005, Tomari and Zamore 2005, Wang, Sheng et al. 2008, Parker, Parizotto et al. 2009, Nakanishi, Weinberg et al. 2012, Schirle and MacRae 2012, Wee, Flores-Jasso et al. 2012, Nakanishi, Ascano et al. 2013, Park, Phan et al. 2017, Park, Araya-Secchi et al. 2019). Ago2 further shapes miRNA-target interactions by creating a central cleft that includes two chambers with space for target pairing to miRNA seed and supplementary (g13-g17) nucleotides, and two narrow regions, which discourage pairing to the miRNA central (g9–g12) and 3' tail (g18– g22) nucleotides (Schirle, Sheu-Gruttadauria et al. 2014, Sheu-Gruttadauria, Xiao et al. 2019). piRNAs are proposed to use a miRNA-like targeting mechanism (Gou, Dai et al. 2014, Shen, Chen et al. 2018). Supporting this notion, seed complementarity is a recurrent feature of piRNA target transcripts identified in vivo (Goh, Falciatori et al. 2015, Zhang, Kang et al. 2015, Shen, Chen et al. 2018, Zhang, Tu et al. 2018, Halbach, Miesen et al. 2020). Additionally, PIWI and Argonaute (AGO) family proteins share a common set of domains and have extensive sequence similarities (Carmell, Xuan et al. 2002). On the other hand, crystal structures of Drosophila Piwi and silkworm Siwi have notable differences in the three-dimensional arrangement of domains compared to known AGO structures, indicating that piRNA-target interactions may be shaped in a distinct manner (Matsumoto, Nishimasu et al. 2016, Yamaguchi, Oe et al. 2020). It has not been possible to distinguish between these possibilities and directly measure piRNA-target interactions because a homogenous source of Piwi-piRNA complex suitable for biochemical and structural analysis has not been available. Here, we describe a recombinant source of Piwi-piRNA complex, derived from the piwi-a gene of the freshwater sponge Ephydatia fluviatilis. The protein (EfPiwi) can be produced in a baculovirus expression system and loaded with a synthetic piRNA, resulting in a stable RNA-protein complex that can be purified to homogeneity. We determined cryo-electron microscopy (cryo-EM) structures of the EfPiwi-piRNA complex with and without target RNAs and performed biochemical analyses of target recognition. Like mammalian Ago2, EfPiwi initially engages targets through seed pairing. However, EfPiwi creates a much weaker seed than Ago2 such that further base pairing is required for high-affinity target interactions. EfPiwi enforces targeting fidelity by extensively probing the piRNA-target duplex in the seed region and employing a structural element, termed the “seed-gate”, to block duplex propagation in the absence of faithful seed pairing. Beyond the seed-gate, EfPiwi 3 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.07.413112; this version posted December 7, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC 4.0 International license. creates central cleft wide enough for unencumbered piRNA-target pairing, enabling EfPiwi-piRNA-target interactions that are long-lived and tolerant of mismatches. Target cleavage, however, requires an extended piRNA-target duplex of sufficient length and rigidity to drive EfPiwi into a fully open conformation and activate endonuclease activity. We propose the overall targeting mechanism allows Piwi-piRNA complexes to accumulate on and/or destroy target transcripts a manner that is sufficiently specific to avoid off-targeting cellular RNAs yet malleable enough to adapt to new and evolving threats to the genome. Recombinant Source of homogenous PIWI-piRNA complexes PIWI proteins derived from natural sources co-purify with heterogenous mixtures of endogenous piRNAs (Aravin, Gaidatzis et al. 2006, Girard, Sachidanandam et al. 2006, Grivna, Beyret et al. 2006, Lau, Seto et al. 2006, Vagin, Sigova et al.
Load more

Recommended publications
  • Epigenetic Roles of PIWI‑Interacting Rnas (Pirnas) in Cancer Metastasis (Review)
    ONCOLOGY REPORTS 40: 2423-2434, 2018 Epigenetic roles of PIWI‑interacting RNAs (piRNAs) in cancer metastasis (Review) JIA LIU1, SHUJUN ZHANG2 and BINGLIN CHENG1 1Department of Integrated Traditional Chinese and Western Medicine Oncology, The First Affiliated Hospital of Harbin Medical University; 2Department of Pathology, The Fourth Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150000, P.R. China Received March 19, 2018; Accepted September 3, 2018 DOI: 10.3892/or.2018.6684 Abstract. P-element-induced wimpy testis (PIWI)-interacting 7. Epigenetics of ncRNAs in cancer RNAs (piRNAs) are epigenetic-related short ncRNAs that 8. Discussion participate in chromatin regulation, transposon silencing, and modification of specific gene sites. These epigenetic factors or alterations are also involved in the growth of a variety 1. Introduction of human cancers, including lung, breast, and colon cancer. Accumulating evidence has revealed that tumor metastasis P-element-induced wimpy testis (PIWI)-interacting RNAs and invasion involve genetic and epigenetic factors. Cancer (piRNAs) belong to a new class of ncRNAs that have been asso- metastasis is characterized by epigenetic alterations including ciated with many cancers (1). piRNAs are involved in the gene DNA methylation and histone modification. Changes in DNA regulation process in which certain nucleotides bind coding methylation, H3K9me3 heterochromatin and transposable regions in gene promoters (2). piRNAs function in the epigen- elements have been detected in several cancers. piRNAs may etic regulation of DNA methylation (3), transposable silencing function in gene silencing and gene modification upstream and chromatin modification (4). PIWI is a type of Argonaute or downstream of oncogenes in cancer cell lines or cancer protein that binds to piRNAs and carries out unique functions tissues.
    [Show full text]
  • Piwi-Interacting Rnas and PIWI Genes As Novel Prognostic Markers for Breast Cancer
    www.impactjournals.com/oncotarget/ Oncotarget, Vol. 7, No. 25 Research Paper Piwi-interacting RNAs and PIWI genes as novel prognostic markers for breast cancer Preethi Krishnan1, Sunita Ghosh2,3, Kathryn Graham2,3, John R. Mackey2,3, Olga Kovalchuk4, Sambasivarao Damaraju1,3 1Department of Laboratory Medicine and Pathology, University of Alberta, Edmonton, Alberta, Canada 2Department of Oncology, University of Alberta, Edmonton, Alberta, Canada 3Cross Cancer Institute, Alberta Health Services, Edmonton, Alberta, Canada 4Department of Biological Sciences, University of Lethbridge, Lethbridge, Alberta, Canada Correspondence to: Sambasivarao Damaraju, email: [email protected] Keywords: piRNA, PIWI, breast cancer, prognostic marker, TCGA Received: January 13, 2016 Accepted: April 28, 2016 Published: May 10, 2016 ABSTRACT Piwi-interacting RNAs (piRNAs), whose role in germline maintenance has been established, are now also being classified as post-transcriptional regulators of gene expression in somatic cells. PIWI proteins, central to piRNA biogenesis, have been identified as genetic and epigenetic regulators of gene expression. piRNAs/PIWIs have emerged as potential biomarkers for cancer but their relevance to breast cancer has not been comprehensively studied. piRNAs and mRNAs were profiled from normal and breast tumor tissues using next generation sequencing and Agilent platforms, respectively. Gene targets for differentially expressed piRNAs were identified from mRNA expression dataset. piRNAs and PIWI genes were independently assessed for their prognostic significance (outcomes: Overall Survival, OS and Recurrence Free Survival, RFS). We discovered eight piRNAs as novel independent prognostic markers and their association with OS was confirmed in an external dataset (The Cancer Genome Atlas). Further, PIWIL3 and PIWIL4 genes showed prognostic relevance. 306 gene targets exhibited reciprocal relationship with piRNA expression.
    [Show full text]
  • The Other Face of Piwi Plant Gene Editing Improved
    RESEARCH HIGHLIGHTS NON-CODING RNA The other face of PIWI Spermiogenesis involves gradual with 3ʹUTRs of the target mRNAs; Credit: S. Bradbrook/Springer Nature Limited chromatin compaction and trans- reporter protein levels but not cription shut-down. mRNAs that mRNA levels increased, implicating activation. Translation of hundreds are transcribed in spermatocytes translation in reporter activation. of mRNAs co-targeted by piRNA and early-round spermatids are Activation of the target-mRNA and HuR was dependent on MIWI, stored as translationally inactive reporters required piRNA–3ʹUTR indicating that they are direct targets ribonucleoproteins until later during base-pairing and 3ʹUTR binding by of this selective mechanism of spermiogenesis, when their trans- functional MIWI. Screening for translation activation. lation is activated, but how this MIWI-interacting proteins revealed The proteins encoded by activation occurs is largely unknown. that eukaryotic translation initiation two of the five original target PIWI proteins and PIWI-interacting factor 3f (eIF3f) directly interacted mRNAs are essential for sperm RNAs (piRNAs) are essential for with MIWI and was also required MIWI– acrosome formation. Indeed, gametogenesis as they suppress for reporter activation. The activated piRNAs severe acrosome defects were found the expression of transposons and 3ʹUTRs included AU-rich elements … in MIWI-depleted spermatids mRNAs. Dai et al. now show that (AREs) that are bound by HuR, interact with owing to considerable decrease in mouse PIWI (MIWI)–piRNAs which is an RNA-binding protein eIF3f–HuR and the levels of the two proteins. are the core of a complex required known to interact with another other proteins Thus, although MIWI–piRNAs for selective mRNA translation translation factor, eIF4G3, for trans- are mostly known for gene silencing, in spermatids.
    [Show full text]
  • The PIWI Protein Aubergine Recruits Eif3 to Activate Translation in the Germ Plasm
    www.nature.com/cr www.cell-research.com ARTICLE OPEN The PIWI protein Aubergine recruits eIF3 to activate translation in the germ plasm Anne Ramat1, Maria-Rosa Garcia-Silva1, Camille Jahan1, Rima Naït-Saïdi1, Jérémy Dufourt 1,5, Céline Garret1, Aymeric Chartier1, Julie Cremaschi1, Vipul Patel1, Mathilde Decourcelle 2, Amandine Bastide3, François Juge 4 and Martine Simonelig 1 Piwi-interacting RNAs (piRNAs) and PIWI proteins are essential in germ cells to repress transposons and regulate mRNAs. In Drosophila, piRNAs bound to the PIWI protein Aubergine (Aub) are transferred maternally to the embryo and regulate maternal mRNA stability through two opposite roles. They target mRNAs by incomplete base pairing, leading to their destabilization in the soma and stabilization in the germ plasm. Here, we report a function of Aub in translation. Aub is required for translational activation of nanos mRNA, a key determinant of the germ plasm. Aub physically interacts with the poly(A)-binding protein (PABP) and the translation initiation factor eIF3. Polysome gradient profiling reveals the role of Aub at the initiation step of translation. In the germ plasm, PABP and eIF3d assemble in foci that surround Aub-containing germ granules, and Aub acts with eIF3d to promote nanos translation. These results identify translational activation as a new mode of mRNA regulation by Aub, highlighting the versatility of PIWI proteins in mRNA regulation. Cell Research (2020) 30:421–435; https://doi.org/10.1038/s41422-020-0294-9 1234567890();,: INTRODUCTION directly interacts with the Oskar (Osk) protein that is specifically Translational control is a widespread mechanism to regulate synthesized at the posterior pole of oocytes and embryos and gene expression in many biological contexts.
    [Show full text]
  • Argonaute Proteins at a Glance Christine Ender and Gunter Meister
    Cell Science at a Glance 1819 Argonaute proteins at a RNAs) – ncRNAs that are characteristically Biogenesis of miRNAs and siRNAs ~20-35 nucleotides long – are required for miRNAs glance the regulation of gene expression in many Generally, miRNAs are transcribed by RNA different organisms. Most small RNA species polymerase II or III to form stem-loop-structured Christine Ender1 and Gunter fall into one of the following three classes: primary miRNA transcripts (pri-miRNAs). pri- Meister1,2,* microRNAs (miRNAs), short-interfering RNAs miRNAs are processed in the nucleus by the 1 Center for Integrated Protein Science Munich (siRNAs) and Piwi-interacting RNAs (piRNAs) microprocessor complex, which contains (CIPSM), Laboratory of RNA Biology, Max-Planck- Institute of Biochemistry, Am Klopferspitz 18, 82152 (Carthew and Sontheimer, 2009). Although the RNAse III enzyme Drosha and its DiGeorge Martinsried, Germany different small RNA classes have different syndrome critical region gene 8 (DGCR8) 2University of Regensburg, Universitätsstraße 31, 93053 Regensburg, Germany bio genesis pathways and exert different cofactor. The transcripts are cleaved at the stem *Author for correspondence functions, all of them must associate with a of the hairpin to produce a stem-loop- ([email protected]) member of the Argonaute protein family for structured miRNA precursor (pre-miRNA) of Journal of Cell Science 123, 1819-1823 activity. This article and its accompanying ~70 nucleotides. After this first processing step, © 2010. Published by The Company of Biologists Ltd poster provide an overview of the different pre-miRNAs are exported into the cytoplasm by doi:10.1242/jcs.055210 classes of small RNAs and the manner in which exportin-5, where they are further processed they interact with Argonaute protein family by the RNAse III enzyme Dicer and its TRBP Although a large portion of the human genome members during small-RNA-guided gene (HIV transactivating response RNA-binding is actively transcribed into RNA, less than 2% silencing.
    [Show full text]
  • Genetic Insight Into the Domain Structure and Functions of Dicer-Type Ribonucleases
    International Journal of Molecular Sciences Review Genetic Insight into the Domain Structure and Functions of Dicer-Type Ribonucleases Kinga Ciechanowska, Maria Pokornowska and Anna Kurzy ´nska-Kokorniak* Department of Ribonucleoprotein Biochemistry, Institute of Bioorganic Chemistry Polish Academy of Sciences, Noskowskiego 12/14, 61-704 Poznan, Poland; [email protected] (K.C.); [email protected] (M.P.) * Correspondence: [email protected]; Tel.: +48-61-852-85-03 (ext. 1264) Abstract: Ribonuclease Dicer belongs to the family of RNase III endoribonucleases, the enzymes that specifically hydrolyze phosphodiester bonds found in double-stranded regions of RNAs. Dicer enzymes are mostly known for their essential role in the biogenesis of small regulatory RNAs. A typical Dicer-type RNase consists of a helicase domain, a domain of unknown function (DUF283), a PAZ (Piwi-Argonaute-Zwille) domain, two RNase III domains, and a double-stranded RNA binding domain; however, the domain composition of Dicers varies among species. Dicer and its homologues developed only in eukaryotes; nevertheless, the two enzymatic domains of Dicer, helicase and RNase III, display high sequence similarity to their prokaryotic orthologs. Evolutionary studies indicate that a combination of the helicase and RNase III domains in a single protein is a eukaryotic signature and is supposed to be one of the critical events that triggered the consolidation of the eukaryotic RNA interference. In this review, we provide the genetic insight into the domain organization and structure of Dicer proteins found in vertebrate and invertebrate animals, plants and fungi. We also discuss, in the context of the individual domains, domain deletion variants and partner proteins, a variety of Dicers’ functions not only related to small RNA biogenesis pathways.
    [Show full text]
  • Function of Piwi, a Nuclear Piwi/Argonaute Protein, Is Independent of Its Slicer Activity
    Function of Piwi, a nuclear Piwi/Argonaute protein, is independent of its slicer activity Nicole Darricarrère, Na Liu, Toshiaki Watanabe, and Haifan Lin1 Yale Stem Cell Center and Department of Cell Biology, Yale University School of Medicine, New Haven, CT 06509 Edited by James A. Birchler, University of Missouri, Columbia, MO, and approved December 11, 2012 (received for review August 1, 2012) The Piwi protein subfamily is essential for Piwi-interacting RNA classical epigenetic phenotypes in Drosophila (15, 17, 18). These (piRNA) biogenesis, transposon silencing, and germ-line develop- observations led us to propose that piRNA molecules can guide ment, all of which have been proposed to require Piwi endonuclease Piwi to specific genomic loci by sequence complementarity, activity, as validated for two cytoplasmic Piwi proteins in mice. where Piwi recruits epigenetic modifiers (17–19). In this epige- However, recent evidence has led to questioning of the generality netic model, the slicer activity of Piwi seems superfluous. Indeed, of this mechanism for the Piwi members that reside in the nucleus. the involvement of the slicer activity of Piwi has been questioned Drosophila offers a distinct opportunity to study the function of recently based on experiments performed in an ovarian somatic nuclear Piwi proteins because, among three Drosophila Piwi pro- cell line (20). However, this analysis was not definitive. It was teins—called Piwi, Aubergine, and Argonaute 3—Piwi is the only based on a single cultured cell type, relied on RNAi that only member of this subfamily that is localized in the nucleus and incompletely reduced endogenous levels of Piwi and tested only expressed in both germ-line and somatic cells in the gonad, where one transposon element.
    [Show full text]
  • Aberrant Expression of Select Pirna-Pathway Genes Does Not
    Aberrant expression of select piRNA-pathway genes BRIEF REPORT does not reactivate piRNA silencing in cancer cells Pavol Genzora, Seth C. Cordtsa, Neha V. Bokila, and Astrid D. Haasea,1 aNational Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892 Edited by Brigid L. M. Hogan, Duke University Medical Center, Durham, NC, and approved April 30, 2019 (received for review March 15, 2019) Germline genes that are aberrantly expressed in nongermline the slicer activity of PIWIL2 (HILI) and the RNA helicase DDX4/ cancer cells have the potential to be ideal targets for diagnosis and VASA (13) (Fig. 1E). We did not observe aberrant expression of therapy due to their restricted physiological expression, their DDX4/VASA in any nongermline cancer (Fig. 1 B–D), eliminating the broad reactivation in various cancer types, and their immunogenic possibility of reactivated ping-pong processing, and thus focused our properties. Among such cancer/testis genes, components of the analysis on potential PLD6/ZUC-dependent piRNA production. Be- PIWI-interacting small RNA (piRNA) pathway are of particular inter- cause of high sequence diversity and lack of sequence conservation, est, as they control mobile genetic elements (transposons) in germ piRNAs are best defined by their interaction with PIWI proteins, cells and thus hold great potential to counteract genome instability according to their initial definition as PIWI-interacting RNAs (11, 14). in cancer. Here, we systematically investigate the potential reac- Echoing the phenomenon of cancer/germline gene-expression in pri- mary tumor samples [The Cancer Genome Atlas (TCGA)], some tivation of functional piRNA-silencing mechanisms in the aberrant cancer cell lines exhibit aberrant expression of piRNA-pathway com- context.
    [Show full text]
  • PIWI Proteins and Cancer Survival: a Meta-Analysis
    bioRxiv preprint doi: https://doi.org/10.1101/663468; this version posted June 14, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. PIWI proteins as prognostic markers in cancer: a systematic review and meta-analysis Running title: PIWI proteins and cancer survival: a meta-analysis Alexios-Fotios A. Mentis1,2, #, Efthimios Dardiotis3, #, Athanassios G. Papavassiliou4, * 1 Public Health Laboratories, Hellenic Pasteur Institute, Athens, Greece 2 Department of Microbiology, University Hospital of Thessaly, Larissa, Greece 3 Department of Neurology, University Hospital of Thessaly, Larissa, Greece 4 Department of Biological Chemistry, National and Kapodistrian University of Athens Medical School, Athens, Greece # Equal contribution *Corresponding author: Prof. Athanasios G. Papavassiliou, M.D., Ph.D. Professor, Department of Biological Chemistry National and Kapodistrian University of Athens Medical School 115 21 Athens, Greece Telephone: +30-210-7462508; E-mail: [email protected] Financial support: None Conflict of Interest: None Word count: 6,166 words Total number of figures: 8 (and 4 supplementary figures) Total number of Tables: 3 (and 1 supplementary table) bioRxiv preprint doi: https://doi.org/10.1101/663468; this version posted June 14, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. ABSTRACT Background: PIWI proteins, which interact with piRNAs, are implicated in stem cell and germ cell regulation, but have been detected in various cancers, as well. Objectives: In this systematic review, we explored, for the first time in the literature (to our knowledge), the association between prognosis in patients with cancer and intratumoral expression of PIWI proteins.
    [Show full text]
  • Title Comprehensive Classification of the Rnase H-Like Domain-Containing Proteins in Plants
    bioRxiv preprint doi: https://doi.org/10.1101/572842; this version posted January 13, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Title Comprehensive Classification of the RNase H-like domain-containing Proteins in Plants Shuai Li, Kunpeng Liu, and Qianwen Sun# Tsinghua-Peking Joint Center for Life Sciences and Center for Plant Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China # Corresponding author Qianwen Sun: [email protected]. Abstract Background R-loop is a nucleic acid structure containing an RNA-DNA hybrid and a displaced single-stranded DNA. Recently, accumulated evidence showed that R-loops widely present in various organisms’ genomes and are involved in many physiological processes, including DNA replication, RNA transcription, and DNA repair. RNase H-like superfamily (RNHLS) domain-containing proteins, such as RNase H enzymes, are essential in restricting R-loop levels. However, little is known about the function and relationship of other RNHLS proteins on R-loop regulation, especially in plants. Results In this study, we characterized 6193 RNHLS proteins from 13 representative plant species and clustered these proteins into 27 clusters, among which reverse transcriptases and exonucleases are the two largest groups. Moreover, we found 691 RNHLS proteins in Arabidopsis with a conserved catalytic alpha-helix and beta-sheet motif. Interestingly, each of the Arabidopsis RNHLS proteins is composed of not only an RNHLS domain but also another different protein domain.
    [Show full text]
  • Abundant Primary Pirnas, Endo-Sirnas, and Micrornas in a Drosophila Ovary Cell Line
    Downloaded from genome.cshlp.org on October 5, 2021 - Published by Cold Spring Harbor Laboratory Press Letter Abundant primary piRNAs, endo-siRNAs, and microRNAs in a Drosophila ovary cell line Nelson C. Lau,1,2,6,7,8 Nicolas Robine,3,6 Raquel Martin,3 Wei-Jen Chung,3 Yuzo Niki,4 Eugene Berezikov,5 and Eric C. Lai3,8 1Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts 02114, USA; 2Department of Genetics, Harvard Medical School. Boston, Massachusetts 02115, USA; 3Department of Developmental Biology, Sloan-Kettering Institute, New York, New York 10065, USA; 4Department of Sciences, Ibaraki University, Mito 310-8512, Japan; 5Hubrecht Institute and University Medical Center Utrecht, Uppsalalaan 8, 3584CT Utrecht, The Netherlands Piwi proteins, a subclass of Argonaute-family proteins, carry ;24–30-nt Piwi-interacting RNAs (piRNAs) that mediate gonadal defense against transposable elements (TEs). We analyzed the Drosophila ovary somatic sheet (OSS) cell line and found that it expresses miRNAs, endogenous small interfering RNAs (endo-siRNAs), and piRNAs in abundance. In contrast to intact gonads, which contain mixtures of germline and somatic cell types that express different Piwi-class proteins, OSS cells are a homogenous somatic cell population that expresses only PIWI and primary piRNAs. Detailed examination of its TE-derived piRNAs and endo-siRNAs revealed aspects of TE defense that do not rely upon ping-pong amplification. In particular, we provide evidence that a subset of piRNA master clusters, including flamenco, are specifically expressed in OSS and ovarian follicle cells. These data indicate that the restriction of certain TEs in somatic gonadal cells is largely mediated by a primary piRNA pathway.
    [Show full text]
  • S Nap S Hot: S Ma Ll
    SnapShot: Small RNA-Mediated RNA-Mediated SnapShot: Small Modifications Epigenetic Carthew and Richard Bei, Sigal Pressman, Yanxia Evanston, IL 60208, USA Northwestern University, Molecular Biology and Cell Biology, Department of Biochemistry, 756 Cell 130, August 24, 2007 ©2007 Elsevier Inc. DOI 10.1016/j.cell.2007.08.015 See online version for legend, abbreviations, and references. SnapShot: Small RNA-Mediated Epigenetic Modifications Yanxia Bei, Sigal Pressman, and Richard Carthew Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, IL 60208, USA (A) Heterochromatin Formation Although heterochromatin formation occurs in the nucleus, it is not clear in which cellular compartment the events leading up to small RNA-directed heterochromatin formation occur. A1: Heterochromatin Formation in Schizosaccharomyces pombe. DNA repeats produce double-stranded (ds)RNAs through bidirectional transcription or RNA- dependent RNA synthesis. dsRNAs are cut into small-interfering (si)RNAs that are loaded into an RNA-induced transcriptional silencing complex (RITS) that consists of Ago, Tas3, an S. pombe specific protein, and Chp1, a chromodomain containing protein. RITS finds the DNA repeats through siRNA base pairing with the nascent transcript and recruits the RNA-directed RNA polymerase complex (RDRC) and Clr4, a histone methyltransferase that methylates histone H3 at lysine 9 (H3K9me). RdRP in RDRC uses the Ago-cut nascent RNA as template to synthesize more dsRNA, which in turn will be cut into siRNAs to reinforce heterochromatin formation. Chp1 in the RITS complex binds to H3K9me, resulting in stable interaction of RITS and heterochromatic DNA. H3K9me also binds to another chromodomain protein, Swi6, an HP1 homolog, leading to the spreading of heterochromatin.
    [Show full text]