Non-coding RNAs have diverse functions in eukaryotic cells. Use of these non-coding RNAs in therapeutic approaches is a promising but rather unexplored direction in biomedical research. We discovered a new class of small non-coding RNAs, piwi-interacting RNAs (piRNAs), that together with their protein partners, Piwi proteins, recognize and silence endogenous genomic parasites called transposable elements. The silencing of transposons is critical in germline cells and the failure of piRNA-mediated repression leads to sterility in both Drosophila and mice. The mechanism of biogenesis of piRNAs appears to be distinct from that of other classes of small non-coding RNAs, microRNA and siRNA. piRNAs are encoded in distinct genomic regions dubbed piRNA clusters that work as memory banks to store inactive copies of past transposon invaders. piRNA clusters produce long ncRNA transcripts, pre-piRNAs, that are further processed to mature piRNAs, which work as guides to recognize and repress transposon sequences. We showed that piRNA clusters have a unique chromatin signature that is essential for their expression and identified a protein complex that directly recognizes these chromatin marks. We further show that this complex is directly involved in transcription of pre-piRNA precursors and that transcription of clusters is unusual in a number of ways. Now we want to capitalize on our findings and understand how transcription and early steps of piRNA biogenesis are regulated. To meet these objectives we will dissect the molecular mechanism of piRNA biogenesis using Drosophila melanogaster. We will explore initiation and termination of pre-piRNA transcription using the combination of a new generation of transcriptome analysis tools as well as transgenic and genome editing approaches. We will also test the role of two complexes, Rhino/Cutoff and TREX, in transcription and processing of pre-piRNA. Our studies will help to advance our understanding of the mechanism of TE silencing, which is important for both fertility and for genomic stability in somatic cells. It will also provide the basis for futur use of the piRNA pathway as a tool in research and therapy. Importantly, the significance of the proposed research extends well beyond answering important questions in the non-coding RNA field. Our studies will provide clues to how co-transcriptional sorting of different types of RNAs into distinct processing pathways operates. It explores fundamental mechanisms that control transcription and early post- transcriptional processes.

Public Health Relevance

The aim of this application is to understand the molecular mechanism of the piRNA pathway that recognizes and silences genomic parasites known as transposable elements. Transposon repression is critical in germline cells, where the failure of piRNA-mediated repression leads to sterility, but transposon repression also might be important in aging and cancer progression. Investigation of the molecular mechanism of the piRNA pathway is of great importance to our understanding of transposon control in health and disease and will provide the basis for directing piRNAs to new targets for use in research and therapy.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM097363-06
Application #
9230395
Study Section
Molecular Genetics A Study Section (MGA)
Program Officer
Bender, Michael T
Project Start
2011-06-06
Project End
2019-12-31
Budget Start
2017-01-01
Budget End
2017-12-31
Support Year
6
Fiscal Year
2017
Total Cost
Indirect Cost
Name
California Institute of Technology
Department
Type
Schools of Arts and Sciences
DUNS #
009584210
City
Pasadena
State
CA
Country
United States
Zip Code
91125
Chen, Yung-Chia Ariel; Stuwe, Evelyn; Luo, Yicheng et al. (2016) Cutoff Suppresses RNA Polymerase II Termination to Ensure Expression of piRNA Precursors. Mol Cell 63:97-109
Hur, Junho K; Luo, Yicheng; Moon, Sungjin et al. (2016) Splicing-independent loading of TREX on nascent RNA is required for efficient expression of dual-strand piRNA clusters in Drosophila. Genes Dev 30:840-55
Webster, Alexandre; Li, Sisi; Hur, Junho K et al. (2015) Aub and Ago3 Are Recruited to Nuage through Two Mechanisms to Form a Ping-Pong Complex Assembled by Krimper. Mol Cell 59:564-75
Chen, Yung-Chia Ariel; Aravin, Alexei A (2015) Non-Coding RNAs in Transcriptional Regulation: The review for Current Molecular Biology Reports. Curr Mol Biol Rep 1:10-18
Le Thomas, Adrien; Stuwe, Evelyn; Li, Sisi et al. (2014) Transgenerationally inherited piRNAs trigger piRNA biogenesis by changing the chromatin of piRNA clusters and inducing precursor processing. Genes Dev 28:1667-80
Hur, Junho K; Olovnikov, Ivan; Aravin, Alexei A (2014) Prokaryotic Argonautes defend genomes against invasive DNA. Trends Biochem Sci 39:257-9
Pezic, Dubravka; Manakov, Sergei A; Sachidanandam, Ravi et al. (2014) piRNA pathway targets active LINE1 elements to establish the repressive H3K9me3 mark in germ cells. Genes Dev 28:1410-28
Le Thomas, Adrien; Tóth, Katalin Fejes; Aravin, Alexei A (2014) To be or not to be a piRNA: genomic origin and processing of piRNAs. Genome Biol 15:204
Olovnikov, Ivan; Le Thomas, Adrien; Aravin, Alexei A (2014) A framework for piRNA cluster manipulation. Methods Mol Biol 1093:47-58
Stuwe, Evelyn; Tóth, Katalin Fejes; Aravin, Alexei A (2014) Small but sturdy: small RNAs in cellular memory and epigenetics. Genes Dev 28:423-31

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