A Network of Noncoding Regulatory RNAs Acts in the Mammalian Brain

Noncoding RNAs (ncRNAs) play increasingly appreciated gene-regulatory roles. Here, we describe a regulatory network centered on four ncRNAs-a long ncRNA, a circular RNA, and two microRNAs-using gene editing in mice to probe the molecular consequences of disrupting key components of this network. The long ncRNA Cyrano uses an extensively paired site to miR-7 to trigger destruction of this microRNA. Cyrano-directed miR-7 degradation is much more effective than previously described examples of target-directed microRNA degradation, which come primarily from studies of artificial and viral RNAs. By reducing miR-7 levels, Cyrano prevents repression of miR-7-targeted mRNAs and enables accumulation of Cdr1as, a circular RNA known to regulate neuronal activity. Without Cyrano, excess miR-7 causes cytoplasmic destruction of Cdr1as in neurons, in part through enhanced slicing of Cdr1as by a second miRNA, miR-671. Thus, several types of ncRNAs can collaborate to establish a sophisticated regulatory network.

[1]  Loyal A Goff,et al.  Differential regulation of microRNA stability. , 2010, RNA.

[2]  Anton J. Enright,et al.  MicroRNA degradation by a conserved target RNA regulates animal behavior , 2018, Nature Structural & Molecular Biology.

[3]  A. Oudenaarden,et al.  Single-molecule mRNA detection and counting in mammalian tissue , 2013, Nature Protocols.

[4]  Daehyun Baek,et al.  Selective degradation of host MicroRNAs by an intergenic HCMV noncoding RNA accelerates virus production. , 2013, Cell host & microbe.

[5]  Chao Zhang,et al.  Growth of the developing cerebral cortex is controlled by microRNA-7 through the p53 pathway. , 2014, Cell reports.

[6]  D. Bartel,et al.  MicroRNA destabilization enables dynamic regulation of the miR-16 family in response to cell-cycle changes. , 2011, Molecular cell.

[7]  Jun Liu,et al.  Characterization of the mammalian miRNA turnover landscape , 2015, Nucleic acids research.

[8]  M. Stoffel,et al.  Loss of microRNA-7a2 induces hypogonadotropic hypogonadism and infertility , 2017, The Journal of clinical investigation.

[9]  Kevin R. Parker,et al.  ciRS-7 exonic sequence is embedded in a long non-coding RNA locus , 2017, bioRxiv.

[10]  D. Tollervey,et al.  Murine cytomegalovirus encodes a miR-27 inhibitor disguised as a target , 2011, Proceedings of the National Academy of Sciences.

[11]  J. Kjems,et al.  Natural RNA circles function as efficient microRNA sponges , 2013, Nature.

[12]  Karl Mechtler,et al.  Molecular basis for cytoplasmic RNA surveillance by uridylation‐triggered decay in Drosophila , 2016, The EMBO journal.

[13]  Jyothi Arikkath,et al.  Culturing pyramidal neurons from the early postnatal mouse hippocampus and cortex , 2012, Nature Protocols.

[14]  Ravi Sachidanandam,et al.  Kinetic Analysis Reveals the Fate of a MicroRNA following Target Regulation in Mammalian Cells , 2011, Current Biology.

[15]  Stefan L Ameres,et al.  Target RNA-directed tailing and trimming purifies the sorting of endo-siRNAs between the two Drosophila Argonaute proteins. , 2011, RNA.

[16]  Michael B. Stadler,et al.  Characterizing Light-Regulated Retinal MicroRNAs Reveals Rapid Turnover as a Common Property of Neuronal MicroRNAs , 2010, Cell.

[17]  Allan R. Jones,et al.  Genome-wide atlas of gene expression in the adult mouse brain , 2007, Nature.

[18]  L. Lim,et al.  MicroRNA targeting specificity in mammals: determinants beyond seed pairing. , 2007, Molecular cell.

[19]  Daehyun Baek,et al.  mRNA destabilization is the dominant effect of mammalian microRNAs by the time substantial repression ensues. , 2014, Molecular cell.

[20]  Carmen Birchmeier,et al.  Loss of a mammalian circular RNA locus causes miRNA deregulation and affects brain function , 2017, Science.

[21]  Thomas R. Gingeras,et al.  STAR: ultrafast universal RNA-seq aligner , 2013, Bioinform..

[22]  Xuemei Chen,et al.  Regulation of small RNA stability: methylation and beyond , 2012, Cell Research.

[23]  J. Richter,et al.  Opposing polymerase-deadenylase activities regulate cytoplasmic polyadenylation. , 2006, Molecular cell.

[24]  M. Greenberg,et al.  A functional screen implicates microRNA-138-dependent regulation of the depalmitoylation enzyme APT1 in dendritic spine morphogenesis , 2009, Nature Cell Biology.

[25]  Michael B. Stadler,et al.  Potent degradation of neuronal miRNAs induced by highly complementary targets , 2015, EMBO reports.

[26]  Sanjay Tyagi,et al.  Single molecule imaging of RNA in situ. , 2011, Methods in molecular biology.

[27]  T. Katoh,et al.  Destabilization of microRNAs in human cells by 3′ deadenylation mediated by PARN and CUGBP1 , 2015, Nucleic acids research.

[28]  Piero Carninci,et al.  PAPD5-mediated 3′ adenylation and subsequent degradation of miR-21 is disrupted in proliferative disease , 2014, Proceedings of the National Academy of Sciences.

[29]  Philippe Soriano,et al.  Context-specific requirements for Fgfr1 signaling through Frs2 and Frs3 during mouse development , 2006, Development.

[30]  Luke A. Gilbert,et al.  CRISPR-Mediated Modular RNA-Guided Regulation of Transcription in Eukaryotes , 2013, Cell.

[31]  D. Bartel,et al.  Predicting effective microRNA target sites in mammalian mRNAs , 2015, eLife.

[32]  E. Izaurralde,et al.  Towards a molecular understanding of microRNA-mediated gene silencing , 2015, Nature Reviews Genetics.

[33]  Ola Snøve,et al.  Distance constraints between microRNA target sites dictate efficacy and cooperativity , 2007, Nucleic acids research.

[34]  D. Bartel.  Metazoan MicroRNAs , 2018, Cell.

[35]  D. Bartel,et al.  Conserved Function of lincRNAs in Vertebrate Embryonic Development despite Rapid Sequence Evolution , 2011, Cell.

[36]  W. Huber,et al.  Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2 , 2014, Genome Biology.

[37]  Stefan L Ameres,et al.  Long-term, efficient inhibition of microRNA function in mice using rAAV vectors , 2012, Nature Methods.

[38]  Paul J. Hertzog,et al.  Analysis of microRNA turnover in mammalian cells following Dicer1 ablation , 2011, Nucleic acids research.

[39]  D. Bartel,et al.  The Menu of Features that Define Primary MicroRNAs and Enable De Novo Design of MicroRNA Genes. , 2015, Molecular cell.

[40]  Paul Theodor Pyl,et al.  HTSeq—a Python framework to work with high-throughput sequencing data , 2014, bioRxiv.

[41]  J. Mendell,et al.  Functional Classification and Experimental Dissection of Long Noncoding RNAs , 2018, Cell.

[42]  S. Pfeffer,et al.  Degradation of Cellular miR-27 by a Novel, Highly Abundant Viral Transcript Is Important for Efficient Virus Replication In Vivo , 2012, PLoS pathogens.

[43]  Marcel Martin.  Cutadapt removes adapter sequences from high-throughput sequencing reads , 2011 .

[44]  R. Gregory,et al.  A role for the Perlman syndrome exonuclease Dis3l2 in the Lin28-let-7 pathway , 2013, Nature.

[45]  S. Pfeffer,et al.  Identification of factors involved in target RNA-directed microRNA degradation , 2016, Nucleic acids research.

[46]  J. Steitz,et al.  Four novel U RNAs are encoded by a herpesvirus , 1988, Cell.

[47]  Zhiping Weng,et al.  Target RNA–Directed Trimming and Tailing of Small Silencing RNAs , 2010, Science.

[48]  C. Betsholtz,et al.  Reverse genetic screening reveals poor correlation between morpholino-induced and mutant phenotypes in zebrafish. , 2015, Developmental cell.

[49]  J. Kjems,et al.  Insights into circular RNA biology , 2017, RNA biology.

[50]  Mark B Gerstein,et al.  Tracking Distinct RNA Populations Using Efficient and Reversible Covalent Chemistry. , 2015, Molecular cell.

[51]  S. Gujja,et al.  Gld2-catalyzed 3′ monoadenylation of miRNAs in the hippocampus has no detectable effect on their stability or on animal behavior , 2016, RNA.

[52]  J. Steitz,et al.  Down-Regulation of a Host MicroRNA by a Herpesvirus saimiri Noncoding RNA , 2010, Science.

[53]  T. Katoh,et al.  Selective stabilization of mammalian microRNAs by 3' adenylation mediated by the cytoplasmic poly(A) polymerase GLD-2. , 2009, Genes & development.

[54]  Jørgen Kjems,et al.  miRNA‐dependent gene silencing involving Ago2‐mediated cleavage of a circular antisense RNA , 2011, The EMBO journal.

[55]  Neil R. Smalheiser,et al.  Expression of microRNAs and Other Small RNAs in Prefrontal Cortex in Schizophrenia, Bipolar Disorder and Depressed Subjects , 2014, PloS one.

[56]  Eran Segal,et al.  EGF Decreases the Abundance of MicroRNAs That Restrain Oncogenic Transcription Factors , 2010, Science Signaling.

[57]  K. Pollard,et al.  Detection of nonneutral substitution rates on mammalian phylogenies. , 2010, Genome research.

[58]  M. Zavolan,et al.  MicroRNA-7a regulates pancreatic β cell function. , 2014, The Journal of clinical investigation.

[59]  E. Wentzel,et al.  A Hexanucleotide Element Directs MicroRNA Nuclear Import , 2007, Science.

[60]  Sebastian D. Mackowiak,et al.  Circular RNAs are a large class of animal RNAs with regulatory potency , 2013, Nature.

[61]  Vikram Agarwal,et al.  Impact of MicroRNA Levels, Target-Site Complementarity, and Cooperativity on Competing Endogenous RNA-Regulated Gene Expression , 2016, Molecular cell.

[62]  D. Haussler,et al.  Evolutionarily conserved elements in vertebrate, insect, worm, and yeast genomes. , 2005, Genome research.

[63]  Gregor Eichele,et al.  GenePaint.org: an atlas of gene expression patterns in the mouse embryo , 2004, Nucleic Acids Res..

[64]  Helga Thorvaldsdóttir,et al.  Integrative Genomics Viewer (IGV): high-performance genomics data visualization and exploration , 2012, Briefings Bioinform..

[65]  Jerilyn A. Walker,et al.  Preparation of PCR-quality mouse genomic DNA with hot sodium hydroxide and tris (HotSHOT). , 2000, BioTechniques.

[66]  T. Tuschl,et al.  Functional anatomy of siRNAs for mediating efficient RNAi in Drosophila melanogaster embryo lysate , 2001, The EMBO journal.

[67]  Max A. Horlbeck,et al.  Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation , 2014, Cell.

[68]  L. Greene,et al.  Isolation and Culture of Post-Natal Mouse Cerebellar Granule Neuron Progenitor Cells and Neurons , 2009, Journal of visualized experiments : JoVE.

[69]  Shan Zong,et al.  Comprehensive Identification of Long Non-coding RNAs in Purified Cell Types from the Brain Reveals Functional LncRNA in OPC Fate Determination , 2015, PLoS genetics.