Degradation dynamics of microRNAs revealed by a novel pulse-chase approach

The regulation of miRNAs is critical to the definition of cell identity and behavior in normal physiology and disease. To date, the dynamics of miRNA degradation and the mechanisms involved in remain largely obscure, in particular, in higher organisms. Here, we developed a pulse-chase approach based on metabolic RNA labeling to calculate miRNA decay rates at genome-wide scale in mammalian cells. Our analysis revealed heterogeneous miRNA half-lives, with many species behaving as stable molecules (T1/2> 24 h), while others, including passenger miRNAs and a number (25/129) of guide miRNAs, are quickly turned over (T1/2= 4-14 h). Decay rates were coupled with other features, including genomic organization, transcription rates, structural heterogeneity (isomiRs), and target abundance, measured through quantitative experimental approaches. This comprehensive analysis highlighted functional mechanisms that mediate miRNA degradation, as well as the importance of decay dynamics in the regulation of the miRNA pool under both steady-state conditions and during cell transitions.

[1]  M. Selbach,et al.  Global quantification of mammalian gene expression control , 2011, Nature.

[2]  R. Gregory,et al.  MicroRNA biogenesis pathways in cancer , 2015, Nature Reviews Cancer.

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

[4]  S. Lowe,et al.  The microcosmos of cancer , 2012, Nature.

[5]  S. Tavazoie,et al.  Control of metastatic progression by microRNA regulatory networks , 2013, Nature Cell Biology.

[6]  D. Bartel.  MicroRNAs: Target Recognition and Regulatory Functions , 2009, Cell.

[7]  W. Filipowicz,et al.  The widespread regulation of microRNA biogenesis, function and decay , 2010, Nature Reviews Genetics.

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

[9]  M. Magnasco,et al.  Decay rates of human mRNAs: correlation with functional characteristics and sequence attributes. , 2003, Genome research.

[10]  L. Zon,et al.  Selective microRNA uridylation by Zcchc6 (TUT7) and Zcchc11 (TUT4) , 2014, Nucleic acids research.

[11]  E. Sontheimer,et al.  Origins and Mechanisms of miRNAs and siRNAs , 2009, Cell.

[12]  Jian-Rong Yang,et al.  MicroRNA-26a/b and their host genes cooperate to inhibit the G1/S transition by activating the pRb protein , 2011, Nucleic acids research.

[13]  D. Haber,et al.  Dual Role for Argonautes in MicroRNA Processing and Posttranscriptional Regulation of MicroRNA Expression , 2007, Cell.

[14]  M. Malumbres,et al.  Multiple E2F-Induced MicroRNAs Prevent Replicative Stress in Response to Mitogenic Signaling , 2010, Molecular and Cellular Biology.

[15]  R. Sachidanandam,et al.  High-throughput assessment of microRNA activity and function using microRNA sensor and decoy libraries , 2012, Nature Methods.

[16]  C. Bracken,et al.  IsomiRs--the overlooked repertoire in the dynamic microRNAome. , 2012, Trends in genetics : TIG.

[17]  Xiaoxia Qi,et al.  Control of Stress-Dependent Cardiac Growth and Gene Expression by a MicroRNA , 2007, Science.

[18]  B. Cullen,et al.  Differential RISC association of endogenous human microRNAs predicts their inhibitory potential , 2014, Nucleic acids research.

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

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

[21]  I. MacRae,et al.  Highly complementary target RNAs promote release of guide RNAs from human Argonaute2. , 2013, Molecular cell.

[22]  Heiko Muller,et al.  IsomiRage: From Functional Classification to Differential Expression of miRNA Isoforms , 2014, Front. Bioeng. Biotechnol..

[23]  Caroline C. Friedel,et al.  Conserved principles of mammalian transcriptional regulation revealed by RNA half-life , 2009, Nucleic acids research.

[24]  H. Grosshans,et al.  MicroRNA turnover: when, how, and why. , 2012, Trends in biochemical sciences.

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

[26]  Saikat Chakraborty,et al.  Pri-miR-17-92a transcript folds into a tertiary structure and autoregulates its processing. , 2012, RNA.

[27]  D. Patel,et al.  Adenylation of maternally inherited microRNAs by Wispy. , 2014, Molecular cell.

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

[29]  Phillip A Sharp,et al.  Endogenous miRNA and target concentrations determine susceptibility to potential ceRNA competition. , 2014, Molecular cell.

[30]  A. D’Ambrogio,et al.  CPEB and two poly(A) polymerases control miR-122 stability and p53 mRNA translation , 2011, Nature.

[31]  Xuemei Chen,et al.  Degradation of microRNAs by a Family of Exoribonucleases in Arabidopsis , 2008, Science.

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

[33]  S. Wyman,et al.  Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity. , 2011, Genome research.

[34]  H. Grosshans,et al.  Active turnover modulates mature microRNA activity in Caenorhabditis elegans , 2009, Nature.

[35]  F. Camargo,et al.  Hippo Signaling Regulates Microprocessor and Links Cell-Density-Dependent miRNA Biogenesis to Cancer , 2014, Cell.

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

[37]  J. Boothroyd,et al.  Biosynthetic labeling of RNA with uracil phosphoribosyltransferase allows cell-specific microarray analysis of mRNA synthesis and decay , 2005, Nature Biotechnology.

[38]  E. Izaurralde,et al.  Getting to the Root of miRNA-Mediated Gene Silencing , 2008, Cell.

[39]  L. Zon,et al.  Selective microRNA uridylation by Zcchc 6 ( TUT 7 ) and Zcchc 11 ( TUT 4 ) , 2014 .

[40]  V. Kim,et al.  Modifications of Small RNAs and Their Associated Proteins , 2010, Cell.

[41]  Grace X. Y. Zheng,et al.  MicroRNAs can generate thresholds in target gene expression , 2011, Nature Genetics.

[42]  S. Cohen,et al.  microRNA functions. , 2007, Annual review of cell and developmental biology.

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

[44]  R. Zimmer,et al.  High-resolution gene expression profiling for simultaneous kinetic parameter analysis of RNA synthesis and decay. , 2008, RNA.

[45]  Haifan Lin,et al.  MicroRNAs: key regulators of stem cells , 2009, Nature Reviews Molecular Cell Biology.

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

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

[48]  E. Wentzel,et al.  Cell–cell contact globally activates microRNA biogenesis , 2009, Proceedings of the National Academy of Sciences.

[49]  N. Friedman,et al.  Metabolic labeling of RNA uncovers principles of RNA production and degradation dynamics in mammalian cells , 2011, Nature Biotechnology.

[50]  L. Dölken,et al.  4-thiouridine inhibits rRNA synthesis and causes a nucleolar stress response , 2013, RNA biology.

[51]  Marco J. Morelli,et al.  INSPEcT: a computational tool to infer mRNA synthesis, processing and degradation dynamics from RNA- and 4sU-seq time course experiments , 2015, Bioinform..

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

[53]  V. Kim,et al.  Regulation of microRNA biogenesis , 2014, Nature Reviews Molecular Cell Biology.

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