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Systemic RNAi in C. elegans from the Viewpoint of RNA as Extracellular Signals

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Extracellular Nucleic Acids

Part of the book series: Nucleic Acids and Molecular Biology ((NUCLEIC,volume 25))

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Abstract

The elegantly simple organism Caenorhabditis elegans has played an important role in the RNAi field. In 1997, the first RNAi experiment was performed in this nematode by engineering a specific sequence of double-stranded RNA (dsRNA) and injecting the molecules into the organism. Subsequently, an RNAi response was noted not only in the treated animal but also in its progeny. In later experiments that utilized Green Fluorescent Protein to assess RNAi in individual cells, systemic RNAi phenocopies were observed in most nonneuronal tissues. These early observations of systemic RNAi revealed a hitherto unknown feature of RNA molecules: an ability of locally delivered dsRNAs to traffic to distant somatic and germline cells and to gain entry into those cells – an ability not generally observed for structurally similar DNA molecules. Understanding the precise nature of the RNA molecules that facilitate RNAi as well as the cellular mechanisms that respond to them will provide insights into the unusually robust nature of RNAi in C. elegans as well as highlight potential trafficking routes for endogenous RNAs.

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References

  • Alcazar RM, Lin R, Fire AZ (2008) Transmission dynamics of heritable silencing induced by double-stranded RNA in Caenorhabditis elegans. Genetics 180:1275–1288

    Article  PubMed  CAS  Google Scholar 

  • Ambros V, Lee RC, Lavanway A, Williams PT, Jewell D (2003) MicroRNAs and other tiny endogenous RNAs in C. elegans. Curr Biol 13:807–818

    Article  PubMed  CAS  Google Scholar 

  • Aoki K, Moriguchi H, Yoshioka T, Okawa K, Tabara H (2007) In vitro analyses of the production and activity of secondary small interfering RNAs in C. elegans. EMBO J 26:5007–5019

    Article  PubMed  CAS  Google Scholar 

  • Batista PJ, Ruby JG, Claycomb JM, Chiang R, Fahlgren N, Kasschau KD, Chaves DA, Gu W, Vasale JJ, Duan S, Conte D Jr, Luo S, Schroth GP, Carrington JC, Bartel DP, Mello CC (2008) PRG-1 and 21U-RNAs interact to form the piRNA complex required for fertility in C. elegans. Mol Cell 31:67–78

    Article  PubMed  CAS  Google Scholar 

  • Boisvert ME, Simard MJ (2008) RNAi pathway in C. elegans: the argonautes and collaborators. Curr Top Microbiol Immunol 320:21–36

    Article  PubMed  CAS  Google Scholar 

  • Charier G, Couprie J, Alpha-Bazin B, Meyer V, Quemeneur E, Guerois R, Callebaut I, Gilquin B, Zinn-Justin S (2004) The Tudor tandem of 53BP1: a new structural motif involved in DNA and RG-rich peptide binding. Structure 12:1551–1562

    Article  PubMed  CAS  Google Scholar 

  • Claycomb JM, Batista PJ, Pang KM, Gu W, Vasale JJ, van Wolfswinkel JC, Chaves DA, Shirayama M, Mitani S, Ketting RF, Conte D Jr, Mello CC (2009) The Argonaute CSR-1 and its 22G-RNA cofactors are required for holocentric chromosome segregation. Cell 139:123–134

    Article  PubMed  CAS  Google Scholar 

  • Cote J, Richard S (2005) Tudor domains bind symmetrical dimethylated arginines. J Biol Chem 280:28476–28483

    Article  PubMed  CAS  Google Scholar 

  • Duchaine TF, Wohlschlegel JA, Kennedy S, Bei Y, Conte D Jr, Pang K, Brownell DR, Harding S, Mitani S, Ruvkun G, Yates JR 3rd, Mello CC (2006) Functional proteomics reveals the biochemical niche of C. elegans DCR-1 in multiple small-RNA-mediated pathways. Cell 124:343–354

    Article  PubMed  CAS  Google Scholar 

  • Farazi TA, Juranek SA, Tuschl T (2008) The growing catalog of small RNAs and their association with distinct Argonaute/Piwi family members. Development 135:1201–1214

    Article  PubMed  CAS  Google Scholar 

  • Feinberg EH, Hunter CP (2003) Transport of dsRNA into cells by the transmembrane protein SID-1. Science 301:1545–1547

    Article  PubMed  CAS  Google Scholar 

  • Filipowicz W (2005) RNAi: the nuts and bolts of the RISC machine. Cell 122:17–20

    Article  PubMed  CAS  Google Scholar 

  • Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC (1998) Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391:806–811

    Article  PubMed  CAS  Google Scholar 

  • Fischer SE, Butler MD, Pan Q, Ruvkun G (2008) Trans-splicing in C. elegans generates the negative RNAi regulator ERI-6/7. Nature 455:491–496

    Article  PubMed  CAS  Google Scholar 

  • Fritz JH, Girardin SE, Philpott DJ (2006) Innate immune defense through RNA interference. Sci STKE 2006:pe27

    Google Scholar 

  • Grad Y, Aach J, Hayes GD, Reinhart BJ, Church GM, Ruvkun G, Kim J (2003) Computational and experimental identification of C. elegans microRNAs. Mol Cell 11:1253–1263

    Article  PubMed  CAS  Google Scholar 

  • Grant BD, Caplan S (2008) Mechanisms of EHD/RME-1 protein function in endocytic transport. Traffic 9:2043–2052

    Article  PubMed  CAS  Google Scholar 

  • Greenstein D (2005) Control of oocyte meiotic maturation and fertilization. (December 28, 2005), WormBook, ed. The C. elegans Research Community, WormBook, doi/10.1895/wormbook.1.53.1, http://www.wormbook.org

  • Grishok A (2005) RNAi mechanisms in Caenorhabditis elegans. FEBS Lett 579:5932–5939

    Article  PubMed  CAS  Google Scholar 

  • Grishok A, Mello CC (2002) RNAi (Nematodes: Caenorhabditis elegans). Adv Genet 46:339–360

    Article  PubMed  CAS  Google Scholar 

  • Grishok A, Pasquinelli AE, Conte D, Li N, Parrish S, Ha I, Baillie DL, Fire A, Ruvkun G, Mello CC (2001) Genes and mechanisms related to RNA interference regulate expression of the small temporal RNAs that control C. elegans developmental timing. Cell 106:23–34

    Article  PubMed  CAS  Google Scholar 

  • Gu W, Shirayama M, Conte D Jr, Vasale J, Batista PJ, Claycomb JM, Moresco JJ, Youngman EM, Keys J, Stoltz MJ, Chen CC, Chaves DA, Duan S, Kasschau KD, Falgren N, Yates JR 3rd, Mitani S, Carrington JC, Mello CC (2009) Distinct argonaute-mediated 22G-RNA pathways direct genome surveillance in the C. elegans germline. Mol Cell 36(2):231–244

    Article  PubMed  CAS  Google Scholar 

  • Guang S, Bochner AF, Pavelec DM, Burkhart KB, Harding S, Lachowiec J, Kennedy S (2008) An Argonaute transports siRNAs from the cytoplasm to the nucleus. Science 321:537–541

    Article  PubMed  CAS  Google Scholar 

  • Hall DH, Altun Z (2008) C. elegans atlas. Cold Spring Harbor Laboratory Press, New York

    Google Scholar 

  • Hall DH, Winfrey VP, Blaeuer G, Hoffman LH, Furuta T, Rose KL, Hobert O, Greenstein D (1999) Ultrastructural features of the adult hermaphrodite gonad of Caenorhabditis elegans: relations between the germ line and soma. Dev Biol 212:101–123

    Article  PubMed  CAS  Google Scholar 

  • Hamilton AJ, Baulcombe DC (1999) A species of small antisense RNA in posttranscriptional gene silencing in plants. Science 286:950–952

    Article  PubMed  CAS  Google Scholar 

  • Hammond SM, Bernstein E, Beach D, Hannon GJ (2000) An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells. Nature 404:293–296

    Article  PubMed  CAS  Google Scholar 

  • Han W, Sundaram P, Kenjale H, Grantham J, Timmons L (2008) The Caenorhabditis elegans rsd-2 and rsd-6 genes are required for chromosome functions during exposure to unfavorable environments. Genetics 178:1875–1893

    Article  PubMed  Google Scholar 

  • Hubbard E J, Greenstein D (2005) Introduction to the germline. (September 1, 2005), WormBook, ed. The C. elegans Research Community, WormBook, doi/10.1895/wormbook.1.18.1, http://www.wormbook.org

  • Hull D, Timmons L (2004) Methods for delivery of double-stranded RNA into Caenorhabditis elegans. Methods Mol Biol 265:23–58

    PubMed  CAS  Google Scholar 

  • Hutvagner G, Simard MJ (2008) Argonaute proteins: key players in RNA silencing. Nat Rev Mol Cell Biol 9:22–32

    Article  PubMed  CAS  Google Scholar 

  • Huyen Y, Zgheib O, Ditullio RA Jr, Gorgoulis VG, Zacharatos P, Petty TJ, Sheston EA, Mellert HS, Stavridi ES, Halazonetis TD (2004) Methylated lysine 79 of histone H3 targets 53BP1 to DNA double-strand breaks. Nature 432:406–411

    Article  PubMed  CAS  Google Scholar 

  • Jaskiewicz L, Filipowicz W (2008) Role of Dicer in posttranscriptional RNA silencing. Curr Top Microbiol Immunol 320:77–97

    Article  PubMed  CAS  Google Scholar 

  • Johnstone IL (1994) The cuticle of the nematode Caenorhabditis elegans: a complex collagen structure. Bioessays 16:171–178

    Article  PubMed  CAS  Google Scholar 

  • Jose AM, Smith JJ, Hunter CP (2009) Export of RNA silencing from C. elegans tissues does not require the RNA channel SID-1. Proc Natl Acad Sci U S A 106:2283–2288

    Article  PubMed  CAS  Google Scholar 

  • Joshua-Tor L (2006) The Argonautes. Cold Spring Harb Symp Quant Biol 71:67–72

    Article  PubMed  CAS  Google Scholar 

  • Kennedy S, Wang D, Ruvkun G (2004) A conserved siRNA-degrading RNase negatively regulates RNA interference in C. elegans. Nature 427:645–649

    Article  PubMed  CAS  Google Scholar 

  • Knight SW, Bass BL (2001) A role for the RNase III enzyme DCR-1 in RNA interference and germ line development in Caenorhabditis elegans. Science 293:2269–2271

    Article  PubMed  CAS  Google Scholar 

  • Knight SW, Bass BL (2002) The role of RNA editing by ADARs in RNAi. Mol Cell 10:809–817

    Article  PubMed  CAS  Google Scholar 

  • Lagos-Quintana M, Rauhut R, Lendeckel W, Tuschl T (2001) Identification of novel genes coding for small expressed RNAs. Science 294:853–858

    Article  PubMed  CAS  Google Scholar 

  • Lau NC, Lim LP, Weinstein EG, Bartel DP (2001) An abundant class of tiny RNAs with probable regulatory roles in Caenorhabditis elegans. Science 294:858–862

    Article  PubMed  CAS  Google Scholar 

  • Lee RC, Ambros V (2001) An extensive class of small RNAs in Caenorhabditis elegans. Science 294:862–864

    Article  PubMed  CAS  Google Scholar 

  • Lee RC, Hammell CM, Ambros V (2006) Interacting endogenous and exogenous RNAi pathways in Caenorhabditis elegans. RNA 12:589–597

    Article  PubMed  CAS  Google Scholar 

  • Lehner B, Calixto A, Crombie C, Tischler J, Fortunato A, Chalfie M, Fraser AG (2006) Loss of LIN-35, the Caenorhabditis elegans ortholog of the tumor suppressor p105Rb, results in enhanced RNA interference. Genome Biol 7:R4

    Article  PubMed  Google Scholar 

  • Li Z, Kim SW, Lin Y, Moore PS, Chang Y, John B (2009) Characterization of viral and human RNAs smaller than canonical microRNAs. J Virol 83(24):12751–12758

    Article  PubMed  CAS  Google Scholar 

  • Lim LP, Lau NC, Weinstein EG, Abdelhakim A, Yekta S, Rhoades MW, Burge CB, Bartel DP (2003) The microRNAs of Caenorhabditis elegans. Genes Dev 17:991–1008

    Article  PubMed  CAS  Google Scholar 

  • Liu Z, Zheng Y (2009) A requirement for epsin in mitotic membrane and spindle organization. J Cell Biol 186:473–480

    Article  PubMed  CAS  Google Scholar 

  • Lu R, Maduro M, Li F, Li HW, Broitman-Maduro G, Li WX, Ding SW (2005) Animal virus replication and RNAi-mediated antiviral silencing in Caenorhabditis elegans. Nature 436:1040–1043

    Article  PubMed  CAS  Google Scholar 

  • Montgomery MK, Fire A (1998) Double-stranded RNA as a mediator in sequence-specific genetic silencing and co-suppression. Trends Genet 14:255–258

    Article  PubMed  CAS  Google Scholar 

  • Montgomery MK, Xu S, Fire A (1998) RNA as a target of double-stranded RNA-mediated genetic interference in Caenorhabditis elegans. Proc Natl Acad Sci U S A 95:15502–15507

    Article  PubMed  CAS  Google Scholar 

  • Ohler U, Yekta S, Lim LP, Bartel DP, Burge CB (2004) Patterns of flanking sequence conservation and a characteristic upstream motif for microRNA gene identification. RNA 10:1309–1322

    Article  PubMed  CAS  Google Scholar 

  • Pak J, Fire A (2007) Distinct populations of primary and secondary effectors during RNAi in C. elegans. Science 315:241–244

    Article  PubMed  CAS  Google Scholar 

  • Ruby JG, Jan C, Player C, Axtell MJ, Lee W, Nusbaum C, Ge H, Bartel DP (2006) Large-scale sequencing reveals 21U-RNAs and additional microRNAs and endogenous siRNAs in C. elegans. Cell 127:1193–1207

    Article  PubMed  CAS  Google Scholar 

  • Saleh MC, van Rij RP, Hekele A, Gillis A, Foley E, O'Farrell PH, Andino R (2006) The endocytic pathway mediates cell entry of dsRNA to induce RNAi silencing. Nat Cell Biol 8:793–802

    Article  PubMed  CAS  Google Scholar 

  • Schwarz DS, Hutvagner G, Du T, Xu Z, Aronin N, Zamore PD (2003) Asymmetry in the assembly of the RNAi enzyme complex. Cell 115:199–208

    Article  PubMed  CAS  Google Scholar 

  • Shih JD, Fitzgerald MC, Sutherlin M, Hunter CP (2009) The SID-1 double-stranded RNA transporter is not selective for dsRNA length. RNA 15:384–390

    Article  PubMed  CAS  Google Scholar 

  • Sijen T, Steiner FA, Thijssen KL, Plasterk RH (2007) Secondary siRNAs result from unprimed RNA synthesis and form a distinct class. Science 315:244–247

    Article  PubMed  CAS  Google Scholar 

  • Simmer F, Tijsterman M, Parrish S, Koushika SP, Nonet ML, Fire A, Ahringer J, Plasterk RH (2002) Loss of the putative RNA-directed RNA polymerase RRF-3 makes C. elegans hypersensitive to RNAi. Curr Biol 12:1317–1319

    Article  PubMed  CAS  Google Scholar 

  • Siomi H, Siomi MC (2009) On the road to reading the RNA-interference code. Nature 457:396–404

    Article  PubMed  CAS  Google Scholar 

  • Steiner FA, Okihara KL, Hoogstrate SW, Sijen T, Ketting RF (2009) RDE-1 slicer activity is required only for passenger-strand cleavage during RNAi in Caenorhabditis elegans. Nat Struct Mol Biol 16:207–211

    Article  PubMed  CAS  Google Scholar 

  • Sulston JE, Horvitz HR (1977) Post-embryonic cell lineages of the nematode, Caenorhabditis elegans. Dev Biol 56:110–156

    Article  PubMed  CAS  Google Scholar 

  • Sulston JE, Schierenberg E, White JG, Thomson JN (1983) The embryonic cell lineage of the nematode Caenorhabditis elegans. Dev Biol 100:64–119

    Article  PubMed  CAS  Google Scholar 

  • Sundaram P, Echalier B, Han W, Hull D, Timmons L (2006) ATP-binding cassette transporters are required for efficient RNAi interference in Caenorhabditis elegans Mol Biol Cell 17(8):3678–88

    CAS  Google Scholar 

  • Tabara H, Grishok A, Mello CC (1998) RNAi in C. elegans: soaking in the genome sequence. Science 282:430–431

    Article  PubMed  CAS  Google Scholar 

  • Tabara H, Sarkissian M, Kelly WG, Fleenor J, Grishok A, Timmons L, Fire A, Mello CC (1999) The rde-1 gene, RNA interference, and transposon silencing in C. elegans. Cell 99:123–132

    Article  PubMed  CAS  Google Scholar 

  • Tabara H, Yigit E, Siomi H, Mello CC (2002) The dsRNA binding protein RDE-4 interacts with RDE-1, DCR-1, and a DExH-box helicase to direct RNAi in C. elegans. Cell 109:861–871

    Article  PubMed  CAS  Google Scholar 

  • Tavernarakis N, Wang SL, Dorovkov M, Ryazanov A, Driscoll M (2000) Heritable and inducible genetic interference by double-stranded RNA encoded by transgenes. Nat Genet 24:180–183

    Article  PubMed  CAS  Google Scholar 

  • Tijsterman M, Plasterk RH (2004) Dicers at RISC; the mechanism of RNAi. Cell 117:1–3

    Article  PubMed  CAS  Google Scholar 

  • Tijsterman M, Ketting RF, Okihara KL, Sijen T, Plasterk RH (2002) RNA helicase MUT-14-dependent gene silencing triggered in C. elegans by short antisense RNAs. Science 295:694–697

    Article  PubMed  CAS  Google Scholar 

  • Tijsterman M, May RC, Simmer F, Okihara KL, Plasterk RH (2004) Genes required for systemic RNA interference in Caenorhabditis elegans. Curr Biol 14:111–116

    Article  PubMed  CAS  Google Scholar 

  • Timmons LD (2007) ABC transporters and RNAi in Caenorhabditis elegans. J Bioenerg Biomembr 39:459–463

    Article  PubMed  CAS  Google Scholar 

  • Timmons L, Fire A (1998) Specific interference by ingested dsRNA. Nature 395:854

    Article  PubMed  CAS  Google Scholar 

  • Timmons L, Court DL, Fire A (2001) Ingestion of bacterially expressed dsRNAs can produce specific and potent genetic interference in Caenorhabditis elegans. Gene 263:103–112

    Article  PubMed  CAS  Google Scholar 

  • Timmons L, Tabara H, Mello CC, Fire AZ (2003) Inducible systemic RNA silencing in Caenorhabditis elegans. Mol Biol Cell 14:2972–2983

    Article  PubMed  CAS  Google Scholar 

  • Tomoyasu Y, Miller SC, Tomita S, Schoppmeier M, Grossmann D, Bucher G (2008) Exploring systemic RNA interference in insects: a genome-wide survey for RNAi genes in Tribolium. Genome Biol 9:R10

    Article  PubMed  Google Scholar 

  • Vastenhouw NL, Brunschwig K, Okihara KL, Muller F, Tijsterman M, Plasterk RH (2006) Gene expression: long-term gene silencing by RNAi. Nature 442:882

    Article  PubMed  CAS  Google Scholar 

  • Wang HW, Noland C, Siridechadilok B, Taylor DW, Ma E, Felderer K, Doudna JA, Nogales E (2009) Structural insights into RNA processing by the human RISC-loading complex. Nat Struct Mol Biol 16(11):1148–1153

    Article  PubMed  CAS  Google Scholar 

  • Winston WM, Molodowitch C, Hunter CP (2002) Systemic RNAi in C. elegans requires the putative transmembrane protein SID-1. Science 295:2456–2459

    Article  PubMed  CAS  Google Scholar 

  • Winston WM, Sutherlin M, Wright AJ, Feinberg EH, Hunter CP (2007) Caenorhabditis elegans SID-2 is required for environmental RNA interference. Proc Natl Acad Sci U S A 104:10565–10570

    Article  PubMed  CAS  Google Scholar 

  • Yigit E, Batista PJ, Bei Y, Pang KM, Chen CC, Tolia NH, Joshua-Tor L, Mitani S, Simard MJ, Mello CC (2006) Analysis of the C. elegans Argonaute family reveals that distinct Argonautes act sequentially during RNAi. Cell 127:747–757

    Article  PubMed  CAS  Google Scholar 

  • Zamore PD, Tuschl T, Sharp PA, Bartel DP (2000) RNAi: double-stranded RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell 101:25–33

    Article  PubMed  CAS  Google Scholar 

  • Zgheib O, Pataky K, Brugger J, Halazonetis TD (2009) An oligomerized 53BP1 tudor domain suffices for recognition of DNA double-strand breaks. Mol Cell Biol 29:1050–1058

    Article  PubMed  CAS  Google Scholar 

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Acknowledgment

My special thanks are due to A. Fire for the integrated gfp reporter used in Fig. 6.2 and to members of the laboratory for their helpful comments. My sincere apologies to those whose work was not given adequate credit. I gratefully acknowledge the American Cancer Society for funding the research in our lab.

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Correspondence to Lisa Timmons .

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Timmons, L. (2010). Systemic RNAi in C. elegans from the Viewpoint of RNA as Extracellular Signals. In: Kikuchi, Y., Rykova, E. (eds) Extracellular Nucleic Acids. Nucleic Acids and Molecular Biology, vol 25. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-12617-8_6

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