Endogenous RNAi pathways in C. elegans* Allison C. Billi1, Sylvia E. J. Fischer2, and John K. Kim1§ 1Life Sciences Institute, Department of Human Genetics, University of Michigan, Ann Arbor MI, USA 2Department of Molecular Biology, Massachusetts General Hospital, Department of Genetics, Harvard Medical School, Boston MA, USA Table of Contents 1. Introduction ............................................................................................................................3 2. Features and targets of 26G RNAs .............................................................................................. 4 3. Biogenesis of 26G RNAs: The ERI complex ................................................................................. 5 3.1. RRF-3 .........................................................................................................................8 3.2. DRH-3 ........................................................................................................................8 3.3. ERI-5 ..........................................................................................................................9 3.4. DCR-1 ........................................................................................................................9 3.5. DCR-1 helicase domain .................................................................................................. 9 3.6. ERI-1b ...................................................................................................................... 10 3.7. ERI-3 ........................................................................................................................ 10 3.8. RDE-4 ...................................................................................................................... 10 4. Other factors implicated in 26G RNA accumulation ..................................................................... 11 4.1. ERI-9 ........................................................................................................................ 11 4.2. ERI-6/7 ..................................................................................................................... 11 4.3. MUT-16 .................................................................................................................... 11 4.4. Other Mutator (MUT) proteins ....................................................................................... 11 5. 26G RNA Argonaute loading ................................................................................................... 11 5.1. ALG-3/4 .................................................................................................................... 12 5.2. ERGO-1 .................................................................................................................... 12 5.3. HENN-1 .................................................................................................................... 12 6. 26G RNA effector function ...................................................................................................... 13 6.1. Sperm-origin ts sterility due to loss of ALG-3/4 class 26G RNAs ......................................... 15 6.2. Enhanced RNAi sensitivity due to loss of ERGO-1 class 26G RNAs ..................................... 16 * Edited by Julie Ahringer. Last revised Novermber 10, 2013. Published May 7, 2014. This chapter should be cited as: Billi et al. Endogenous RNAi pathways in C. elegans (May 7, 2014), WormBook, ed. The C. elegans Research Community, WormBook, doi/10.1895/wormbook.1.170.1, http://www.wormbook.org. Copyright: © 2014 Allison C. Billi, Sylvia E. J. Fischer, John K. Kim. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. §To whom correspondence should be addressed. E-mail: [email protected] 1 Endogenous RNAi pathways in C. elegans 7. 22G RNAs comprise two major small RNA pathways ................................................................... 16 8. 22G RNA biogenesis: The 22G RNA RdRP module ..................................................................... 16 8.1. RRF-1 ....................................................................................................................... 17 8.2. DRH-3 ...................................................................................................................... 17 8.3. EKL-1 ....................................................................................................................... 18 9. Triggering of WAGO 22G RNAs .............................................................................................. 18 9.1. Triggering by 26G RNAs .............................................................................................. 19 9.2. Triggering by primary exo-siRNAs ................................................................................. 19 9.3. Triggering by RDE-1-scavenged small RNAs ................................................................... 19 9.4. Triggering by 21U RNAs .............................................................................................. 19 10. Germline amplification of WAGO 22G RNAs in perinuclear Mutator foci ...................................... 20 10.1. Dynamics of Mutator foci ........................................................................................... 20 10.2. Assembly of Mutator foci ............................................................................................ 20 10.3. Function of Mutator foci ............................................................................................. 21 11. Additional amplification of WAGO 22G RNAs by RDE-10/RDE-11 ............................................. 21 11.1. RDE-10/RDE-11 amplifies 22G RNAs triggered by exo-RNAi and ERGO-1 ........................ 21 11.2. RDE-10/RDE-11-mediated target degradation ................................................................. 22 11.3. RSD-2, RSD-6, and/or HAF-6 may potentiate RDE-10/RDE-11 function ............................. 22 12. WAGO 22G RNA Argonaute loading ...................................................................................... 22 13. WAGO 22G RNA-mediated post-transcriptional gene silencing .................................................... 23 14. WAGO 22G RNA-mediated transcriptional gene silencing: The nuclear RNAi pathway .................... 24 14.1. Early evidence for transcriptional gene silencing ............................................................. 24 14.2. Early evidence for heritable and multigenerational gene silencing ....................................... 24 14.3. NRDE-3 and HRDE-1 WAGO Argonautes mediate nuclear RNAi ...................................... 24 14.4. NRDE-3 collaborates with other WAGO Argonautes to silence somatic targets ..................... 25 14.5. HRDE-1 transgenerationally silences diverse targets to promote germline immortality ............ 25 15. NRDE-3 target recognition .................................................................................................... 26 16. The downstream nuclear RNAi machinery ................................................................................ 26 16.1. NRDE-2 .................................................................................................................. 27 16.2. NRDE-1 .................................................................................................................. 27 16.3. NRDE-4 .................................................................................................................. 27 17. Nuclear RNAi inhibits transcriptional elongation and drives H3K9 trimethylation ............................ 27 18. NRDE-3 directs single-generation epigenetic inheritance of silencing ............................................ 28 19. HRDE-1 directs multigenerational epigenetic inheritance of silencing ............................................ 29 20. Features and targets of CSR-1 22G RNAs ................................................................................. 30 21. The CSR-1 22G RNA pathway molecular machinery .................................................................. 31 21.1. CSR-1 ..................................................................................................................... 31 22. Phenotypes associated with CSR-1 22G RNA pathway compromise .............................................. 31 22.1. Germline phenotypes .................................................................................................. 31 22.2. Embryonic phenotypes ............................................................................................... 32 23. Localization and assembly of the CSR-1 22G RNA complex ........................................................ 32 23.1. Germline localization ................................................................................................. 32 23.2. Embryo localization ................................................................................................... 32 24. CSR-1 22G RNA effector function .......................................................................................... 33 25. CDE-1 regulates the CSR-1 22G RNA pathway ......................................................................... 34 26. 21U RNA features and loci ...................................................................................................