Analysis of the C. elegans Germline Stem Cell Pool

  • Sarah L. CrittendenEmail author
  • Hannah S. Seidel
  • Judith Kimble
Part of the Methods in Molecular Biology book series (MIMB, volume 1463)


The Caenorhabditis elegans germline is an excellent model for studying the regulation of a pool of stem cells and progression of cells from a stem cell state to a differentiated state. At the tissue level, the germline is organized in an assembly line with the germline stem cell (GSC) pool at one end and differentiated cells at the other. A simple mesenchymal niche caps the GSC region of the germline and maintains GSCs in an undifferentiated state by signaling through the conserved Notch pathway. Downstream of Notch signaling, key regulators include novel LST-1 and SYGL-1 proteins and a network of RNA regulatory proteins. In this chapter we present methods for characterizing the C. elegans GSC pool and early germ cell differentiation. The methods include examination of the germline in living and fixed worms, cell cycle analysis, and analysis of markers. We also discuss assays to separate mutants that affect the stem cell vs. differentiation decision from those that affect germ cell processes more generally.

Key words

Stem cells Progenitor cells C. elegans Germline Proliferation Meiosis Mitosis EdU Cell cycle 


  1. 1.
    Kershner A, Crittenden SL, Friend K, Sorensen EB, Porter DF, Kimble J (2013) Germline stem cells and their regulation in the nematode Caenorhabditis elegans. Adv Exp Med Biol 786:29–46. doi: 10.1007/978-94-007-6621-1_3 PubMedCrossRefGoogle Scholar
  2. 2.
    Hansen D, Schedl T (2013) Stem cell proliferation versus meiotic fate decision in Caenorhabditis elegans. Adv Exp Med Biol 757:71–99. doi: 10.1007/978-1-4614-4015-4_4 PubMedPubMedCentralCrossRefGoogle Scholar
  3. 3.
    Hubbard EJ, Korta DZ, Dalfo D (2013) Physiological control of germline development. Adv Exp Med Biol 757:101–131. doi: 10.1007/978-1-4614-4015-4_5 PubMedPubMedCentralCrossRefGoogle Scholar
  4. 4.
    Kimble J, Seidel H (2013) C. elegans germline stem cells and their niche. Stembook. doi: 10.3824/stembook.1.95.1
  5. 5.
    Lander AD, Kimble J, Clevers H, Fuchs E, Montarras D, Buckingham M, Calof AL, Trumpp A, Oskarsson T (2012) What does the concept of the stem cell niche really mean today? BMC Biol 10:19. doi: 10.1186/1741-7007-10-19 PubMedPubMedCentralCrossRefGoogle Scholar
  6. 6.
    Joshi PM, Riddle MR, Djabrayan NJ, Rothman JH (2010) Caenorhabditis elegans as a model for stem cell biology. Dev Dyn 239(5):1539–1554. doi: 10.1002/dvdy.22296 PubMedPubMedCentralCrossRefGoogle Scholar
  7. 7.
    Kimble J (2011) Molecular regulation of the mitosis/meiosis decision in multicellular organisms. Cold Spring Harb Perspect Biol 3(8):a0002683. doi: 10.1101/cshperspect.a002683 CrossRefGoogle Scholar
  8. 8.
    Hubbard EJ (2007) Caenorhabditis elegans germ line: a model for stem cell biology. Dev Dyn 236(12):3343–3357. doi: 10.1002/dvdy.21335 PubMedPubMedCentralCrossRefGoogle Scholar
  9. 9.
    Cinquin O (2009) Purpose and regulation of stem cells: a systems-biology view from the Caenorhabditis elegans germ line. J Pathol 217(2):186–198. doi: 10.1002/path.2481 PubMedPubMedCentralCrossRefGoogle Scholar
  10. 10.
    Biedermann B, Hotz HR, Ciosk R (2010) The Quaking family of RNA-binding proteins: coordinators of the cell cycle and differentiation. Cell Cycle 9(10):1929–1933PubMedCrossRefGoogle Scholar
  11. 11.
    Kipreos ET (2005) C. elegans cell cycles: invariance and stem cell divisions. Nat Rev Mol Cell Biol 6(10):766–776PubMedCrossRefGoogle Scholar
  12. 12.
    Waters KA, Reinke V (2011) Extrinsic and intrinsic control of germ cell proliferation in Caenorhabditis elegans. Mol Reprod Dev 78(3):151–160. doi: 10.1002/mrd.21289 PubMedPubMedCentralCrossRefGoogle Scholar
  13. 13.
    Kimble J, Crittenden SL (2007) Controls of germline stem cells, entry into meiosis, and the sperm/oocyte decision in Caenorhabditis elegans. Annu Rev Cell Dev Biol 23:405–433. doi: 10.1146/annurev.cellbio.23.090506.123326 PubMedCrossRefGoogle Scholar
  14. 14.
    Crittenden SL, Leonhard KA, Byrd DT, Kimble J (2006) Cellular analyses of the mitotic region in the Caenorhabditis elegans adult germ line. Mol Biol Cell 17(7):3051–3061PubMedPubMedCentralCrossRefGoogle Scholar
  15. 15.
    Cinquin O, Crittenden SL, Morgan DE, Kimble J (2010) Progression from a stem cell-like state to early differentiation in the C. elegans germ line. Proc Natl Acad Sci U S A 107(5):2048–2053. doi: 10.1073/pnas.0912704107 PubMedPubMedCentralCrossRefGoogle Scholar
  16. 16.
    Kimble JE, White JG (1981) On the control of germ cell development in Caenorhabditis elegans. Dev Biol 81:208–219PubMedCrossRefGoogle Scholar
  17. 17.
    Kershner AM, Kimble J (2010) Genome-wide analysis of mRNA targets for Caenorhabditis elegans FBF, a conserved stem cell regulator. Proc Natl Acad Sci U S A 107(8):3936–3941. doi: 10.1073/pnas.1000495107 PubMedPubMedCentralCrossRefGoogle Scholar
  18. 18.
    Racher H, Hansen D (2010) Translational control in the C. elegans hermaphrodite germ line. Genome 53(2):83–102. doi: 10.1139/g09-090 PubMedCrossRefGoogle Scholar
  19. 19.
    Moore FL, Jaruzelska J, Fox MS, Urano J, Firpo MT, Turek PJ, Dorfman DM, Reijo Pera RA (2003) Human Pumilio-2 is expressed in embryonic stem cells and germ cells and interacts with DAZ (Deleted in AZoospermia) and DAZ-like proteins. Proc Natl Acad Sci U S A 100(2):538–543. doi: 10.1073/pnas.0234478100 PubMedPubMedCentralCrossRefGoogle Scholar
  20. 20.
    Kimble J, Hirsh D (1979) The postembryonic cell lineages of the hermaphrodite and male gonads in Caenorhabditis elegans. Dev Biol 70(2):396–417PubMedCrossRefGoogle Scholar
  21. 21.
    Angelo G, Van Gilst MR (2009) Starvation protects germline stem cells and extends reproductive longevity in C. elegans. Science 326(5955):954–958. doi: 10.1126/science.1178343 PubMedCrossRefGoogle Scholar
  22. 22.
    Seidel HS, Kimble J (2011) The oogenic germline starvation response in C. elegans. PLoS One 6(12):e28074. doi: 10.1371/journal.pone.0028074 PubMedPubMedCentralCrossRefGoogle Scholar
  23. 23.
    Morgan DE, Crittenden SL, Kimble J (2010) The C. elegans adult male germline: stem cells and sexual dimorphism. Dev Biol 346(2):204–214. doi: 10.1016/j.ydbio.2010.07.022 PubMedPubMedCentralCrossRefGoogle Scholar
  24. 24.
    Hubbard EJ (2011) Insulin and germline proliferation in Caenorhabditis elegans. Vitam Horm 87:61–77. doi: 10.1016/B978-0-12-386015-6.00024-X PubMedPubMedCentralCrossRefGoogle Scholar
  25. 25.
    Salinas LS, Maldonado E, Navarro RE (2006) Stress-induced germ cell apoptosis by a p53 independent pathway in Caenorhabditis elegans. Cell Death Differ 13(12):2129–2139. doi: 10.1038/sj.cdd.4401976 PubMedCrossRefGoogle Scholar
  26. 26.
    Gracida X, Eckmann CR (2013) Fertility and germline stem cell maintenance under different diets requires nhr-114/HNF4 in C. elegans. Curr Biol 23(7):607–613. doi: 10.1016/j.cub.2013.02.034 PubMedCrossRefGoogle Scholar
  27. 27.
    Qin Z, Hubbard EJ (2015) Non-autonomous DAF-16/FOXO activity antagonizes age-related loss of C. elegans germline stem/progenitor cells. Nat Commun 6:7107. doi: 10.1038/ncomms8107 PubMedPubMedCentralCrossRefGoogle Scholar
  28. 28.
    Seidel HS, Kimble J (2015) Cell-cycle quiescence maintains germline stem cells independent of GLP-1/Notch. Elife 4. doi: 10.7554/eLife.10832
  29. 29.
    Lopez AL 3rd, Chen J, Joo HJ, Drake M, Shidate M, Kseib C, Arur S (2013) DAF-2 and ERK couple nutrient availability to meiotic progression during Caenorhabditis elegans oogenesis. Dev Cell 27(2):227–240. doi: 10.1016/j.devcel.2013.09.008 PubMedCrossRefGoogle Scholar
  30. 30.
    Gupta P, Leahul L, Wang X, Wang C, Bakos B, Jasper K, Hansen D (2015) Proteasome regulation of the chromodomain protein MRG-1 controls the balance between proliferative fate and differentiation in the C. elegans germ line. Development 142(2):291–302. doi: 10.1242/dev.115147 PubMedCrossRefGoogle Scholar
  31. 31.
    Millonigg S, Minasaki R, Nousch M, Eckmann CR (2014) GLD-4-mediated translational activation regulates the size of the proliferative germ cell pool in the adult C. elegans germ line. PLoS Genet 10(9):e1004647. doi: 10.1371/journal.pgen.1004647 PubMedPubMedCentralCrossRefGoogle Scholar
  32. 32.
    Novak P, Wang X, Ellenbecker M, Feilzer S, Voronina E (2015) Splicing machinery facilitates post-transcriptional regulation by FBFs and other RNA-binding proteins in Caenorhabditis elegans germline. G3 (Bethesda). doi: 10.1534/g3.115.019315
  33. 33.
    Jaramillo-Lambert A, Ellefson M, Villeneuve AM, Engebrecht J (2007) Differential timing of S phases, X chromosome replication, and meiotic prophase in the C. elegans germ line. Dev Biol 308(1):206–221. doi: 10.1016/j.ydbio.2007.05.019 PubMedCrossRefGoogle Scholar
  34. 34.
    Fox PM, Vought VE, Hanazawa M, Lee MH, Maine EM, Schedl T (2011) Cyclin E and CDK-2 regulate proliferative cell fate and cell cycle progression in the C. elegans germline. Development 138(11):2223–2234. doi: 10.1242/dev.059535 PubMedPubMedCentralCrossRefGoogle Scholar
  35. 35.
    Chiang M, Cinquin A, Paz A, Meeds E, Price CA, Welling M, Cinquin O (2015) Control of Caenorhabditis elegans germ-line stem-cell cycling speed meets requirements of design to minimize mutation accumulation. BMC Biol 13(1):51. doi: 10.1186/s12915-015-0148-y PubMedPubMedCentralCrossRefGoogle Scholar
  36. 36.
    Kimble J, Crittenden SL (2005) Germline proliferation and its control. WormBook. doi: 10.1895/wormbook.1.13.1 PubMedPubMedCentralGoogle Scholar
  37. 37.
    Hansen D, Schedl T (2006) The regulatory network controlling the proliferation-meiotic entry decision in the Caenorhabditis elegans germ line. Curr Top Dev Biol 76:185–215PubMedCrossRefGoogle Scholar
  38. 38.
    Fox PM, Schedl T (2015) Analysis of germline stem cell differentiation following loss of GLP-1 Notch activity in Caenorhabditis elegans. Genetics 201:167–184. doi: 10.1534/genetics.115.178061 PubMedPubMedCentralCrossRefGoogle Scholar
  39. 39.
    Morgan CT, Noble D, Kimble J (2013) Mitosis-meiosis and sperm-oocyte fate decisions are separable regulatory events. Proc Natl Acad Sci U S A 110(9):3411–3416. doi: 10.1073/pnas.1300928110 PubMedPubMedCentralCrossRefGoogle Scholar
  40. 40.
    Hansen D, Wilson-Berry L, Dang T, Schedl T (2004) Control of the proliferation versus meiotic development decision in the C. elegans germline through regulation of GLD-1 protein accumulation. Development 131:93–104. doi: 10.1242/dev.00916 PubMedCrossRefGoogle Scholar
  41. 41.
    Hansen D, Hubbard EJA, Schedl T (2004) Multi-pathway control of the proliferation versus meiotic development decision in the Caenorhabditis elegans germline. Dev Biol 268(2):342–357PubMedCrossRefGoogle Scholar
  42. 42.
    Zetka MC, Kawasaki I, Strome S, Müller F (1999) Synapsis and chiasma formation in Caenorhabditis elegans require HIM-3, a meiotic chromosome core component that functions in chromosome segregation. Genes Dev 13(17):2258–2270PubMedPubMedCentralCrossRefGoogle Scholar
  43. 43.
    Thompson BE, Bernstein DS, Bachorik JL, Petcherski AG, Wickens M, Kimble J (2005) Dose-dependent control of proliferation and sperm specification by FOG-1/CPEB. Development 132(15):3471–3481. doi: 10.1242/dev.01921 PubMedPubMedCentralCrossRefGoogle Scholar
  44. 44.
    Maciejowski J, Ugel N, Mishra B, Isopi M, Hubbard EJA (2006) Quantitative analysis of germline mitosis in adult C. elegans. Dev Biol 292:142–151. doi: 10.1016/j.ydbio.2005.12.046 PubMedCrossRefGoogle Scholar
  45. 45.
    Snow JJ, Lee MH, Verheyden J, Kroll-Conner PL, Kimble J (2013) C. elegans FOG-3/Tob can either promote or inhibit germline proliferation, depending on gene dosage and genetic context. Oncogene 32(21):2614–2621. doi: 10.1038/onc.2012.291 PubMedCrossRefGoogle Scholar
  46. 46.
    Lamont LB, Kimble J (2007) Developmental expression of FOG-1/CPEB protein and its control in the Caenorhabditis elegans hermaphrodite germ line. Dev Dyn 236(3):871–879PubMedPubMedCentralCrossRefGoogle Scholar
  47. 47.
    Jones AR, Francis R, Schedl T (1996) GLD-1, a cytoplasmic protein essential for oocyte differentiation, shows stage- and sex-specific expression during Caenorhabditis elegans germline development. Dev Biol 180(1):165–183PubMedCrossRefGoogle Scholar
  48. 48.
    Pazdernik N, Schedl T (2013) Introduction to germ cell development in Caenorhabditis elegans. Adv Exp Med Biol 757:1–16. doi: 10.1007/978-1-4614-4015-4_1 PubMedPubMedCentralCrossRefGoogle Scholar
  49. 49.
    Rothman JH, Singson A (eds) (2012) Caenorhabditis elegans: cell biology and physiology, vol 107. Methods in cell biology. ElsevierGoogle Scholar
  50. 50.
    Rothman JH, Singson A (2011) Caenorhabditis elegans: molecular genetics and development. Methods Cell Biol 106:xv–xviiiPubMedCrossRefGoogle Scholar
  51. 51.
    Ghazi A, Yanowitz J, Silverman GA (eds) (2014) C. elegans: methods, vol 68(3). ElsevierGoogle Scholar
  52. 52.
    Ito K, McGhee JD (1987) Parental DNA strands segregate randomly during embryonic development of Caenorhabditis elegans. Cell 49(3):329–336PubMedCrossRefGoogle Scholar
  53. 53.
    Dernburg AF, McDonald K, Moulder G, Barstead R, Dresser M, Villeneuve AM (1998) Meiotic recombination in C. elegans initiates by a conserved mechanism and is dispensable for homologous chromosome synapsis. Cell 94(3):387–398PubMedCrossRefGoogle Scholar
  54. 54.
    Francis R, Barton MK, Kimble J, Schedl T (1995) gld-1, a tumor suppressor gene required for oocyte development in Caenorhabditis elegans. Genetics 139(2):579–606PubMedPubMedCentralGoogle Scholar
  55. 55.
    Gumienny TL, Lambie E, Hartwieg E, Horvitz HR, Hengartner MO (1999) Genetic control of programmed cell death in the Caenorhabditis elegans hermaphrodite germline. Development 126(5):1011–1022PubMedGoogle Scholar
  56. 56.
    Sulston J, Hodgkin J (1988) Methods. In: Wood WB (ed) The nematode Caenorhabditis elegans, vol 17, Cold Spring Harbor monograph series. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp 587–606Google Scholar
  57. 57.
    Porta-de-la-Riva M, Fontrodona L, Villanueva A, Ceron J (2012) Basic Caenorhabditis elegans methods: synchronization and observation. J Vis Exp 64:e4019. doi: 10.3791/4019 PubMedGoogle Scholar
  58. 58.
    Hutter H (2012) Fluorescent protein methods: strategies and applications. Methods Cell Biol 107:67–92. doi: 10.1016/B978-0-12-394620-1.00003-5 PubMedCrossRefGoogle Scholar
  59. 59.
    McCarter J, Bartlett B, Dang T, Schedl T (1999) On the control of oocyte meiotic maturation and ovulation in Caenorhabditis elegans. Dev Biol 205(1):111–128PubMedCrossRefGoogle Scholar
  60. 60.
    Shakes DC, Miller DM 3rd, Nonet ML (2012) Immunofluorescence microscopy. Methods Cell Biol 107:35–66. doi: 10.1016/B978-0-12-394620-1.00002-3 PubMedCrossRefGoogle Scholar
  61. 61.
    Duerr JS (2006) Immunohistochemistry. WormBook. doi: 10.1895/wormbook.1.105.1 PubMedPubMedCentralGoogle Scholar
  62. 62.
    Byrd DT, Knobel K, Affeldt K, Crittenden SL, Kimble J (2014) A DTC niche plexus surrounds the germline stem cell pool in Caenorhabditis elegans. PLoS One 9(2):e88372. doi: 10.1371/journal.pone.0088372 PubMedPubMedCentralCrossRefGoogle Scholar
  63. 63.
    Eckmann CR, Crittenden SL, Suh N, Kimble J (2004) GLD-3 and control of the mitosis/meiosis decision in the germline of Caenorhabditis elegans. Genetics 168:147–160. doi: 10.1534/genetics.104.029264 PubMedPubMedCentralCrossRefGoogle Scholar
  64. 64.
    Vogel JL, Michaelson D, Santella A, Hubbard EJ, Bao Z (2014) Irises: a practical tool for image-based analysis of cellular DNA content. Worm 3:e29041. doi: 10.4161/worm.29041 PubMedPubMedCentralCrossRefGoogle Scholar
  65. 65.
    Kershner AM, Shin H, Hansen TJ, Kimble J (2014) Discovery of two GLP-1/Notch target genes that account for the role of GLP-1/Notch signaling in stem cell maintenance. Proc Natl Acad Sci U S A 111(10):3739–3744. doi: 10.1073/pnas.1401861111 PubMedPubMedCentralCrossRefGoogle Scholar
  66. 66.
    Shirayama M, Seth M, Lee HC, Gu W, Ishidate T, Conte D Jr, Mello CC (2012) piRNAs initiate an epigenetic memory of nonself RNA in the C. elegans germline. Cell 150(1):65–77. doi: 10.1016/j.cell.2012.06.015 PubMedPubMedCentralCrossRefGoogle Scholar
  67. 67.
    Wedeles CJ, Wu MZ, Claycomb JM (2013) Protection of germline gene expression by the C. elegans Argonaute CSR-1. Dev Cell 27(6):664–671. doi: 10.1016/j.devcel.2013.11.016 PubMedCrossRefGoogle Scholar
  68. 68.
    Wedeles CJ, Wu MZ, Claycomb JM (2014) Silent no more: endogenous small RNA pathways promote gene expression. Worm 3:e28641. doi: 10.4161/worm.28641 PubMedPubMedCentralCrossRefGoogle Scholar
  69. 69.
    Crittenden SL, Troemel ER, Evans TC, Kimble J (1994) GLP-1 is localized to the mitotic region of the C. elegans germ line. Development 120:2901–2911PubMedGoogle Scholar
  70. 70.
    Crittenden SL, Bernstein DS, Bachorik JL, Thompson BE, Gallegos M, Petcherski AG, Moulder G, Barstead R, Wickens M, Kimble J (2002) A conserved RNA-binding protein controls germline stem cells in Caenorhabditis elegans. Nature 417:660–663. doi: 10.1038/nature754 PubMedCrossRefGoogle Scholar
  71. 71.
    Lamont LB, Crittenden SL, Bernstein D, Wickens M, Kimble J (2004) FBF-1 and FBF-2 regulate the size of the mitotic region in the C. elegans germline. Dev Cell 7(5):697–707PubMedCrossRefGoogle Scholar
  72. 72.
    Voronina E, Paix A, Seydoux G (2012) The P granule component PGL-1 promotes the localization and silencing activity of the PUF protein FBF-2 in germline stem cells. Development 139(20):3732–3740. doi: 10.1242/dev.083980 PubMedPubMedCentralCrossRefGoogle Scholar
  73. 73.
    Schumacher B, Hanazawa M, Lee M-H, Nayak S, Volkmann K, Hofmann R, Hengartner M, Schedl T, Gartner A (2005) Translational repression of C. elegans p53 by GLD-1 regulates DNA damage-induced apoptosis. Cell 120(3):357–368PubMedCrossRefGoogle Scholar
  74. 74.
    Blelloch R, Santa Anna-Arriola S, Gao D, Li Y, Hodgkin J, Kimble J (1999) The gon-1 gene is required for gonadal morphogenesis in Caenorhabditis elegans. Dev Biol 216:382–393PubMedCrossRefGoogle Scholar
  75. 75.
    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(1):101–123. doi: 10.1006/dbio.1999.9356 PubMedCrossRefGoogle Scholar
  76. 76.
    Wong BG, Paz A, Corrado MA, Ramos BR, Cinquin A, Cinquin O, Hui EE (2013) Live imaging reveals active infiltration of mitotic zone by its stem cell niche. Integr Biol (Cambridge) 5(7):976–982. doi: 10.1039/c3ib20291g CrossRefGoogle Scholar
  77. 77.
    Nadarajan S, Govindan JA, McGovern M, Hubbard EJA, Greenstein D (2009) MSP and GLP-1/Notch signaling coordinately regulate actomyosin-dependent cytoplasmic streaming and oocyte growth in C. elegans. Development 136(13):2223–2234. doi: 10.1242/dev.034603 PubMedPubMedCentralCrossRefGoogle Scholar
  78. 78.
    Korta DZ, Hubbard EJ (2010) Soma-germline interactions that influence germline proliferation in Caenorhabditis elegans. Dev Dyn 239(5):1449–1459. doi: 10.1002/dvdy.22268 PubMedPubMedCentralGoogle Scholar
  79. 79.
    van den Heuvel S (2005) Cell-cycle regulation. WormBook. doi: 10.1895/wormbook.1.28.1 PubMedPubMedCentralGoogle Scholar
  80. 80.
    Hendzel MJ, Wei Y, Mancini MA, Van Hooser A, Ranalli T, Brinkley BR, Bazett-Jones DP, Allis CD (1997) Mitosis-specific phosphorylation of histone H3 initiates primarily within pericentromeric heterochromatin during G2 and spreads in an ordered fashion coincident with mitotic chromosome condensation. Chromosoma 106(6):348–360PubMedCrossRefGoogle Scholar
  81. 81.
    Crittenden SL, Kimble J (2008) Analysis of the C. elegans germline stem cell region. In: Hou S, Singh SR (eds) Germline stem cell protocols, vol 450, Methods in molecular biology. Humana, Totowa, NJ, pp 27–44CrossRefGoogle Scholar
  82. 82.
    Jantsch V, Tang L, Pasierbek P, Penkner A, Nayak S, Baudrimont A, Schedl T, Gartner A, Loidl J (2007) Caenorhabditis elegans prom-1 is required for meiotic prophase progression and homologous chromosome pairing. Mol Biol Cell 18(12):4911–4920. doi: 10.1091/mbc.E07-03-0243 PubMedPubMedCentralCrossRefGoogle Scholar
  83. 83.
    Sato A, Isaac B, Phillips CM, Rillo R, Carlton PM, Wynne DJ, Kasad RA, Dernburg AF (2009) Cytoskeletal forces span the nuclear envelope to coordinate meiotic chromosome pairing and synapsis. Cell 139(5):907–919. doi: 10.1016/j.cell.2009.10.039
  84. 84.
    Salic A, Mitchison TJ (2008) A chemical method for fast and sensitive detection of DNA synthesis in vivo. Proc Natl Acad Sci U S A 105(7):2415–2420PubMedPubMedCentralCrossRefGoogle Scholar
  85. 85.
    Jeong J, Verheyden JM, Kimble J (2011) Cyclin E and Cdk2 control GLD-1, the mitosis/meiosis decision, and germline stem cells in Caenorhabditis elegans. PLoS Genet 7(3):e1001348. doi: 10.1371/journal.pgen.1001348 PubMedPubMedCentralCrossRefGoogle Scholar
  86. 86.
    Aherne WA, Camplejohn RS, Wright NA (1977) An introduction to cell population kinetics. Edward Arnold, LondonGoogle Scholar
  87. 87.
    Fay DS (2013) Classical genetic methods. WormBook. pp 1–58. doi: 10.1895/wormbook.1.165.1
  88. 88.
    Ahringer J. (ed) (2006) Reverse genetics. WormBook. doi: 10.1895/wormbook.1.47.1
  89. 89.
    Kutscher LM, Shaham S (2014) Forward and reverse mutagenesis in C. elegans. WormBook. pp 1–26. doi: 10.1895/wormbook.1.167.1
  90. 90.
    Huang L, Sternberg PW (2006) Genetic dissection of developmental pathways. WormBook. doi: 10.1895/wormbook.1.88.2 PubMedPubMedCentralGoogle Scholar
  91. 91.
    Kemphues K (2005) Essential genes. WormBook. pp 1–7. doi: 10.1895/wormbook.1.57.1
  92. 92.
    Lambie EJ, Kimble J (1991) Two homologous regulatory genes, lin-12 and glp-1, have overlapping functions. Development 112:231–240PubMedGoogle Scholar
  93. 93.
    Hubbard EJ (2014) FLP/FRT and Cre/lox recombination technology in C. elegans. Methods 68(3):417–424. doi: 10.1016/j.ymeth.2014.05.007 PubMedPubMedCentralCrossRefGoogle Scholar
  94. 94.
    Kage-Nakadai E, Imae R, Suehiro Y, Yoshina S, Hori S, Mitani S (2014) A conditional knockout toolkit for Caenorhabditis elegans based on the Cre/loxP recombination. PLoS One 9(12):e114680. doi: 10.1371/journal.pone.0114680 PubMedPubMedCentralCrossRefGoogle Scholar
  95. 95.
    Wei X, Potter CJ, Luo L, Shen K (2012) Controlling gene expression with the Q repressible binary expression system in Caenorhabditis elegans. Nat Methods 9(4):391–395. doi: 10.1038/nmeth.1929 PubMedPubMedCentralCrossRefGoogle Scholar
  96. 96.
    Zeiser E, Frokjaer-Jensen C, Jorgensen E, Ahringer J (2011) MosSCI and gateway compatible plasmid toolkit for constitutive and inducible expression of transgenes in the C. elegans germline. PLoS One 6(5):e20082. doi: 10.1371/journal.pone.0020082 PubMedPubMedCentralCrossRefGoogle Scholar
  97. 97.
    Cho U, Zimmerman SM, Chen LC, Owen E, Kim JV, Kim SK, Wandless TJ (2013) Rapid and tunable control of protein stability in Caenorhabditis elegans using a small molecule. PLoS One 8(8):e72393. doi: 10.1371/journal.pone.0072393 PubMedPubMedCentralCrossRefGoogle Scholar
  98. 98.
    Toettcher JE, Weiner OD, Lim WA (2013) Using optogenetics to interrogate the dynamic control of signal transmission by the Ras/Erk module. Cell 155(6):1422–1434. doi: 10.1016/j.cell.2013.11.004 PubMedPubMedCentralCrossRefGoogle Scholar
  99. 99.
    Yumerefendi H, Dickinson DJ, Wang H, Zimmerman SP, Bear JE, Goldstein B, Hahn K, Kuhlman B (2015) Control of protein activity and cell fate specification via light-mediated nuclear translocation. PLoS One 10(6):e0128443. doi: 10.1371/journal.pone.0128443 PubMedPubMedCentralCrossRefGoogle Scholar
  100. 100.
    Wedeles CJ, Wu MZ, Claycomb JM (2013) A multitasking Argonaute: exploring the many facets of C. elegans CSR-1. Chromosome Res 21(6-7):573–586. doi: 10.1007/s10577-013-9383-7 PubMedCrossRefGoogle Scholar
  101. 101.
    Merritt C, Seydoux G (2010) Transgenic solutions for the germline. WormBook. doi: 10.1895/wormbook.1.148.1 PubMedPubMedCentralGoogle Scholar
  102. 102.
    Fire A, Alcazar R, Tan F (2006) Unusual DNA structures associated with germline genetic activity in Caenorhabditis elegans. Genetics 173(3):1259–1273PubMedPubMedCentralCrossRefGoogle Scholar
  103. 103.
    Green RA, Audhya A, Pozniakovsky A, Dammermann A, Pemble H, Monen J, Portier N, Hyman A, Desai A, Oegema K (2008) Expression and imaging of fluorescent proteins in the C. elegans gonad and early embryo. Methods Cell Biol 85:179–218. doi: 10.1016/S0091-679X(08)85009-1 PubMedCrossRefGoogle Scholar
  104. 104.
    Austin J, Kimble J (1987) glp-1 is required in the germ line for regulation of the decision between mitosis and meiosis in C. elegans. Cell 51:589–599PubMedCrossRefGoogle Scholar
  105. 105.
    Crittenden SL, Eckmann CR, Wang L, Bernstein DS, Wickens M, Kimble J (2003) Regulation of the mitosis/meiosis decision in the Caenorhabditis elegans germline. Philos Trans R Soc Lond B Biol Sci 358:1359–1362. doi: 10.1098/rstb.2003.1333 PubMedPubMedCentralCrossRefGoogle Scholar
  106. 106.
    Berry LW, Westlund B, Schedl T (1997) Germ-line tumor formation caused by activation of glp-1, a Caenorhabditis elegans member of the Notch family of receptors. Development 124(4):925–936PubMedGoogle Scholar
  107. 107.
    Kadyk LC, Kimble J (1998) Genetic regulation of entry into meiosis in Caenorhabditis elegans. Development 125(10):1803–1813PubMedGoogle Scholar
  108. 108.
    Wright JE, Ciosk R (2013) RNA-based regulation of pluripotency. Trends Genet 29(2):99–107. doi: 10.1016/j.tig.2012.10.007 PubMedCrossRefGoogle Scholar
  109. 109.
    Biedermann B, Wright J, Senften M, Kalchhauser I, Sarathy G, Lee MH, Ciosk R (2009) Translational repression of cyclin E prevents precocious mitosis and embryonic gene activation during C. elegans meiosis. Dev Cell 17(3):355–364. doi: 10.1016/j.devcel.2009.08.003 PubMedCrossRefGoogle Scholar
  110. 110.
    Updike DL, Knutson AK, Egelhofer TA, Campbell AC, Strome S (2014) Germ-granule components prevent somatic development in the C. elegans germline. Curr Biol 24(9):970–975. doi: 10.1016/j.cub.2014.03.015 PubMedPubMedCentralCrossRefGoogle Scholar
  111. 111.
    Tursun B, Patel T, Kratsios P, Hobert O (2011) Direct conversion of C. elegans germ cells into specific neuron types. Science 331(6015):304–308. doi: 10.1126/science.1199082 PubMedCrossRefGoogle Scholar
  112. 112.
    Patel T, Tursun B, Rahe DP, Hobert O (2012) Removal of Polycomb repressive complex 2 makes C. elegans germ cells susceptible to direct conversion into specific somatic cell types. Cell Rep 2(5):1178–1186. doi: 10.1016/j.celrep.2012.09.020 PubMedPubMedCentralCrossRefGoogle Scholar
  113. 113.
    Robert VJ, Garvis S, Palladino F (2015) Repression of somatic cell fate in the germline. Cell Mol Life Sci. doi: 10.1007/s00018-015-1942-y PubMedGoogle Scholar
  114. 114.
    Killian DJ, Hubbard EJA (2004) C. elegans pro-1 activity is required for soma/germline interactions that influence proliferation and differentiation in the germ line. Development 131(6):1267–1278PubMedCrossRefGoogle Scholar
  115. 115.
    Pepper AS-R, Lo T-W, Killian DJ, Hall DH, Hubbard EJA (2003) The establishment of Caenorhabditis elegans germline pattern is controlled by overlapping proximal and distal somatic gonad signals. Dev Biol 259(2):336–350PubMedCrossRefGoogle Scholar
  116. 116.
    McGovern M, Voutev R, Maciejowski J, Corsi AK, Hubbard EJ (2009) A “latent niche” mechanism for tumor initiation. Proc Natl Acad Sci U S A 106(28):11617–11622. doi: 10.1073/pnas.0903768106 PubMedPubMedCentralCrossRefGoogle Scholar
  117. 117.
    Subramaniam K, Seydoux G (2003) Dedifferentiation of primary spermatocytes into germ cell tumors in C. elegans lacking the Pumilio-like protein PUF-8. Curr Biol 13(2):134–139PubMedCrossRefGoogle Scholar
  118. 118.
    Pepper AS-R, Killian DJ, Hubbard EJA (2003) Genetic analysis of Caenorhabditis elegans glp-1 mutants suggests receptor interaction or competition. Genetics 163(1):115–132PubMedPubMedCentralGoogle Scholar
  119. 119.
    Kraemer B, Crittenden S, Gallegos M, Moulder G, Barstead R, Kimble J, Wickens M (1999) NANOS-3 and FBF proteins physically interact to control the sperm-oocyte switch in Caenorhabditis elegans. Curr Biol 9(18):1009–1018PubMedCrossRefGoogle Scholar
  120. 120.
    Subramaniam K, Seydoux G (1999) nos-1 and nos-2, two genes related to Drosophila nanos, regulate primordial germ cell development and survival in Caenorhabditis elegans. Development 126(21):4861–4871PubMedGoogle Scholar
  121. 121.
    Killian DJ, Hubbard EJA (2005) Caenorhabditis elegans germline patterning requires coordinated development of the somatic gonadal sheath and the germ line. Dev Biol 279(2):322–335PubMedCrossRefGoogle Scholar
  122. 122.
    McCarter J, Bartlett B, Dang T, Schedl T (1997) Soma – germ cell interactions in Caenorhabditis elegans: multiple events of hermaphrodite germline development require the somatic sheath and spermathecal lineages. Dev Biol 181(2):121–143PubMedCrossRefGoogle Scholar
  123. 123.
    Reinke V, Gil IS, Ward S, Kazmer K (2004) Genome-wide germline-enriched and sex-biased expression profiles in Caenorhabditis elegans. Development 131(2):311–323. doi: 10.1242/dev.00914 PubMedCrossRefGoogle Scholar
  124. 124.
    Wang X, Zhao Y, Wong K, Ehlers P, Kohara Y, Jones SJ, Marra MA, Holt RA, Moerman DG, Hansen D (2009) Identification of genes expressed in the hermaphrodite germ line of C. elegans using SAGE. BMC Genomics 10:213. doi: 10.1186/1471-2164-10-213 PubMedPubMedCentralCrossRefGoogle Scholar
  125. 125.
    Ortiz MA, Noble D, Sorokin EP, Kimble J (2014) A new dataset of spermatogenic vs. oogenic transcriptomes in the nematode Caenorhabditis elegans. G3 (Bethesda) 4(9):1765–1772. doi: 10.1534/g3.114.012351 CrossRefGoogle Scholar
  126. 126.
    Lee M-H, Schedl T (2006) RNA in situ hybridization of dissected gonads. WormBook. doi: 10.1895/wormbook.1.107.1 Google Scholar
  127. 127.
    Suh N, Crittenden SL, Goldstrohm AC, Hook B, Thompson B, Wickens M, Kimble J (2009) FBF and its dual control of gld-1 expression in the Caenorhabditis elegans germline. Genetics 181(4):1249–1260. doi: 10.1534/genetics.108.099440 PubMedPubMedCentralCrossRefGoogle Scholar
  128. 128.
    Sijen T, Fleenor J, Simmer F, Thijssen KL, Parrish S, Timmons L, Plasterk RHA, Fire A (2001) On the role of RNA amplification in dsRNA-triggered gene silencing. Cell 107:465–476PubMedCrossRefGoogle Scholar
  129. 129.
    Kumsta C, Hansen M (2012) C. elegans rrf-1 mutations maintain RNAi efficiency in the soma in addition to the germline. PLoS One 7(5):e35428. doi: 10.1371/journal.pone.0035428 PubMedPubMedCentralCrossRefGoogle Scholar
  130. 130.
    Maine EM, Hansen D, Springer D, Vought VE (2004) Caenorhabditis elegans atx-2 promotes germline proliferation and the oocyte fate. Genetics 168(2):817–830PubMedPubMedCentralCrossRefGoogle Scholar
  131. 131.
    Kim E, Sun L, Gabel CV, Fang-Yen C (2013) Long-term imaging of Caenorhabditis elegans using nanoparticle-mediated immobilization. PLoS One 8(1):e53419. doi: 10.1371/journal.pone.0053419 PubMedPubMedCentralCrossRefGoogle Scholar
  132. 132.
    Rog O, Dernburg AF (2015) Direct visualization reveals kinetics of meiotic chromosome synapsis. Cell Rep. doi: 10.1016/j.celrep.2015.02.032 PubMedPubMedCentralGoogle Scholar
  133. 133.
    Gerhold AR, Ryan J, Vallee-Trudeau JN, Dorn JF, Labbe JC, Maddox PS (2015) Investigating the regulation of stem and progenitor cell mitotic progression by in situ imaging. Curr Biol 25(9):1123–1134. doi: 10.1016/j.cub.2015.02.054 PubMedCrossRefGoogle Scholar
  134. 134.
    San-Miguel A, Lu H (2013) Microfluidics as a tool for C. elegans research. WormBook. pp 1–19. doi: 10.1895/wormbook.1.162.1
  135. 135.
    Shaham S (2006) WormBook: methods in cell biology. WormBook. doi: 10.1895/wormbook.1.41.1 Google Scholar
  136. 136.
    Ji N, van Oudenaarden A (2012) Single molecule fluorescent in situ hybridization (smFISH) of C. elegans worms and embryos. WormBook. pp 1–16. doi: 10.1895/wormbook.1.153.1
  137. 137.
    Kawasaki I, Shim Y-H, Kirchner J, Kaminker J, Wood WB, Strome S (1998) PGL-1, a predicted RNA-binding component of germ granules, is essential for fertility in C. elegans. Cell 94(5):635–645PubMedCrossRefGoogle Scholar
  138. 138.
    Ward S, Roberts TM, Strome S, Pavalko FM, Hogan E (1986) Monoclonal antibodies that recognize a polypeptide antigenic determinant shared by multiple Caenorhabditis elegans sperm-specific proteins. J Cell Biol 102(5):1778–1786PubMedCrossRefGoogle Scholar
  139. 139.
    Sorokin EP, Gasch AP, Kimble J (2014) Competence for chemical reprogramming of sexual fate correlates with an intersexual molecular signature in Caenorhabditis elegans. Genetics 198(2):561–575. doi: 10.1534/genetics.114.169409 PubMedPubMedCentralCrossRefGoogle Scholar
  140. 140.
    Kulkarni M, Shakes DC, Guevel K, Smith HE (2012) SPE-44 implements sperm cell fate. PLoS Genet 8(4):e1002678. doi: 10.1371/journal.pgen.1002678 PubMedPubMedCentralCrossRefGoogle Scholar
  141. 141.
    Grant B, Hirsh D (1999) Receptor-mediated endocytosis in the Caenorhabditis elegans oocyte. Mol Biol Cell 10(12):4311–4326PubMedPubMedCentralCrossRefGoogle Scholar
  142. 142.
    Pasierbek P, Jantsch M, Melcher M, Schleiffer A, Schweizer D, Loidl J (2001) A Caenorhabditis elegans cohesion protein with functions in meiotic chromosome pairing and disjunction. Genes Dev 15(11):1349–1360PubMedPubMedCentralCrossRefGoogle Scholar
  143. 143.
    Cinquin A, Zheng L, Taylor PH, Paz A, Zhang L, Chiang M, Snow JJ, Nie Q, Cinquin O (2015) Semi-permeable diffusion barriers enhance patterning robustness in the C. elegans germline. Dev Cell 35(4):405–417. doi: 10.1016/j.devcel.2015.10.027

Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Sarah L. Crittenden
    • 1
    Email author
  • Hannah S. Seidel
    • 1
  • Judith Kimble
    • 1
  1. 1.HHMI/Department of BiochemistryHoward Hughes Medical Institute and University of Wisconsin-MadisonMadisonUSA

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