Archives of Toxicology

, Volume 88, Issue 3, pp 543–551 | Cite as

Dispersed crude oil amplifies germ cell apoptosis in Caenorhabditis elegans, followed a CEP-1-dependent pathway



The Deepwater Horizon oil spill is among the most severe environmental disasters in US history. The extent of crude oil released and the subsequent dispersant used for cleanup was unprecedented. The dispersed crude oil represents a unique form of environmental contaminant that warrants investigations of its environmental and human health impacts. Lines of evidence have demonstrated that dispersed oil affects reproduction in various organisms, in a more potent manner than oil- and dispersant-only exposures. However, the action mechanism of dispersed oil remains largely unknown. In this study, we utilized the model organism Caenorhabditis elegans to investigate impacts of dispersed oil exposure on sex cell apoptosis and related gene expressions. Worms were exposed to different diluted levels of crude oil–dispersant (oil–dis) mixtures (20:1, v/v; at 500×, 2,000×, and 5,000× dilutions). The dispersed crude oil significantly increases the number of apoptotic germ cells in treated worms when compared with control at all exposure levels (p < 0.05). Genes involved in the apoptosis pathway were dysregulated, which include ced-13,ced-3, ced-4, ced-9, cep-1,dpl-1, efl-1, efl-2, egl-1, egl-38, lin-35, pax-2, and sir-2.1. Many aberrant expressed genes encoding for core components in apoptosis machinery (cep-1/p53, ced-13/BH3, ced-9/Bcl-2, ced-4/Apaf-1, and ced-3/caspase) displayed consistent expression patterns across all exposure levels. Significantly ced-3/caspase was upregulated at all dispersed oil-treated groups, consistent with the observed apoptosis phenotype. Given cep-1/p53 was activated at all dispersed oil treatments and the germ cell apoptosis was suppressed in the CEP-1 loss of function mutant, the increased apoptosis is likely CEP-1 dependent. In addition, the anti-apoptotic ced-9/Bcl-2 was activated in response to the increase in cell death. This study provides a mechanism understanding of dispersed crude oil-induced reproductive toxicity.


Deepwater Horizon (DWH) oil spill C. elegans Apoptosis Gene expression cep-1 pathway 


  1. Aballay A, Ausubel FM (2001) Programmed cell death mediated by ced-3 and ced-4 protects Caenorhabditis elegans from Salmonella typhimurium-mediated killing. Proc Natl Acad Sci USA 98(5):2735–2739. doi:10.1073/pnas.041613098 PubMedCrossRefGoogle Scholar
  2. Almeda R, Wambaugh Z, Wang Z, Hyatt C, Liu Z, Buskey EJ (2013) Interactions between zooplankton and crude oil: toxic effects and bioaccumulation of polycyclic aromatic hydrocarbons. PLoS ONE 8(6) doi:10.1371/journal.pone.0067212
  3. Anderson SE, Franko J, Lukomska E, Meade BJ (2011) Potential immunotoxicological health effects following exposure to COREXIT 9500A during cleanup of the Deepwater Horizon oil spill. J Toxicol Environ Health Part A 74(21):1419–1430. doi:10.1080/15287394.2011.606797 PubMedCrossRefGoogle Scholar
  4. Bailly A, Gartner A (2013) Germ cell apoptosis and DNA damage responses. Adv Exp Med Biol 757:249–276. doi:10.1007/978-1-4614-4015-4_9 PubMedCrossRefGoogle Scholar
  5. Bossy-Wetzel E, Newmeyer DD, Green DR (1998) Mitochondrial cytochrome c release in apoptosis occurs upstream of DEVD-specific caspase activation and independently of mitochondrial transmembrane depolarization. EMBO J 17(1):37–49. doi:10.1093/emboj/17.1.37 PubMedCrossRefGoogle Scholar
  6. Boxem M, van den Heuvel S (2002) C. elegans class B synthetic multivulva genes act in G(1) regulation. Curr Biol CB 12(11):906–911CrossRefGoogle Scholar
  7. Brenner S (1974) The genetics of Caenorhabditis elegans. Genetics 77(1):71–94PubMedGoogle Scholar
  8. Brodigan TM, Liu J, Park M, Kipreos ET, Krause M (2003) Cyclin E expression during development in Caenorhabditis elegans. Dev Biol 254(1):102–115PubMedCrossRefGoogle Scholar
  9. C. elegans Sequencing Consortium (1998) Genome sequence of the nematode C. elegans: a platform for investigating biology. Science 282(5396):2012–2018Google Scholar
  10. Ceol CJ, Horvitz HR (2001) dpl-1 DP and efl-1 E2F act with lin-35 Rb to antagonize Ras signaling in C. elegans vulval development. Mol Cell 7(3):461–473PubMedCrossRefGoogle Scholar
  11. Chamberlin HM, Palmer RE, Newman AP, Sternberg PW, Baillie DL, Thomas JH (1997) The PAX gene egl-38 mediates developmental patterning in Caenorhabditis elegans. Development 124(20):3919–3928PubMedGoogle Scholar
  12. Chen F, Hersh BM, Conradt B et al (2000) Translocation of C. elegans CED-4 to nuclear membranes during programmed cell death. Science 287(5457):1485–1489PubMedCrossRefGoogle Scholar
  13. Chi W, Reinke V (2009) DPL-1 (DP) acts in the germ line to coordinate ovulation and fertilization in C. elegans. Mech Dev 126(5–6):406–416. doi:10.1016/j.mod.2009.01.008 PubMedCentralPubMedCrossRefGoogle Scholar
  14. Conradt B, Horvitz HR (1998) The C-elegans protein EGL-1 is required for programmed cell death and interacts with the Bcl-2-like protein CED-9. Cell 93(4):519–529. doi:10.1016/S0092-8674(00)81182-4 PubMedCrossRefGoogle Scholar
  15. Cutter AD, Dey A, Murray RL (2009) Evolution of the Caenorhabditis elegans genome. Mol Biol Evol 26(6):1199–1234. doi:10.1093/molbev/msp048 PubMedCrossRefGoogle Scholar
  16. De Flora S, Bagnasco M, Zanacchi P (1991) Genotoxic, carcinogenic, and teratogenic hazards in the marine environment, with special reference to the Mediterranean Sea. Mutat Res 258(3):285–320PubMedCrossRefGoogle Scholar
  17. de Soysa TY, Ulrich A, Friedrich T et al (2012) Macondo crude oil from the Deepwater Horizon oil spill disrupts specific developmental processes during zebrafish embryogenesis. BMC Biol 10:40. doi:10.1186/1741-7007-10-40 PubMedCentralPubMedCrossRefGoogle Scholar
  18. Denning DP, Hatch V, Horvitz HR (2013) Both the Caspase CSP-1 and a Caspase-independent pathway promote programmed cell death in parallel to the canonical pathway for apoptosis in Caenorhabditis elegans. Plos Genet 9(3) doi:10.1371/journal.pgen.1003341
  19. Derry WB, Putzke AP, Rothman JH (2001) Caenorhabditis elegans p53: role in apoptosis, meiosis, and stress resistance. Science 294(5542):591–595. doi:10.1126/science.1065486 PubMedCrossRefGoogle Scholar
  20. Finch BE, Wooten KJ, Faust DR, Smith PN (2012) Embryotoxicity of mixtures of weathered crude oil collected from the Gulf of Mexico and Corexit 9500 in mallard ducks (Anas platyrhynchos). Sci Total Environ 426:155–159. doi:10.1016/j.scitotenv.2012.03.070 PubMedCrossRefGoogle Scholar
  21. Gartner A, Boag PR, Blackwell TK (2008) Germline survival and apoptosis. WormBook: the online review of C. elegans biology:1–20 doi:10.1895/wormbook.1.145.1
  22. Goodbody-Gringley G, Wetzel DL, Gillon D, Pulster E, Miller A, Ritchie KB (2013) Toxicity of Deepwater Horizon source oil and the chemical dispersant, Corexit(R) 9500, to coral larvae. PLoS ONE 8(1):e45574. doi:10.1371/journal.pone.0045574 PubMedCentralPubMedCrossRefGoogle Scholar
  23. 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
  24. Hall DH, Altun ZF (2008) C. elegans atlas. Cold Spring Harbor Laboratory Press, Cold Spring HarborGoogle Scholar
  25. Hall PA, Lane DP (1997) Tumor suppressors: a developing role for p53? Curr Biol CB 7(3):R144–R147CrossRefGoogle Scholar
  26. Hengartner MO (1997) Genetic control of programmed cell death and aging in the nematode Caenorhabditis elegans. Exp Gerontol 32(4–5):363–374PubMedCrossRefGoogle Scholar
  27. Hockenbery DM (1994) bcl-2 in cancer, development and apoptosis. J Cell Sci Supplement 18:51–55CrossRefGoogle Scholar
  28. Jiang X, Wang X (2004) Cytochrome C-mediated apoptosis. Annu Rev Biochem 73:87–106. doi:10.1146/annurev.biochem.73.011303.073706 PubMedCrossRefGoogle Scholar
  29. Kaletta T, Hengartner MO (2006) Finding function in novel targets: C. elegans as a model organism. Nat Rev Drug Discov 5(5):387–398. doi:10.1038/nrd2031 Google Scholar
  30. Kenyon CJ (2010) The genetics of ageing. Nature 464(7288):504–512. doi:10.1038/nature08980 PubMedCrossRefGoogle Scholar
  31. L’Hernault SW (2009) The genetics and cell biology of spermatogenesis in the nematode C. elegans. Mol Cell Endocrinol 306(1–2):59–65. doi:10.1016/j.mce.2009.01.008 PubMedCrossRefGoogle Scholar
  32. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods 25(4):402–408. doi:10.1006/meth 2001.1262PubMedCrossRefGoogle Scholar
  33. Lord CE, Gunawardena AH (2012) Programmed cell death in C. elegans, mammals and plants. Eur J Cell Biol 91(8):603–613. doi:10.1016/j.ejcb.2012.02.002 PubMedCrossRefGoogle Scholar
  34. Ludewig AH, Izrayelit Y, Park D et al (2013) Pheromone sensing regulates Caenorhabditis elegans lifespan and stress resistance via the deacetylase SIR-2.1. Proc Natl Acad Sci USA 110(14):5522–5527. doi:10.1073/pnas.1214467110 PubMedCrossRefGoogle Scholar
  35. Park D, Jia H, Rajakumar V, Chamberlin HM (2006) Pax2/5/8 proteins promote cell survival in C. elegans. Development 133(21):4193–4202. doi:10.1242/dev.02614 PubMedCrossRefGoogle Scholar
  36. Pourkarimi E, Greiss S (2011) Gartner A (2012), Evidence that CED-9/Bcl2 and CED-4/Apaf-1 localization is not consistent with the current model for C. elegans apoptosis induction. Cell Death Differ 19(3):406–415. doi:10.1038/Cdd.104 PubMedCrossRefGoogle Scholar
  37. Radniecki TS, Schneider MC, Semprini L (2013) The influence of Corexit 9500A and weathering on Alaska North Slope crude oil toxicity to the ammonia oxidizing bacterium, Nitrosomonas europaea. Mar Pollut Bullet 68(1–2):64–70. doi:10.1016/j.marpolbul.2012.12.022 CrossRefGoogle Scholar
  38. Reddien PW, Andersen EC, Huang MC, Horvitz HR (2007) DPL-1 DP, LIN-35 Rb and EFL-1 E2F act with the MCD-1 zinc-finger protein to promote programmed cell death in Caenorhabditis elegans. Genetics 175(4):1719–1733. doi:10.1534/genetics.106.068148 PubMedCrossRefGoogle Scholar
  39. Ruan QL, Ju JJ, Li YH et al (2012) Chlorpyrifos exposure reduces reproductive capacity owing to a damaging effect on gametogenesis in the nematode Caenorhabditis elegans. J Appl Toxicol JAT 32(7):527–535. doi:10.1002/jat.1783 CrossRefGoogle Scholar
  40. 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
  41. Schertel C, Conradt B (2007) C. elegans orthologs of components of the RB tumor suppressor complex have distinct pro-apoptotic functions. Development 134(20):3691–3701. doi:10.1242/dev.004606 PubMedCrossRefGoogle Scholar
  42. Schumacher B, Hofmann K, Boulton S, Gartner A (2001) The C. elegans homolog of the p53 tumor suppressor is required for DNA damage-induced apoptosis. Curr Biol 11(21):1722–1727. doi:10.1016/S0960-9822(01)00534-6 PubMedCrossRefGoogle Scholar
  43. Schumacher B, Schertel C, Wittenburg N et al (2005) C-elegans ced-13 can promote apoptosis and is induced in response to DNA damage. Cell Death Differ 12(2):153–161. doi:10.1038/sj.cdd.4401539 PubMedCrossRefGoogle Scholar
  44. Shaham S, Horvitz HR (1996) An alternatively spliced C-elegans ced-4 RNA encodes a novel cell death inhibitor. Cell 86(2):201–208. doi:10.1016/S0092-8674(00)80092-6 PubMedCrossRefGoogle Scholar
  45. Shaham S, Reddien PW, Davies B, Horvitz HR (1999) Mutational analysis of the Caenorhabditis elegans cell-death gene ced-3. Genetics 153(4):1655–1671PubMedGoogle Scholar
  46. Speight JG (1999) The chemistry and technology of petroleum, 3rd edn. Marcel Dekker, New YorkCrossRefGoogle Scholar
  47. Xue D, Shaham S, Horvitz HR (1996) The Caenorhabditis elegans cell-death protein CED-3 is a cysteine protease with substrate specificities similar to those of the human CPP32 protease. Genes Dev 10(9):1073–1083PubMedCrossRefGoogle Scholar
  48. Zhang Y, Chen D, Smith MA, Zhang B, Pan X (2012) Selection of reliable reference genes in Caenorhabditis elegans for analysis of nanotoxicity. PLoS ONE 7(3):e31849. doi:10.1371/journal.pone.0031849 PubMedCentralPubMedCrossRefGoogle Scholar
  49. Zhang YQ, Chen DL, Ennis AC et al (2013) Chemical dispersant potentiates crude oil impacts on growth, reproduction, and gene expression in Caenorhabditis elegans. Arch Toxicol 87(2):371–382. doi:10.1007/s00204-012-0936-x PubMedCrossRefGoogle Scholar
  50. Zou H, Henzel WJ, Liu X, Lutschg A, Wang X (1997) Apaf-1, a human protein homologous to C. elegans CED-4, participates in cytochrome c-dependent activation of caspase-3. Cell 90(3):405–413PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Joseph Ryan Polli
    • 1
  • Yanqiong Zhang
    • 1
  • Xiaoping Pan
    • 1
  1. 1.Department of BiologyEast Carolina UniversityGreenvilleUSA

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