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Assessing Ovulation in Drosophila melanogaster

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Drosophila Oogenesis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2626))

Abstract

Ovulation is a critical reproductive process by which a mature oocyte is released from the ovary for fertilization. This process requires the coordination of multiple cellular and molecular events including the spatiotemporal breakdown of the follicle wall. Recent work has shown that ovulation in Drosophila utilizes conserved cellular processes and molecular pathways as in mammals. Thus, Drosophila ovulation can serve as a good model to decipher the fundamental mechanisms of ovulation utilized across species. In past decades, several methods have been developed to study Drosophila ovulation, but all of them have drawbacks. This chapter offers a strategy and detailed protocols for performing and analyzing the necessary assays to evaluate the ovulation process in Drosophila.

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References

  1. Espey LL (2006) Comprehensive analysis of ovarian gene expression during ovulation using differential display. Methods Mol Biol 317:219–241. https://doi.org/10.1385/1-59259-968-0:219

  2. Duffy DM, Ko C, Jo M, Brannstrom M, Curry TE (2019) Ovulation: parallels with inflammatory processes. Endocr Rev 40(2):369–416. https://doi.org/10.1210/er.2018-00075

    Article  Google Scholar 

  3. Fan HY, Liu Z, Mullany LK, Richards JS (2012) Consequences of RAS and MAPK activation in the ovary: the good, the bad and the ugly. Mol Cell Endocrinol 356(1–2):74–79. https://doi.org/10.1016/j.mce.2011.12.005

    Article  CAS  Google Scholar 

  4. Robker RL, Hennebold JD, Russell DL (2018) Coordination of ovulation and oocyte maturation: a good egg at the right time. Endocrinology 159(9):3209–3218. https://doi.org/10.1210/en.2018-00485

    Article  CAS  Google Scholar 

  5. Deady LD, Shen W, Mosure SA, Spradling AC, Sun J (2015) Matrix metalloproteinase 2 is required for ovulation and corpus luteum formation in Drosophila. PLoS Genet 11(2):e1004989. https://doi.org/10.1371/journal.pgen.1004989

    Article  CAS  Google Scholar 

  6. Deady LD, Sun J (2015) A follicle rupture assay reveals an essential role for follicular adrenergic signaling in Drosophila ovulation. PLoS Genet 11(10):e1005604. https://doi.org/10.1371/journal.pgen.1005604

    Article  CAS  Google Scholar 

  7. Li W, Young JF, Sun J (2018) NADPH oxidase-generated reactive oxygen species in mature follicles are essential for Drosophila ovulation. Proc Natl Acad Sci U S A 115(30):7765–7770. https://doi.org/10.1073/pnas.1800115115

    Article  CAS  Google Scholar 

  8. Knapp E, Sun J (2017) Steroid signaling in mature follicles is important for Drosophila ovulation. Proc Natl Acad Sci U S A 114(4):699–704. https://doi.org/10.1073/pnas.1614383114

    Article  CAS  Google Scholar 

  9. Jiang K, Zhang J, Huang Y, Wang Y, Xiao S, Hadden MK, Woodruff TK, Sun J (2021) A platform utilizing Drosophila ovulation for nonhormonal contraceptive screening. Proc Natl Acad Sci U S A 118(28):e2026403118. https://doi.org/10.1073/pnas.2026403118

    Article  CAS  Google Scholar 

  10. Aigaki T, Fleischmann I, Chen PS, Kubli E (1991) Ectopic expression of sex peptide alters reproductive behavior of female D. melanogaster. Neuron 7(4):557–563. https://doi.org/10.1016/0896-6273(91)90368-a

    Article  CAS  Google Scholar 

  11. Heifetz Y, Lung O, Frongillo EA Jr, Wolfner MF (2000) The Drosophila seminal fluid protein Acp26Aa stimulates release of oocytes by the ovary. Curr Biol 10(2):99–102. https://doi.org/10.1016/s0960-9822(00)00288-8

    Article  CAS  Google Scholar 

  12. Lee HG, Rohila S, Han KA (2009) The octopamine receptor OAMB mediates ovulation via Ca2+/calmodulin-dependent protein kinase II in the Drosophila oviduct epithelium. PLoS One 4(3):e4716. https://doi.org/10.1371/journal.pone.0004716

    Article  CAS  Google Scholar 

  13. Lim J, Sabandal PR, Fernandez A, Sabandal JM, Lee HG, Evans P, Han KA (2014) The octopamine receptor Octbeta2R regulates ovulation in Drosophila melanogaster. PLoS One 9(8):e104441. https://doi.org/10.1371/journal.pone.0104441

    Article  CAS  Google Scholar 

  14. Cole SH, Carney GE, McClung CA, Willard SS, Taylor BJ, Hirsh J (2005) Two functional but noncomplementing Drosophila tyrosine decarboxylase genes: distinct roles for neural tyramine and octopamine in female fertility. J Biol Chem 280(15):14948–14955. https://doi.org/10.1074/jbc.M414197200

    Article  CAS  Google Scholar 

  15. Monastirioti M (2003) Distinct octopamine cell population residing in the CNS abdominal ganglion controls ovulation in Drosophila melanogaster. Dev Biol 264(1):38–49. https://doi.org/10.1016/j.ydbio.2003.07.019

    Article  CAS  Google Scholar 

  16. Monastirioti M, Linn CE Jr, White K (1996) Characterization of Drosophila tyramine beta-hydroxylase gene and isolation of mutant flies lacking octopamine. J Neurosci 16(12):3900–3911

    Article  CAS  Google Scholar 

  17. Ritsick DR, Edens WA, Finnerty V, Lambeth JD (2007) Nox regulation of smooth muscle contraction. Free Radic Biol Med 43(1):31–38. https://doi.org/10.1016/j.freeradbiomed.2007.03.006

    Article  CAS  Google Scholar 

  18. Liao S, Nassel DR (2020) Drosophila insulin-like peptide 8 (DILP8) in ovarian follicle cells regulates ovulation and metabolism. Front Endocrinol (Lausanne) 11:461. https://doi.org/10.3389/fendo.2020.00461

    Article  Google Scholar 

  19. Deady LD, Li W, Sun J (2017) The zinc-finger transcription factor Hindsight regulates ovulation competency of Drosophila follicles. Elife 6:e29887. https://doi.org/10.7554/eLife.29887

    Article  Google Scholar 

  20. Luo W, Liu S, Zhang W, Yang L, Huang J, Zhou S, Feng Q, Palli SR, Wang J, Roth S, Li S (2021) Juvenile hormone signaling promotes ovulation and maintains egg shape by inducing expression of extracellular matrix genes. Proc Natl Acad Sci U S A 118(39):e2104461118. https://doi.org/10.1073/pnas.2104461118

    Article  CAS  Google Scholar 

  21. Rubinstein CD, Wolfner MF (2013) Drosophila seminal protein ovulin mediates ovulation through female octopamine neuronal signaling. Proc Natl Acad Sci U S A 110(43):17420–17425. https://doi.org/10.1073/pnas.1220018110

    Article  Google Scholar 

  22. Knapp EM, Li W, Singh V, Sun J (2020) Nuclear receptor Ftz-f1 promotes follicle maturation and ovulation partly via bHLH/PAS transcription factor Sim. Elife 9:e54568. https://doi.org/10.7554/eLife.54568

    Article  CAS  Google Scholar 

  23. Knapp EM, Deady LD, Sun J (2018) Ex vivo follicle rupture and in situ Zymography in Drosophila. Bio Protoc 8(10):e2846. https://doi.org/10.21769/BioProtoc.2846

    Article  CAS  Google Scholar 

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Acknowledgments

We thank members of Sun Lab for their careful reading and suggestions. This work is supported by the National Institute of Child Health and Human Development grants (R01-HD086175 and R01-HD097206) to J.S.

Author information

Authors and Affiliations

  1. Department of Physiology & Neurobiology, University of Connecticut, Storrs, CT, USA

    Andrew Beard, Rebecca Oramas & Jianjun Sun

  2. Institute for Systems Genomics, University of Connecticut, Storrs, CT, USA

    Jianjun Sun

Authors
  1. Andrew Beard
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  2. Rebecca Oramas
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  3. Jianjun Sun
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Corresponding author

Correspondence to Jianjun Sun .

Editor information

Editors and Affiliations

  1. Department of Anatomy & Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, IA, USA

    Michelle S. Giedt

  2. Department of Anatomy & Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, IA, USA

    Tina L. Tootle

1 Electronic Supplementary Material

Supplemental 1

Assessing Ovulation Template. The blank Excel template used in our lab to store and analyze all the ovulation assay data is described in this chapter. There are six sheets within the Excel file: egg-laying, egg-laying summary, mature follicles, egg distribution, egg-laying time, and OA culture. The “egg-laying” sheet is where the experimental conditions and number of eggs counted on each plate are inputted to calculate the average eggs laid per female per day for each genotype. The next sheet “egg-laying summary” is the summary of all the egg-laying data simplified along with the associated graph. The “mature follicles” sheet is where the experimental conditions and number of mature follicles per the top and bottom ovary are entered to calculate the average and generate a graph. The “egg distribution” sheet is where the experimental conditions and scores (e.g., N, C, B, E, and U) from the egg laying time assay are entered for each genotype. The data from the “egg-laying summary” and “egg distribution” sheets are then populated into the “egg-laying time” sheet to calculate ovulation time, uterus time, and oviduct time, respectively, which ultimately generates a graph. The “OA culture” sheet is where the experimental conditions, the total number of follicles, and the number of ruptured follicles are entered for each genotype to calculate the percentage of ruptured follicles in each condition (XLSX 61 kb)

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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Beard, A., Oramas, R., Sun, J. (2023). Assessing Ovulation in Drosophila melanogaster. In: Giedt, M.S., Tootle, T.L. (eds) Drosophila Oogenesis. Methods in Molecular Biology, vol 2626. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2970-3_13

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  • DOI: https://doi.org/10.1007/978-1-0716-2970-3_13

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2969-7

  • Online ISBN: 978-1-0716-2970-3

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