Abstract
Phenotypic small molecule screens in zebrafish have gained popularity as an unbiased approach to probe biological processes. In this chapter we outline basic methods for performing chemical screens with larval zebrafish including breeding large numbers of embryos, plating larval fish into multi-well dishes, and adding small molecules to these wells. We also highlight important considerations when designing and interpreting the results of a phenotypic screen and possible follow-up approaches, including popular methods used to identify the mechanism of action of a chemical compound.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Rennekamp AJ, Peterson RT (2015) 15 years of zebrafish chemical screening. Curr Opin Chem Biol 24C:58–70. doi:10.1016/j.cbpa.2014.10.025
Nath AK, Ryu JH, Jin YN, Roberts LD, Dejam A, Gerszten RE, Peterson RT (2015) PTPMT1 inhibition lowers glucose through succinate dehydrogenase phosphorylation. Cell Rep. doi:10.1016/j.celrep.2015.01.010
Liu Y, Asnani A, Zou L, Bentley VL, Yu M, Wang Y, Dellaire G, Sarkar KS, Dai M, Chen HH, Sosnovik DE, Shin JT, Haber DA, Berman JN, Chao W, Peterson RT (2014) Visnagin protects against doxorubicin-induced cardiomyopathy through modulation of mitochondrial malate dehydrogenase. Sci Transl Med 6(266):266ra170. doi:10.1126/scitranslmed.3010189
Yu PB, Hong CC, Sachidanandan C, Babitt JL, Deng DY, Hoyng SA, Lin HY, Bloch KD, Peterson RT (2008) Dorsomorphin inhibits BMP signals required for embryogenesis and iron metabolism. Nat Chem Biol 4(1):33–41
Ridges S, Heaton WL, Joshi D, Choi H, Eiring A, Batchelor L, Choudhry P, Manos EJ, Sofla H, Sanati A, Welborn S, Agarwal A, Spangrude GJ, Miles RR, Cox JE, Frazer JK, Deininger M, Balan K, Sigman M, Muschen M, Perova T, Johnson R, Montpellier B, Guidos CJ, Jones DA, Trede NS (2012) Zebrafish screen identifies novel compound with selective toxicity against leukemia. Blood 119(24):5621–5631. doi:10.1182/blood-2011-12-398818
Gutierrez A, Pan L, Groen RW, Baleydier F, Kentsis A, Marineau J, Grebliunaite R, Kozakewich E, Reed C, Pflumio F, Poglio S, Uzan B, Clemons P, VerPlank L, An F, Burbank J, Norton S, Tolliday N, Steen H, Weng AP, Yuan H, Bradner JE, Mitsiades C, Look AT, Aster JC (2014) Phenothiazines induce PP2A-mediated apoptosis in T cell acute lymphoblastic leukemia. J Clin Invest 124(2):644–655. doi:10.1172/jci65093
Rihel J, Prober DA, Arvanites A, Lam K, Zimmerman S, Jang S, Haggarty SJ, Kokel D, Rubin LL, Peterson RT, Schier AF (2010) Zebrafish behavioral profiling links drugs to biological targets and rest/wake regulation. Science 327(5963):348–351. doi:10.1126/science.1183090
Kokel D, Bryan J, Laggner C, White R, Cheung CY, Mateus R, Healey D, Kim S, Werdich AA, Haggarty SJ, Macrae CA, Shoichet B, Peterson RT (2010) Rapid behavior-based identification of neuroactive small molecules in the zebrafish. Nat Chem Biol 6(3):231–237
North TE, Goessling W, Walkley CR, Lengerke C, Kopani KR, Lord AM, Weber GJ, Bowman TV, Jang IH, Grosser T, Fitzgerald GA, Daley GQ, Orkin SH, Zon LI (2007) Prostaglandin E2 regulates vertebrate haematopoietic stem cell homeostasis. Nature 447(7147):1007–1011
Hagedorn EJ, Durand EM, Fast EM, Zon LI (2014) Getting more for your marrow: boosting hematopoietic stem cell numbers with PGE2. Exp Cell Res 329(2):220–226. doi:10.1016/j.yexcr.2014.07.030
Zon LI, Peterson RT (2005) In vivo drug discovery in the zebrafish. Nat Rev Drug Discov 4(1):35–44. doi:10.1038/nrd1606
Taylor KL, Grant NJ, Temperley ND, Patton EE (2010) Small molecule screening in zebrafish: an in vivo approach to identifying new chemical tools and drug leads. Cell Commun Signal 8:11. doi:10.1186/1478-811X-8-11
Peterson RT, Fishman MC (2011) Designing zebrafish chemical screens. Methods Cell Biol 105:525–541. doi:10.1016/B978-0-12-381320-6.00023-0
Jing L, Durand EM, Ezzio C, Pagliuca SM, Zon LI (2012) In situ hybridization assay-based small molecule screening in zebrafish. Curr Protoc Chem Biol 4(2):110236. doi:10.1002/9780470559277.ch110236
Adatto I, Lawrence C, Thompson M, Zon LI (2011) A new system for the rapid collection of large numbers of developmentally staged zebrafish embryos. PLoS One 6(6):e21715. doi:10.1371/journal.pone.0021715
Bruni G, Lakhani P, Kokel D (2014) Discovering novel neuroactive drugs through high-throughput behavior-based chemical screening in the zebrafish. Front Pharmacol 5:153. doi:10.3389/fphar.2014.00153
Rovira M, Huang W, Yusuff S, Shim JS, Ferrante AA, Liu JO, Parsons MJ (2011) Chemical screen identifies FDA-approved drugs and target pathways that induce precocious pancreatic endocrine differentiation. Proc Natl Acad Sci U S A 108(48):19264–19269. doi:10.1073/pnas.1113081108
Molina G, Vogt A, Bakan A, Dai W, Queiroz de Oliveira P, Znosko W, Smithgall TE, Bahar I, Lazo JS, Day BW, Tsang M (2009) Zebrafish chemical screening reveals an inhibitor of Dusp6 that expands cardiac cell lineages. Nat Chem Biol 5(9):680–687. doi:10.1038/nchembio.190
Peal DS, Mills RW, Lynch SN, Mosley JM, Lim E, Ellinor PT, January CT, Peterson RT, Milan DJ (2011) Novel chemical suppressors of long QT syndrome identified by an in vivo functional screen. Circulation 123(1):23–30. doi:10.1161/circulationaha.110.003731
Peterson RT, Link BA, Dowling JE, Schreiber SL (2000) Small molecule developmental screens reveal the logic and timing of vertebrate development. Proc Natl Acad Sci U S A 97(24):12965–12969
Yeh JR, Munson KM, Chao YL, Peterson QP, Macrae CA, Peterson RT (2008) AML1-ETO reprograms hematopoietic cell fate by downregulating scl expression. Development 135(2):401–410. doi:10.1242/dev.008904
Gut P, Baeza-Raja B, Andersson O, Hasenkamp L, Hsiao J, Hesselson D, Akassoglou K, Verdin E, Hirschey MD, Stainier DY (2013) Whole-organism screening for gluconeogenesis identifies activators of fasting metabolism. Nat Chem Biol 9(2):97–104. doi:10.1038/nchembio.1136
Rennekamp AJ, Peterson RT (2013) From phenotype to mechanism after zebrafish small molecule screens. Drug Discov Today Dis Models 10(1):e51–e55. doi:10.1016/j.ddmod.2012.02.002
Kokel D, Rennekamp AJ, Shah AH, Liebel U, Peterson RT (2012) Behavioral barcoding in the cloud: embracing data-intensive digital phenotyping in neuropharmacology. Trends Biotechnol 30(8):421–425. doi:10.1016/j.tibtech.2012.05.001
Hwang WY, Fu Y, Reyon D, Maeder ML, Tsai SQ, Sander JD, Peterson RT, Yeh JR, Joung JK (2013) Efficient genome editing in zebrafish using a CRISPR-Cas system. Nat Biotechnol 31(3):227–229. doi:10.1038/nbt.2501
Hwang WY, Fu Y, Reyon D, Maeder ML, Kaini P, Sander JD, Joung JK, Peterson RT, Yeh JR (2013) Heritable and precise zebrafish genome editing using a CRISPR-Cas system. PLoS One 8(7):e68708. doi:10.1371/journal.pone.0068708
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2016 Springer Science+Business Media New York
About this protocol
Cite this protocol
Brady, C.A., Rennekamp, A.J., Peterson, R.T. (2016). Chemical Screening in Zebrafish. In: Kawakami, K., Patton, E., Orger, M. (eds) Zebrafish. Methods in Molecular Biology, vol 1451. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3771-4_1
Download citation
DOI: https://doi.org/10.1007/978-1-4939-3771-4_1
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3769-1
Online ISBN: 978-1-4939-3771-4
eBook Packages: Springer Protocols