Functional assessment of the “two-hit” model for neurodevelopmental defects in Drosophila and X. laevis

Autoři: Lucilla Pizzo aff001;  Micaela Lasser aff002;  Tanzeen Yusuff aff001;  Matthew Jensen aff001;  Phoebe Ingraham aff001;  Emily Huber aff001;  Mayanglambam Dhruba Singh aff001;  Connor Monahan aff002;  Janani Iyer aff001;  Inshya Desai aff001;  Siddharth Karthikeyan aff001;  Dagny J. Gould aff001;  Sneha Yennawar aff001;  Alexis T. Weiner aff001;  Vijay Kumar Pounraja aff001;  Arjun Krishnan aff003;  Melissa M. Rolls aff001;  Laura Anne Lowery aff005;  Santhosh Girirajan aff001
Působiště autorů: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA, United States of America aff001;  Department of Biology, Boston College, Chestnut Hill, MA, United States of America aff002;  Department of Computational Mathematics, Science and Engineering, Michigan State University, East Lansing, MI, United States of America aff003;  Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI, United States of America aff004;  Department of Medicine, Boston University Medical Center, Boston, MA, United States of America aff005;  Department of Anthropology, The Pennsylvania State University, University Park, PA, United States of America aff006
Vyšlo v časopise: Functional assessment of the “two-hit” model for neurodevelopmental defects in Drosophila and X. laevis. PLoS Genet 17(4): e1009112. doi:10.1371/journal.pgen.1009112
Kategorie: Research Article


We previously identified a deletion on chromosome 16p12.1 that is mostly inherited and associated with multiple neurodevelopmental outcomes, where severely affected probands carried an excess of rare pathogenic variants compared to mildly affected carrier parents. We hypothesized that the 16p12.1 deletion sensitizes the genome for disease, while “second-hits” in the genetic background modulate the phenotypic trajectory. To test this model, we examined how neurodevelopmental defects conferred by knockdown of individual 16p12.1 homologs are modulated by simultaneous knockdown of homologs of “second-hit” genes in Drosophila melanogaster and Xenopus laevis. We observed that knockdown of 16p12.1 homologs affect multiple phenotypic domains, leading to delayed developmental timing, seizure susceptibility, brain alterations, abnormal dendrite and axonal morphology, and cellular proliferation defects. Compared to genes within the 16p11.2 deletion, which has higher de novo occurrence, 16p12.1 homologs were less likely to interact with each other in Drosophila models or a human brain-specific interaction network, suggesting that interactions with “second-hit” genes may confer higher impact towards neurodevelopmental phenotypes. Assessment of 212 pairwise interactions in Drosophila between 16p12.1 homologs and 76 homologs of patient-specific “second-hit” genes (such as ARID1B and CACNA1A), genes within neurodevelopmental pathways (such as PTEN and UBE3A), and transcriptomic targets (such as DSCAM and TRRAP) identified genetic interactions in 63% of the tested pairs. In 11 out of 15 families, patient-specific “second-hits” enhanced or suppressed the phenotypic effects of one or many 16p12.1 homologs in 32/96 pairwise combinations tested. In fact, homologs of SETD5 synergistically interacted with homologs of MOSMO in both Drosophila and X. laevis, leading to modified cellular and brain phenotypes, as well as axon outgrowth defects that were not observed with knockdown of either individual homolog. Our results suggest that several 16p12.1 genes sensitize the genome towards neurodevelopmental defects, and complex interactions with “second-hit” genes determine the ultimate phenotypic manifestation.

Klíčová slova:

Drosophila melanogaster – Eyes – Genetic interactions – Larvae – Phenotypes – RNA interference – Xenopus – Morpholino


1. Girirajan S, Campbell CD, Eichler EE. Human copy number variation and complex genetic disease. Annu Rev Genet. 2011;45:203–26. doi: 10.1146/annurev-genet-102209-163544 21854229

2. Wilfert AB, Sulovari A, Turner TN, Coe BP, Eichler EE. Recurrent de novo mutations in neurodevelopmental disorders: properties and clinical implications. Genome Med. 2017;9(1):101. doi: 10.1186/s13073-017-0498-x 29179772

3. Weiss LA, Shen Y, Korn JM, Arking DE, Miller DT, Fossdal R, et al. Association between microdeletion and microduplication at 16p11.2 and autism. N Engl J Med. 2008;358(7):667–75. doi: 10.1056/NEJMoa075974 18184952

4. Zufferey F, Sherr EH, Beckmann ND, Hanson E, Maillard AM, Hippolyte L, et al. A 600 kb deletion syndrome at 16p11. 2 leads to energy imbalance and neuropsychiatric disorders. Journal of medical genetics. 2012;49(10):660–8. doi: 10.1136/jmedgenet-2012-101203 23054248

5. Mulle JG. The 3q29 deletion confers >40-fold increase in risk for schizophrenia. Mol Psychiatry. 2015;20(9):1028–9. doi: 10.1038/mp.2015.76 26055425

6. Helbig I, Mefford HC, Sharp AJ, Guipponi M, Fichera M, Franke A, et al. 15q13.3 microdeletions increase risk of idiopathic generalized epilepsy. Nat Genet. 2009;41(2):160–2. doi: 10.1038/ng.292 19136953

7. Girirajan S, Eichler EE. Phenotypic variability and genetic susceptibility to genomic disorders. Human molecular genetics. 2010;19(R2):R176–R87. doi: 10.1093/hmg/ddq366 20807775

8. Girirajan S, Rosenfeld JA, Cooper GM, Antonacci F, Siswara P, Itsara A, et al. A recurrent 16p12. 1 microdeletion supports a two-hit model for severe developmental delay. Nature genetics. 2010;42(3):203–9. doi: 10.1038/ng.534 20154674

9. Pizzo L, Jensen M, Polyak A, Rosenfeld JA, Mannik K, Krishnan A, et al. Rare variants in the genetic background modulate cognitive and developmental phenotypes in individuals carrying disease-associated variants. Genet Med. 2018. doi: 10.1038/s41436-018-0266-3 30190612

10. Stefansson H, Meyer-Lindenberg A, Steinberg S, Magnusdottir B, Morgen K, Arnarsdottir S, et al. CNVs conferring risk of autism or schizophrenia affect cognition in controls. Nature. 2014;505(7483):361–6. doi: 10.1038/nature12818 24352232

11. Girirajan S, Rosenfeld JA, Coe BP, Parikh S, Friedman N, Goldstein A, et al. Phenotypic heterogeneity of genomic disorders and rare copy-number variants. New England Journal of Medicine. 2012;367(14):1321–31. doi: 10.1056/NEJMoa1200395 22970919

12. Gatto CL, Pereira D, Broadie K. GABAergic circuit dysfunction in the Drosophila Fragile X syndrome model. Neurobiol Dis. 2014;65:142–59. doi: 10.1016/j.nbd.2014.01.008 24423648

13. Jumbo-Lucioni PP, Parkinson WM, Kopke DL, Broadie K. Coordinated movement, neuromuscular synaptogenesis and trans-synaptic signaling defects in Drosophila galactosemia models. Hum Mol Genet. 2016;25(17):3699–714. doi: 10.1093/hmg/ddw217 27466186

14. Sears JC, Broadie K. Fragile X Mental Retardation Protein Regulates Activity-Dependent Membrane Trafficking and Trans-Synaptic Signaling Mediating Synaptic Remodeling. Front Mol Neurosci. 2017;10:440. doi: 10.3389/fnmol.2017.00440 29375303

15. Grossman TR, Gamliel A, Wessells RJ, Taghli-Lamallem O, Jepsen K, Ocorr K, et al. Over-expression of DSCAM and COL6A2 cooperatively generates congenital heart defects. PLoS Genet. 2011;7(11):e1002344. doi: 10.1371/journal.pgen.1002344 22072978

16. Chen SX, Tari PK, She K, Haas K. Neurexin-neuroligin cell adhesion complexes contribute to synaptotropic dendritogenesis via growth stabilization mechanisms in vivo. Neuron. 2010;67(6):967–83. doi: 10.1016/j.neuron.2010.08.016 20869594

17. Lewis BB, Wester MR, Miller LE, Nagarkar MD, Johnson MB, Saha MS. Cloning and characterization of voltage-gated calcium channel alpha1 subunits in Xenopus laevis during development. Dev Dyn. 2009;238(11):2891–902. doi: 10.1002/dvdy.22102 19795515

18. Ishimaru H, Kamboj R, Ambrosini A, Henley JM, Soloviev MM, Sudan H, et al. A unitary non-NMDA receptor short subunit from Xenopus: DNA cloning and expression. Receptors Channels. 1996;4(1):31–49. 8723645

19. Ueno S, Kono R, Iwao Y. PTEN is required for the normal progression of gastrulation by repressing cell proliferation after MBT in Xenopus embryos. Dev Biol. 2006;297(1):274–83. doi: 10.1016/j.ydbio.2006.06.001 16919259

20. Iyer J, Singh MD, Jensen M, Patel P, Pizzo L, Huber E, et al. Pervasive genetic interactions modulate neurodevelopmental defects of the autism-associated 16p11.2 deletion in Drosophila melanogaster. Nat Commun. 2018;9(1):2548. doi: 10.1038/s41467-018-04882-6 29959322

21. Singh MD, Jensen M, Lasser M, Huber E, Yusuff T, Pizzo L, et al. NCBP2 modulates neurodevelopmental defects of the 3q29 deletion in Drosophila and Xenopus laevis models. PLoS Genet. 2020;16(2):e1008590. doi: 10.1371/journal.pgen.1008590 32053595

22. McCammon JM, Blaker-Lee A, Chen X, Sive H. The 16p11.2 homologs fam57ba and doc2a generate certain brain and body phenotypes. Hum Mol Genet. 2017;26(19):3699–712. doi: 10.1093/hmg/ddx255 28934389

23. Qiu Y, Arbogast T, Lorenzo SM, Li H, Tang SC, Richardson E, et al. Oligogenic Effects of 16p11.2 Copy-Number Variation on Craniofacial Development. Cell Rep. 2019;28(13):3320–8 e4. doi: 10.1016/j.celrep.2019.08.071 31553903

24. Hu Y, Flockhart I, Vinayagam A, Bergwitz C, Berger B, Perrimon N, et al. An integrative approach to ortholog prediction for disease-focused and other functional studies. BMC Bioinformatics. 2011;12:357. doi: 10.1186/1471-2105-12-357 21880147

25. Brand AH, Perrimon N. Targeted gene expression as a means of altering cell fates and generating dominant phenotypes. Development. 1993;118(2):401–15. 8223268

26. Strigini M, Cohen SM. A Hedgehog activity gradient contributes to AP axial patterning of the Drosophila wing. Development. 1997;124(22):4697–705. 9409685

27. Yan SJ, Gu Y, Li WX, Fleming RJ. Multiple signaling pathways and a selector protein sequentially regulate Drosophila wing development. Development. 2004;131(2):285–98. doi: 10.1242/dev.00934 14701680

28. Yusuff T, Jensen M, Yennawar S, Pizzo L, Karthikeyan S, Gould DJ, et al. Drosophila models of pathogenic copy-number variant genes show global and non-neuronal defects during development. PLoS Genet. 2020;16(6):e1008792. doi: 10.1371/journal.pgen.1008792 32579612

29. Callan MA, Cabernard C, Heck J, Luois S, Doe CQ, Zarnescu DC. Fragile X protein controls neural stem cell proliferation in the Drosophila brain. Hum Mol Genet. 2010;19(15):3068–79. doi: 10.1093/hmg/ddq213 20504994

30. Kwon CH, Luikart BW, Powell CM, Zhou J, Matheny SA, Zhang W, et al. Pten regulates neuronal arborization and social interaction in mice. Neuron. 2006;50(3):377–88. doi: 10.1016/j.neuron.2006.03.023 16675393

31. Lee A, Li W, Xu K, Bogert BA, Su K, Gao FB. Control of dendritic development by the Drosophila fragile X-related gene involves the small GTPase Rac1. Development. 2003;130(22):5543–52. doi: 10.1242/dev.00792 14530299

32. Parker L, Padilla M, Du Y, Dong K, Tanouye MA. Drosophila as a model for epilepsy: bss is a gain-of-function mutation in the para sodium channel gene that leads to seizures. Genetics. 2011;187(2):523–34. doi: 10.1534/genetics.110.123299 21115970

33. Rujano MA, Sanchez-Pulido L, Pennetier C, le Dez G, Basto R. The microcephaly protein Asp regulates neuroepithelium morphogenesis by controlling the spatial distribution of myosin II. Nat Cell Biol. 2013;15(11):1294–306. doi: 10.1038/ncb2858 24142104

34. Stuss DP, Boyd JD, Levin DB, Delaney KR. MeCP2 mutation results in compartment-specific reductions in dendritic branching and spine density in layer 5 motor cortical neurons of YFP-H mice. PLoS One. 2012;7(3):e31896. doi: 10.1371/journal.pone.0031896 22412847

35. Wang HD, Kazemi-Esfarjani P, Benzer S. Multiple-stress analysis for isolation of Drosophila longevity genes. Proc Natl Acad Sci U S A. 2004;101(34):12610–5. doi: 10.1073/pnas.0404648101 15308776

36. Orr WC, Sohal RS. Extension of life-span by overexpression of superoxide dismutase and catalase in Drosophila melanogaster. Science. 1994;263(5150):1128–30. doi: 10.1126/science.8108730 8108730

37. Copeland JM, Cho J, Lo T Jr., Hur JH, Bahadorani S, Arabyan T, et al. Extension of Drosophila life span by RNAi of the mitochondrial respiratory chain. Curr Biol. 2009;19(19):1591–8. doi: 10.1016/j.cub.2009.08.016 19747824

38. Mahmoudi S, Xu L, Brunet A. Turning back time with emerging rejuvenation strategies. Nat Cell Biol. 2019;21(1):32–43. doi: 10.1038/s41556-018-0206-0 30602763

39. Matthews BJ, Kim ME, Flanagan JJ, Hattori D, Clemens JC, Zipursky SL, et al. Dendrite self-avoidance is controlled by Dscam. Cell. 2007;129(3):593–604. doi: 10.1016/j.cell.2007.04.013 17482551

40. Soba P, Zhu S, Emoto K, Younger S, Yang SJ, Yu HH, et al. Drosophila sensory neurons require Dscam for dendritic self-avoidance and proper dendritic field organization. Neuron. 2007;54(3):403–16. doi: 10.1016/j.neuron.2007.03.029 17481394

41. King KL, Cidlowski JA. Cell cycle and apoptosis: common pathways to life and death. J Cell Biochem. 1995;58(2):175–80. doi: 10.1002/jcb.240580206 7673325

42. King KL, Cidlowski JA. Cell cycle regulation and apoptosis. Annu Rev Physiol. 1998;60:601–17. doi: 10.1146/annurev.physiol.60.1.601 9558478

43. Hunt P, Gulisano M, Cook M, Sham MH, Faiella A, Wilkinson D, et al. A distinct Hox code for the branchial region of the vertebrate head. Nature. 1991;353(6347):861–4. doi: 10.1038/353861a0 1682814

44. Hunt P, Whiting J, Muchamore I, Marshall H, Krumlauf R. Homeobox genes and models for patterning the hindbrain and branchial arches. Dev Suppl. 1991;1:187–96. 1683802

45. Lasser M, Pratt B, Monahan C, Kim SW, Lowery LA. The Many Faces of Xenopus: Xenopus laevis as a Model System to Study Wolf-Hirschhorn Syndrome. Front Physiol. 2019;10:817. doi: 10.3389/fphys.2019.00817 31297068

46. Le Lievre CS, Le Douarin NM. Mesenchymal derivatives of the neural crest: analysis of chimaeric quail and chick embryos. J Embryol Exp Morphol. 1975;34(1):125–54. 1185098

47. Lumsden A, Sprawson N, Graham A. Segmental origin and migration of neural crest cells in the hindbrain region of the chick embryo. Development. 1991;113(4):1281–91. 1811942

48. Mills A, Bearce E, Cella R, Kim SW, Selig M, Lee S, et al. Wolf-Hirschhorn Syndrome-Associated Genes Are Enriched in Motile Neural Crest Cells and Affect Craniofacial Development in Xenopus laevis. Front Physiol. 2019;10:431. doi: 10.3389/fphys.2019.00431 31031646

49. Goldberg JL. How does an axon grow? Genes Dev. 2003;17(8):941–58. doi: 10.1101/gad.1062303 12704078

50. Jensen M, Girirajan S. An interaction-based model for neuropsychiatric features of copy-number variants. PLoS Genet. 2019;15(1):e1007879. doi: 10.1371/journal.pgen.1007879 30653500

51. Thaker HM, Kankel DR. Mosaic analysis gives an estimate of the extent of genomic involvement in the development of the visual system in Drosophila melanogaster. Genetics. 1992;131(4):883–94. 1516819

52. Branco J, Al-Ramahi I, Ukani L, Perez AM, Fernandez-Funez P, Rincon-Limas D, et al. Comparative analysis of genetic modifiers in Drosophila points to common and distinct mechanisms of pathogenesis among polyglutamine diseases. Hum Mol Genet. 2008;17(3):376–90. doi: 10.1093/hmg/ddm315 17984172

53. Cziko AM, McCann CT, Howlett IC, Barbee SA, Duncan RP, Luedemann R, et al. Genetic modifiers of dFMR1 encode RNA granule components in Drosophila. Genetics. 2009;182(4):1051–60. doi: 10.1534/genetics.109.103234 19487564

54. Iyer J, Wang Q, Le T, Pizzo L, Gronke S, Ambegaokar SS, et al. Quantitative Assessment of Eye Phenotypes for Functional Genetic Studies Using Drosophila melanogaster. G3 (Bethesda). 2016;6(5):1427–37. doi: 10.1534/g3.116.027060 26994292

55. Baryshnikova A, Costanzo M, Kim Y, Ding H, Koh J, Toufighi K, et al. Quantitative analysis of fitness and genetic interactions in yeast on a genome scale. Nat Methods. 2010;7(12):1017–24. doi: 10.1038/nmeth.1534 21076421

56. Dixon SJ, Costanzo M, Baryshnikova A, Andrews B, Boone C. Systematic mapping of genetic interaction networks. Annu Rev Genet. 2009;43:601–25. doi: 10.1146/annurev.genet.39.073003.114751 19712041

57. Horn T, Sandmann T, Fischer B, Axelsson E, Huber W, Boutros M. Mapping of signaling networks through synthetic genetic interaction analysis by RNAi. Nat Methods. 2011;8(4):341–6. doi: 10.1038/nmeth.1581 21378980

58. Jonikas MC, Collins SR, Denic V, Oh E, Quan EM, Schmid V, et al. Comprehensive characterization of genes required for protein folding in the endoplasmic reticulum. Science. 2009;323(5922):1693–7. doi: 10.1126/science.1167983 19325107

59. Duncan AM, Ozawa T, Suzuki H, Rozen R. Assignment of the gene for the core protein II (UQCRC2) subunit of the mitochondrial cytochrome bc1 complex to human chromosome 16p12. Genomics. 1993;18(2):455–6. doi: 10.1006/geno.1993.1500 8288258

60. Gomez-Roman N, Grandori C, Eisenman RN, White RJ. Direct activation of RNA polymerase III transcription by c-Myc. Nature. 2003;421(6920):290–4. doi: 10.1038/nature01327 12529648

61. O’Donovan KJ, Diedler J, Couture GC, Fak JJ, Darnell RB. The onconeural antigen cdr2 is a novel APC/C target that acts in mitosis to regulate c-myc target genes in mammalian tumor cells. PLoS One. 2010;5(4):e10045. doi: 10.1371/journal.pone.0010045 20383333

62. Hartman JLt, Garvik B, Hartwell L. Principles for the buffering of genetic variation. Science. 2001;291(5506):1001–4. doi: 10.1126/science.291.5506.1001 11232561

63. Queitsch C, Carlson KD, Girirajan S. Lessons from model organisms: phenotypic robustness and missing heritability in complex disease. PLoS Genet. 2012;8(11):e1003041. doi: 10.1371/journal.pgen.1003041 23166511

64. Grozeva D, Carss K, Spasic-Boskovic O, Parker MJ, Archer H, Firth HV, et al. De novo loss-of-function mutations in SETD5, encoding a methyltransferase in a 3p25 microdeletion syndrome critical region, cause intellectual disability. Am J Hum Genet. 2014;94(4):618–24. doi: 10.1016/j.ajhg.2014.03.006 24680889

65. Hu P, Wu S, Sun Y, Yuan CC, Kobayashi R, Myers MP, et al. Characterization of human RNA polymerase III identifies orthologues for Saccharomyces cerevisiae RNA polymerase III subunits. Mol Cell Biol. 2002;22(22):8044–55. doi: 10.1128/mcb.22.22.8044-8055.2002 12391170

66. Pusapati GV, Kong JH, Patel BB, Krishnan A, Sagner A, Kinnebrew M, et al. CRISPR Screens Uncover Genes that Regulate Target Cell Sensitivity to the Morphogen Sonic Hedgehog. Dev Cell. 2018;44(2):271. doi: 10.1016/j.devcel.2018.01.002 29401421

67. McCamphill PK, Farah CA, Anadolu MN, Hoque S, Sossin WS. Bidirectional regulation of eEF2 phosphorylation controls synaptic plasticity by decoding neuronal activity patterns. J Neurosci. 2015;35(10):4403–17. doi: 10.1523/JNEUROSCI.2376-14.2015 25762683

68. Gildish I, Manor D, David O, Sharma V, Williams D, Agarwala U, et al. Impaired associative taste learning and abnormal brain activation in kinase-defective eEF2K mice. Learn Mem. 2012;19(3):116–25. doi: 10.1101/lm.023937.111 22366775

69. Zhang P, Riazy M, Gold M, Tsai SH, McNagny K, Proud C, et al. Impairing eukaryotic elongation factor 2 kinase activity decreases atherosclerotic plaque formation. Can J Cardiol. 2014;30(12):1684–8. doi: 10.1016/j.cjca.2014.09.019 25475470

70. Monteggia LM, Gideons E, Kavalali ET. The role of eukaryotic elongation factor 2 kinase in rapid antidepressant action of ketamine. Biol Psychiatry. 2013;73(12):1199–203. doi: 10.1016/j.biopsych.2012.09.006 23062356

71. Greene CS, Krishnan A, Wong AK, Ricciotti E, Zelaya RA, Himmelstein DS, et al. Understanding multicellular function and disease with human tissue-specific networks. Nat Genet. 2015;47(6):569–76. doi: 10.1038/ng.3259 25915600

72. Krishnan A, Zhang R, Yao V, Theesfeld CL, Wong AK, Tadych A, et al. Genome-wide prediction and functional characterization of the genetic basis of autism spectrum disorder. Nat Neurosci. 2016;19(11):1454–62. doi: 10.1038/nn.4353 27479844

73. Osipovich AB, Gangula R, Vianna PG, Magnuson MA. Setd5 is essential for mammalian development and the co-transcriptional regulation of histone acetylation. Development. 2016;143(24):4595–607. doi: 10.1242/dev.141465 27864380

74. D’Angelo D, Lebon S, Chen Q, Martin-Brevet S, Snyder LG, Hippolyte L, et al. Defining the Effect of the 16p11.2 Duplication on Cognition, Behavior, and Medical Comorbidities. JAMA Psychiatry. 2016;73(1):20–30. doi: 10.1001/jamapsychiatry.2015.2123 26629640

75. Niarchou M, Chawner S, Doherty JL, Maillard AM, Jacquemont S, Chung WK, et al. Psychiatric disorders in children with 16p11.2 deletion and duplication. Transl Psychiatry. 2019;9(1):8. doi: 10.1038/s41398-018-0339-8 30664628

76. Andrews T, Honti F, Pfundt R, de Leeuw N, Hehir-Kwa J, Vulto-van Silfhout A, et al. The clustering of functionally related genes contributes to CNV-mediated disease. Genome Res. 2015;25(6):802–13. doi: 10.1101/gr.184325.114 25887030

77. Iossifov I, Levy D, Allen J, Ye K, Ronemus M, Lee YH, et al. Low load for disruptive mutations in autism genes and their biased transmission. Proc Natl Acad Sci U S A. 2015;112(41):E5600–7. doi: 10.1073/pnas.1516376112 26401017

78. Kury S, van Woerden GM, Besnard T, Proietti Onori M, Latypova X, Towne MC, et al. De Novo Mutations in Protein Kinase Genes CAMK2A and CAMK2B Cause Intellectual Disability. Am J Hum Genet. 2017;101(5):768–88. doi: 10.1016/j.ajhg.2017.10.003 29100089

79. Krumm N, Turner TN, Baker C, Vives L, Mohajeri K, Witherspoon K, et al. Excess of rare, inherited truncating mutations in autism. Nature genetics. 2015;47(6):582–8. doi: 10.1038/ng.3303 25961944

80. Petrovski S, Wang Q, Heinzen EL, Allen AS, Goldstein DB. Genic intolerance to functional variation and the interpretation of personal genomes. PLoS Genet. 2013;9(8):e1003709. doi: 10.1371/journal.pgen.1003709 23990802

81. Yates AD, Achuthan P, Akanni W, Allen J, Allen J, Alvarez-Jarreta J, et al. Ensembl 2020. Nucleic Acids Res. 2020;48(D1):D682–D8. doi: 10.1093/nar/gkz966 31691826

82. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, et al. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 1997;25(17):3389–402. doi: 10.1093/nar/25.17.3389 9254694

83. Chintapalli VR, Wang J, Dow JA. Using FlyAtlas to identify better Drosophila melanogaster models of human disease. Nat Genet. 2007;39(6):715–20. doi: 10.1038/ng2049 17534367

84. Bowes JB, Snyder KA, Segerdell E, Jarabek CJ, Azam K, Zorn AM, et al. Xenbase: gene expression and improved integration. Nucleic Acids Res. 2010;38(Database issue):D607–12. doi: 10.1093/nar/gkp953 19884130

85. Xu D, Wang Y, Willecke R, Chen Z, Ding T, Bergmann A. The effector caspases drICE and dcp-1 have partially overlapping functions in the apoptotic pathway in Drosophila. Cell Death Differ. 2006;13(10):1697–706. doi: 10.1038/sj.cdd.4401920 16645642

86. Schmid A, Hallermann S, Kittel RJ, Khorramshahi O, Frolich AM, Quentin C, et al. Activity-dependent site-specific changes of glutamate receptor composition in vivo. Nat Neurosci. 2008;11(6):659–66. doi: 10.1038/nn.2122 18469810

87. Xiao C, Mileva-Seitz V, Seroude L, Robertson RM. Targeting HSP70 to motoneurons protects locomotor activity from hyperthermia in Drosophila. Dev Neurobiol. 2007;67(4):438–55. doi: 10.1002/dneu.20344 17443800

88. Ye J, Coulouris G, Zaretskaya I, Cutcutache I, Rozen S, Madden TL. Primer-BLAST: a tool to design target-specific primers for polymerase chain reaction. BMC Bioinformatics. 2012;13:134. doi: 10.1186/1471-2105-13-134 22708584

89. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25(4):402–8. doi: 10.1006/meth.2001.1262 11846609

90. Sun Y, Yolitz J, Wang C, Spangler E, Zhan M, Zou S. Aging studies in Drosophila melanogaster. Methods Mol Biol. 2013;1048:77–93. doi: 10.1007/978-1-62703-556-9_7 23929099

91. Ganetzky B, Wu CF. Indirect Suppression Involving Behavioral Mutants with Altered Nerve Excitability in DROSOPHILA MELANOGASTER. Genetics. 1982;100(4):597–614. 17246073

92. Schindelin J, Arganda-Carreras I, Frise E, Kaynig V, Longair M, Pietzsch T, et al. Fiji: an open-source platform for biological-image analysis. Nat Methods. 2012;9(7):676–82. doi: 10.1038/nmeth.2019 22743772

93. Bolger AM, Lohse M, Usadel B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014;30(15):2114–20. doi: 10.1093/bioinformatics/btu170 24695404

94. Kim D, Pertea G, Trapnell C, Pimentel H, Kelley R, Salzberg SL. TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions. Genome Biol. 2013;14(4):R36. doi: 10.1186/gb-2013-14-4-r36 23618408

95. Anders S, Pyl PT, Huber W. HTSeq—a Python framework to work with high-throughput sequencing data. Bioinformatics. 2015;31(2):166–9. doi: 10.1093/bioinformatics/btu638 25260700

96. Robinson MD, McCarthy DJ, Smyth GK. edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics. 2010;26(1):139–40. doi: 10.1093/bioinformatics/btp616 19910308

97. Thomas PD, Campbell MJ, Kejariwal A, Mi H, Karlak B, Daverman R, et al. PANTHER: a library of protein families and subfamilies indexed by function. Genome Res. 2003;13(9):2129–41. doi: 10.1101/gr.772403 12952881

98. Kircher M, Witten DM, Jain P, O’Roak BJ, Cooper GM, Shendure J. A general framework for estimating the relative pathogenicity of human genetic variants. Nat Genet. 2014;46(3):310–5. doi: 10.1038/ng.2892 24487276

99. Lek M, Karczewski KJ, Minikel EV, Samocha KE, Banks E, Fennell T, et al. Analysis of protein-coding genetic variation in 60,706 humans. Nature. 2016;536(7616):285–91. doi: 10.1038/nature19057 27535533

100. Elena SF, Lenski RE. Test of synergistic interactions among deleterious mutations in bacteria. Nature. 1997;390(6658):395–8. doi: 10.1038/37108 9389477

101. Lowery LA, Faris AE, Stout A, Van Vactor D. Neural Explant Cultures from Xenopus laevis. J Vis Exp. 2012(68):e4232. doi: 10.3791/4232 23295240

102. Nieuwkoop PD FJ. Normal table of Xenopus laevis (Daudin): a systematical and chronological survey of the development from the fertilized egg till the end of metamorphosis: Garland Pub., 1994; 1994.

103. Schneider CA, Rasband WS, Eliceiri KW. NIH Image to ImageJ: 25 years of image analysis. Nat Methods. 2012;9(7):671–5. doi: 10.1038/nmeth.2089 22930834

104. Kennedy AE, Dickinson AJ. Quantitative analysis of orofacial development and median clefts in Xenopus laevis. Anat Rec (Hoboken). 2014;297(5):834–55. doi: 10.1002/ar.22864 24443252

105. Popko J, Fernandes A, Brites D, Lanier LM. Automated analysis of NeuronJ tracing data. Cytometry A. 2009;75(4):371–6. doi: 10.1002/cyto.a.20660 18937344

106. Hagberg AA SD, Swart PJ, editor Exploring network structure, dynamics, and function using NetworkX. 7th Python in Science Conference SciPy 2008; 2008.

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