Understanding the relationship between genetic and phenotypic variation is one of the great outstanding challenges in biology. To meet this challenge, comprehensive genomic variation maps of human as well as of model organism populations are required. Here, we present a nucleotide resolution catalog of single-nucleotide, multi-nucleotide, and structural variants in 39 Drosophila melanogaster Genetic Reference Panel inbred lines. Using an integrative, local assembly-based approach for variant discovery, we identify more than 3.6 million distinct variants, among which were more than 800,000 unique insertions, deletions (indels), and complex variants (1 to 6,000 bp). While the SNP density is higher near other variants, we find that variants themselves are not mutagenic, nor are regions with high variant density particularly mutation-prone. Rather, our data suggest that the elevated SNP density around variants is mainly due to population-level processes. We also provide insights into the regulatory architecture of gene expression variation in adult flies by mapping cis-expression quantitative trait loci (cis-eQTLs) for more than 2,000 genes. Indels comprise around 10% of all cis-eQTLs and show larger effects than SNP cis-eQTLs. In addition, we identified two-fold more gene associations in males as compared to females and found that most cis-eQTLs are sex-specific, revealing a partial decoupling of the genomic architecture between the sexes as well as the importance of genetic factors in mediating sex-biased gene expression. Finally, we performed RNA-seq-based allelic expression imbalance analyses in the offspring of crosses between sequenced lines, which revealed that the majority of strong cis-eQTLs can be validated in heterozygous individuals.
References
[1]
Mackay TFC, Stone EA, Ayroles JF (2009) The genetics of quantitative traits: challenges and prospects. Nat Rev Genet 10: 565–577. doi: 10.1038/nrg2612
[2]
Consortium H (2010) Integrating common and rare genetic variation in diverse human populations. Nature 467: 52–58.
[3]
Mills RE, Walter K, Stewart C, Handsaker RE, Chen K, et al. (2011) Mapping copy number variation by population-scale genome sequencing. Nature 470: 59–65. doi: 10.1038/nature09708
[4]
Keane TM, Goodstadt L, Danecek P, White MA, Wong K, et al. (2011) Mouse genomic variation and its effect on phenotypes and gene regulation. Nature 477: 289–294. doi: 10.1038/nature10413
[5]
Gan X, Stegle O, Behr J, Steffen JG, Drewe P, et al. (2011) Multiple reference genomes and transcriptomes for Arabidopsis thaliana. Nature 477: 419–423. doi: 10.1038/nature10414
[6]
Ayroles JF, Carbone MA, Stone EA, Jordan KW, Lyman RF, et al. (2009) Systems genetics of complex traits in Drosophila melanogaster. Nat Genet 41: 299–307. doi: 10.1038/ng.332
[7]
Mackay TFC, Richards S, Stone EA, Barbadilla A, Ayroles JF, et al. (2012) The Drosophila melanogaster Genetic Reference Panel. Nature 482: 173–178. doi: 10.1038/nature10811
[8]
Rubin GM, Lewis EB (2000) A Brief History of Drosophila's Contributions to Genome Research. Science 287: 2216–2218. doi: 10.1126/science.287.5461.2216
[9]
Consortium Tm, Roy S, Ernst J, Kharchenko PV, Kheradpour P, et al. (2010) Identification of Functional Elements and Regulatory Circuits by Drosophila modENCODE. Science 330: 1787–1797.
[10]
Hens K, Feuz J-D, Isakova A, Iagovitina A, Massouras A, et al. (2011) Automated protein-DNA interaction screening of Drosophila regulatory elements. Nat Meth 8: 1065–1070. doi: 10.1038/nmeth.1763
[11]
Adams MD, Celniker SE, Holt RA, Evans CA, Gocayne JD, et al. (2000) The Genome Sequence of Drosophila melanogaster. Science 287: 2185–2195. doi: 10.1126/science.287.5461.2185
[12]
Moriyama EN, Powell JR (1996) Intraspecific nuclear DNA variation in Drosophila. Mol Biol Evol 13: 261–277. doi: 10.1093/oxfordjournals.molbev.a025563
[13]
Emerson JJ, Cardoso-Moreira M, Borevitz JO, Long M (2008) Natural Selection Shapes Genome-Wide Patterns of Copy-Number Polymorphism in Drosophila melanogaster. Science 320: 1629–1631. doi: 10.1126/science.1158078
[14]
Cridland JM, Thornton KR (2010) Validation of rearrangement break points identified by paired-end sequencing in natural populations of Drosophila melanogaster. Genome Biol Evol 2: 83–101. doi: 10.1093/gbe/evq001
[15]
Langley CH, Stevens K, Cardeno C, Lee YCG, Schrider DR, et al. (2012) Genomic Variation in Natural Populations of Drosophila melanogaster. Genetics doi: 10.1534/genetics.112.142018
[16]
Rebeiz M, Pool JE, Kassner VA, Aquadro CF, Carroll SB (2009) Stepwise Modification of a Modular Enhancer Underlies Adaptation in a Drosophila Population. Science 326: 1663–1667. doi: 10.1126/science.1178357
Carbone MA, Jordan KW, Lyman RF, Harbison ST, Leips J, et al. (2006) Phenotypic variation and natural selection at Catsup, a pleiotropic quantitative trait gene in Drosophila. Current Biology 16: 912–919. doi: 10.1016/j.cub.2006.03.051
[19]
Massouras A, Hens K, Gubelmann C, Uplekar S, Decouttere F, et al. (2010) Primer-initiated sequence synthesis to detect and assemble structural variants. Nat Meth 7: 485–486. doi: 10.1038/nmeth.f.308
[20]
Chen D, Ahlford A, Schnorrer F, Kalchhauser I, Fellner M, et al. (2008) High-resolution, high-throughput SNP mapping in Drosophila melanogaster. Nat Meth 5: 323–329. doi: 10.1038/nmeth.1191
[21]
Yalcin B, Wong K, Agam A, Goodson M, Keane TM, et al. (2011) Sequence-based characterization of structural variation in the mouse genome. Nature 477: 326–329. doi: 10.1038/nature10432
[22]
Mills RE, Pittard WS, Mullaney JM, Farooq U, Creasy TH, et al. (2011) Natural genetic variation caused by small insertions and deletions in the human genome. Genome Research 21: 830–839. doi: 10.1101/gr.115907.110
[23]
Cao J, Schneeberger K, Ossowski S, Gunther T, Bender S, et al. (2011) Whole-genome sequencing of multiple Arabidopsis thaliana populations. Nat Genet 43: 956–963. doi: 10.1038/ng.911
[24]
A map of human genome variation from population-scale sequencing. Nature 467: 1061–1073. doi: 10.1038/nature09991
[25]
Kim PM, Lam HYK, Urban AE, Korbel JO, Affourtit J, et al. (2008) Analysis of copy number variants and segmental duplications in the human genome: Evidence for a change in the process of formation in recent evolutionary history. Genome Research 18: 1865–1874. doi: 10.1101/gr.081422.108
[26]
Fiston-Lavier A-S, Anxolabehere D, Quesneville H (2007) A model of segmental duplication formation in Drosophila melanogaster. Genome Research 17: 000. doi: 10.1101/gr.6208307
[27]
Schrider Daniel R, Hourmozdi Jonathan N, Hahn Matthew W (2011) Pervasive Multinucleotide Mutational Events in Eukaryotes. Current Biology 21: 1051–1054. doi: 10.1016/j.cub.2011.05.013
[28]
Betancourt AJ, Kim Y, Orr HA (2004) A pseudohitchhiking model of X vs. autosomal diversity. Genetics 168: 2261–2269. doi: 10.1534/genetics.104.030999
[29]
Andolfatto P (2001) Contrasting patterns of X-linked and autosomal nucleotide variation in Drosophila melanogaster and Drosophila simulans. Mol Biol Evol 18: 279–290. doi: 10.1093/oxfordjournals.molbev.a003804
[30]
Begun DJ, Aquadro CF (1992) Levels of naturally occurring DNA polymorphism correlate with recombination rates in D. melanogaster. Nature 356: 519–520. doi: 10.1038/356519a0
[31]
Charlesworth B, Morgan MT, Charlesworth D (1993) The effect of deleterious mutations on neutral molecular variation. Genetics 134: 1289–1303.
[32]
Hodgkinson A, Eyre-Walker A (2011) Variation in the mutation rate across mammalian genomes. Nat Rev Genet 12: 756–766. doi: 10.1038/nrg3098
[33]
Tian D, Wang Q, Zhang P, Araki H, Yang S, et al. (2008) Single-nucleotide mutation rate increases close to insertions/deletions in eukaryotes. Nature 455: 105–108. doi: 10.1038/nature07175
[34]
McDonald MJ, Wang W-C, Huang H-D, Leu J-Y (2011) Clusters of Nucleotide Substitutions and Insertion/Deletion Mutations Are Associated with Repeat Sequences. PLoS Biol 9: e1000622 doi:10.1371/journal.pbio.1000622..
[35]
Clark AG, Eisen MB, Smith DR, Bergman CM, Oliver B, et al. (2007) Evolution of genes and genomes on the Drosophila phylogeny. Nature 450: 203–218.
[36]
Mavrich TN, Jiang C, Ioshikhes IP, Li X, Venters BJ, et al. (2008) Nucleosome organization in the Drosophila genome. Nature 453: 358–362. doi: 10.1038/nature06929
[37]
Peckham HE, Thurman RE, Fu Y, Stamatoyannopoulos JA, Noble WS, et al. (2007) Nucleosome positioning signals in genomic DNA. Genome Research 17: 1170–1177. doi: 10.1101/gr.6101007
[38]
Tolstorukov MY, Volfovsky N, Stephens RM, Park PJ (2011) Impact of chromatin structure on sequence variability in the human genome. Nat Struct Mol Biol 18: 510–515. doi: 10.1038/nsmb.2012
[39]
Tanay A, Siggia ED (2008) Sequence context affects the rate of short insertions and deletions in flies and primates. Genome Biol 9: R37. doi: 10.1186/gb-2008-9-2-r37
[40]
Stranger BE, Forrest MS, Dunning M, Ingle CE, Beazley C, et al. (2007) Relative Impact of Nucleotide and Copy Number Variation on Gene Expression Phenotypes. Science 315: 848–853. doi: 10.1126/science.1136678
[41]
Cheung VG, Nayak RR, Wang IX, Elwyn S, Cousins SM, et al. (2010) Polymorphic Cis- and Trans-Regulation of Human Gene Expression. PLoS Biol 8: e1000480 doi:10.1371/journal.pbio.1000480..
[42]
Parisi M, Nuttall R, Naiman D, Bouffard G, Malley J, et al. (2003) Paucity of Genes on the Drosophila X Chromosome Showing Male-Biased Expression. Science 299: 697–700. doi: 10.1126/science.1079190
[43]
Bachtrog D, Toda NRT, Lockton S (2010) Dosage Compensation and Demasculinization of X Chromosomes in Drosophila. Current Biology 20: 1476–1481. doi: 10.1016/j.cub.2010.06.076
[44]
Zhang Y, Sturgill D, Parisi M, Kumar S, Oliver B (2007) Constraint and turnover in sex-biased gene expression in the genus Drosophila. Nature 450: 233–237. doi: 10.1038/nature06323
[45]
Chintapalli VR, Wang J, Dow JAT (2007) Using FlyAtlas to identify better Drosophila melanogaster models of human disease. Nature Genetics 39: 715–720. doi: 10.1038/ng2049
[46]
Graveley BR, Brooks AN, Carlson JW, Duff MO, Landolin JM, et al. (2011) The developmental transcriptome of Drosophila melanogaster. Nature 471: 473–479. doi: 10.1038/nature09715
[47]
Veyrieras J-B, Kudaravalli S, Kim SY, Dermitzakis ET, Gilad Y, et al. (2008) High-Resolution Mapping of Expression-QTLs Yields Insight into Human Gene Regulation. PLoS Genet 4: e1000214 doi:10.1371/journal.pgen.1000214..
[48]
Doss S, Schadt EE, Drake TA, Lusis AJ (2005) Cis-acting expression quantitative trait loci in mice. Genome Res 15: 681–691. doi: 10.1101/gr.3216905
[49]
Ronald J, Brem RB, Whittle J, Kruglyak L (2005) Local Regulatory Variation in Saccharomyces cerevisiae. PLoS Genet 1: e25 doi:10.1371/journal.pgen.0010025.
[50]
Stranger BE, Nica AC, Forrest MS, Dimas A, Bird CP, et al. (2007) Population genomics of human gene expression. Nat Genet 39: 1217–1224. doi: 10.1038/ng2142
[51]
Mikkelsen T, Ku M, Jaffe D, Issac B, Lieberman E, et al. (2007) Genome-wide maps of chromatin state in pluripotent and lineage-committed cells. Nature 448: 553–560. doi: 10.1038/nature06008
[52]
McManus CJ, Coolon JD, Duff MO, Eipper-Mains J, Graveley BR, et al. (2010) Regulatory divergence in Drosophila revealed by mRNA-seq. Genome Res 20: 816–825. doi: 10.1101/gr.102491.109
[53]
Wittkopp P, Haerum B, Clark A (2004) Evolutionary changes in cis and trans gene regulation. Nature 430: 85–88. doi: 10.1038/nature02698
[54]
Pfaff CL, Parra EJ, Bonilla C, Hiester K, McKeigue PM, et al. (2001) Population structure in admixed populations: effect of admixture dynamics on the pattern of linkage disequilibrium. Am J Hum Genet 68: 198–207. doi: 10.1086/316935
[55]
Chippindale AK, Gibson JR, Rice WR (2001) Negative genetic correlation for adult fitness between sexes reveals ontogenetic conflict in Drosophila. Proceedings of the National Academy of Sciences 98: 1671–1675. doi: 10.1073/pnas.98.4.1671
[56]
Hoekstra HE, Coyne JA (2007) The locus of evolution: evo devo and the genetics of adaptation. Evolution 61: 995–1016. doi: 10.1111/j.1558-5646.2007.00105.x
[57]
Carroll SB (2008) Evo-Devo and an Expanding Evolutionary Synthesis: A Genetic Theory of Morphological Evolution. Cell 134: 25–36. doi: 10.1016/j.cell.2008.06.030
[58]
Li H, Durbin R (2009) Fast and accurate short read alignment with Burrows–Wheeler transform. Bioinformatics 25: 1754–1760. doi: 10.1093/bioinformatics/btp324
[59]
Fiston-Lavier A-S, Singh ND, Lipatov M, Petrov DA (2010) Drosophila melanogaster recombination rate calculator. Gene 463: 18–20. doi: 10.1016/j.gene.2010.04.015
