Summary - We are interested mainly in the molecular and population genetics of species and racial differentiation. The three main areas of research given below all center around this theme. The first area consists of genomic approaches to genes of species differentiation. The second area concerns gene expression differences between species, focusing on the roles microRNAs play in expression divergence. The third area of research activities is about interpreting genomic data by developing theories of natural selection and speciation.
I. Molecular genetics of species differentiation - The main interest is in the genetic and molecular basis of species differences. (You can think of human vs. chimpanzee, if you wish. How many genetic differences delineate us from them?) We use sibling species of Drosophila to get to the answers (see Wu, C.-I and C.-T. Ting 2004 Genes and speciation. Nature Review Genetics 5: 114-122). Among the traits of special interest are hybrid incompatibility and sexual behavior divergence. These traits are what define species.
Current approaches
In the past, we took a gene-by-gene approach and have successfully cloned two speciation genes (see the list below, or Wu and Ting 2004). In the beginning of this new century, there is a need and an opportunity to take a system-wide approach in order to identify the majority of genes involved in a particular process of speciation. We shall study the divergence in mating behaviors between the Z (for Zimbabwe) and M (cosmopolitan) behavioral races of Drosophila melanogaster (Wu et al. 1995; Hollocher et al. 1997a, b; Ting et al. 2001). These two races are at the very nascent stage of species formation.
We wish to understand the genetic and transcriptional bases of phenotypic divergence at this early stage of speciation. As shown in the figure below, the studies will be at 3 different levels - genotype, transcriptome and phenotype. The scope will be genomic and the tools will include genotyping tiling array, expression microarrays, large-scale sequencing, behavioral QTL mapping and, finally, precise gene replacement.
Aim 1 – At the genotypic level, we will identify nucleotide sites that are strongly differentiated between the Z and M races. In collaboration with large sequencing facilities in and outside of USA, we plan to sequence multiple Z and M lines in their entirety. (For example, one run of Solexa sequencing, at more than 1/2 gigabases, should yield at least 4X coverage of the Drosophila genome.) These race-differentiating sites will anchor genetic mapping of transcription and phenotypic divergence. We will construct multiply recombined (MR) lines between the Z- and M-races for such mapping.
Aim 2 – At the transcriptional level, we shall study the expression profiles of the MR lines (in heads and reproductive organs of the two sexes) and correlate these profiles with their underlying genotypes. The regulation of transcription is important in linking phenotype and genotype.
Aim 3 – At the phenotypic level, we shall first attempt to find the transcriptional basis of the phenotypic divergence (behavioral isolation), using the MR lines. We shall also link phenotypic divergence to genotypic changes directly. Genotyping will be done by either whole-genome tiling array or by direct sequencing using the new massive sequencing technology. We expect to identify most candidate “speciation genes” between the Z- and M- races and will carry out precise gene replacement (Rong and Golic 2000; Greenberg et al. 2003; Sun et al. 2004) to confirm their effects on the mating behavior.
II. Evolution of microRNAs in relation to expression divergenece -
One of the most intriguing class of regulatory molecules are microRNAs which have attracted great interest only recently (see the picture below). These are small RNA molecules that guide the RISC enzyme to degrade the targeted transcripts. The molecular action of miRNAs is reminiscent of the genetic basis of species differentiation (multiple factors, each with weak effect).
Current approach
The goal is to understand the evolution of microRNAs (miRNAs), the coevolution between miRNAs and their targets, and the role miRNA-target interactions play in the expression divergence between closely related species of Drosophila. miRNAs are small regulatory RNAs which target mRNA transcripts to repress their expression.
Aim 1. The evolution of miRNA genes – Emergence of new miRNA genes and changes in either the sequences of mature miRs or their expression will be studied by extensive sequencing of small RNAs from 6 tissues in 5 species of Drosophila.
Aim 2. The effects of miRNAs on the evolution of gene expression (I. miRNAs) – First, 4 conservative miRNAs will be used to transform D. melanogaster and D. simulans. Second, 4 pairs of evolving miRNAs will be used to transform D. melanogaster. The effects of the transgenes on expression will be assayed by microarrays.
Aim 3. The effects of miRNAs on the evolution of gene expression (II. targets) – We will verify and calibrate the results of D2 by selecting the 3' UTR of 15 target genes for reporter assay.
Aim 4. The fitness effects of the coevolution between miRNAs and targets - If miRNAs and targets are co-adapted, then miRNAs and target transcripts that have been evolving separately would likely show fitness incompatibilities. This hypothesis, in the frame work of Muller-Dobzhansky model, will be tested.
III. Genomics and population genetics -The torrent of genomic data by DNA sequencing and expression microarrays have provided unprecedented opportunities for analyzing natural selection and adaptation. Speciation is the consequence of natural selection driving populations to adapt to different environments. It is also a genome-scale phenomenon.
Current approaches -We continue to carry out data analysis as done in the papers cited below. The data were often obtained from the public domain and we have been developing theoretical tools to analyze these data. At the same time, we also carry out large-scale data collection in collaboration with various sequencing centers. We also continue to develop theories to test the geographical models of speciation (see Osada and Wu 2005; Zhou et al. 2007).
Ting, C.-T., S.-C. Tsaur, M.-L. Wu and C.-I Wu, 1998 A rapidly evolving homeobox at the site of a hybrid sterility gene. Science 282: 1501-1504.
Fay, J. C. and C.-I Wu, 1999 A human population bottleneck is not incompatible with the discordance between patterns of mitochondrial vs. nuclear DNA variation. Molec. Biol. Evol. 16:1003 - 1006.
Wyckoff, G. J., W. Wang and C.-I Wu, 2000 Rapid evolution of male reproductive genes in the descent of man. Nature 403: 304 - 309.
Sawamura, K., A. W. Davis and C.-I Wu, 2000 Genetic analysis by means of introgression into Drosophila melanogaster. Proc. Natl. Acad. Sci. 97:2652-2655.
Ting, T.-C., S. C. Tsaur, S.-C., and C.-I Wu, 2000 The phylogeny of closely related species as revealed by the genealogy of a speciation gene, Odysseus. Proc. Natl. Acad. Sci. 97:5313 - 5316.
Fay, J. C. and C.-I Wu, 2000 Hitchhiking under positive Darwinian selection. Genetics 155: 1405-1413.
Wu, C.-I, 2000 Genetics of species differentiation: What is unknown and what will be unknowable? Evol. Biol. 32: 239 - 248. (M. T. Clegg, M. K. Hecht and R. J. MacIntyre, eds.)
Tsaur, S. C., C.-T. Ting and C.-I Wu, 2001 Sex in Drosophila mauritiana: Extremely high level of replacement polymorphism in a male reproductive gene. Molec. Biol. Evol. 18:22-26
Alipaz, J. A., C-I Wu and T. L. Karr, 2001 Sperm-egg incompatibility between races of Drosophila. Proc. Roy. Soc. 268:789-795.
Takahashi, A, S.C. Tsaur, J. A. Coyne and C.-I Wu, 2001 The nucleotide changes governing pheromonal variation in Drosophila and their evolution. Proc. Natl. Acad. Sci.98:3920-3925.
Fay, J. C., G. J. Wyckoff and C.-I Wu, 2001 Positive and negative selection on the human genome. Genetics 158:1227-1234.
Ting, C.T., A. Takahashi and C.-I Wu, 2001 Incipient speciation by sexual selection: concurrent evolution at multiple loci. Proc. Natl. Acad. Sci. 98: 6709-6713 (see also Nature 412:31-32)
Luo, Z.W. and C.-I Wu, 2001 Modeling linkage disequilibrium between a polymorphic marker locus and a locus affecting dichotomous disease traits in natural populations. Genetics 158: 1785-1800.
Wu, C.-I 2001 The genic view of the process of speciation. J. Evol. Biol. 14: 851-865..
Wu, C.-I 2001 Genes and Speciation - A reply. J. Evol. Biol. 14:889-891.
Fay, J. C. and C.-I Wu, 2001 The neutral theory in the genomic era. Curr. Opinion Gen. Dev. 11:642-646.
Fay, J. C., G. J. Wyckoff and C.-I Wu, 2002 Testing the neutral theory of molecular evolution with genomic data. Nature 415: 1024 - 1026.
Luo, Z.W., C.-I Wu, and M. J. Kearsey, 2002 Precision and high-resolution mapping of quantitative trait loci by use of recurrent selection, backcross or intercross schemes. Genetics 161: 915-929.
Wyckoff, G. J., J. Li and C.-I Wu, 2002 Molecular evolution of functional genes on the mammalian Y chromosome. Molec. Biol. Evol. 19: 1633-1636
Fang, S., A. Takahashi, and C.-I Wu, 2002 A mutation in the promoter of desaturase 2 is correlated with sexual isolation between Drosophila behavioral races. Genetics 168: 781-784.
Wu, C.-I and E. Y. Xu, 2003 Sexual antagonism and X-inactivation - The SAXI hypothesis. Trends in Genetics 19:243-247.
Fay, J. C., and C.-I. Wu, 2003a Sequence divergence, selective constraint and selection in protein evolution. Ann. Rev. Genomics Hum. Genet. 4:213-235.
Fay, J. C., and C.-I. Wu, 2003b Detecting hitchhiking from patterns of DNA polymorphisms in Selective sweeps, edited by D. I. Nurminsky. Landes Bioscience.
Wang, H-Y., H. Tang, C. K. Shen and C.-I Wu 2003 Rapidly evolving genes in human – I. The glycophorins and their possible role in evading malaria parasites. Mol. Biol. Evol. 20:1795-1804.
Greenberg, A. J., J. R. Moran, J. A. Coyne, and Chung-I Wu 2003 Ecological adaptation during incipient speciation as revealed by precise gene replacement. Science 302: 1754-1757.
Lu, J., W.-H. Li and C.-I Wu. 2003 Chromosomal speciation and gene flow. Science 302:988c.
Wu, C.-I, S. Shi and Y. Zhang 2004 A case for conservation. Nature 428:213-214. (A commentary in a supplementary issue).
Wu, C.-I and C.-T. Ting 2004 Genes and speciation. Nature Review Genetics 5: 114122.
He, J et al. (51 authors) 2004. Molecuar evolution of the SARS-conronavirus during the course of SARS-epidemic in China. Science 303: 1666-1669 (C.-I Wu is a communicating author for the data analysis section).
Kohn, M. H., S. Fang and C.-I Wu 2004 Inference of positive and negative selection on the 5' regulatory regions of Drosophila genes. Mol. Biol. Evol. 21:374-383.
Kyoichi Sawamura, John Roote, Chung-I Wu, and Masa-Toshi Yamamoto 2004 Extreme Genetic Complexity Underlying Hybrid Male Sterility in Drosophila. Genetics 166: 789-796.
Tang, H. , G. J. Wyckoff, J Lu, and C.-I Wu, 2004 A Universal Evolutionary Index for Amino Acid Changes. Mol. Biol. Evol. 21: 1548-1556.
Ting, C., S. Tsuar, S. Sun, W. Browne, N. Patel and C.-I Wu 2004 Gene duplication and speciation in Drosophila – Evidence from the Odysseus locus. Proc. Natl. Acad. Sci. 101: 12232-12235.
Sun, S, C. Ting, and C.-I Wu 2004 The normal function of a speciation gene, Odysseus, and its hybrid sterility effect. Science 305: 81-83.
Osada, N. and C.-I Wu 2005 Testing the mode of speciation with genomic data - Examples from the great apes. Genetics 169: 259-264.
Song, H.-D. et al. C.-I Wu and G.-P. Zhao 2005 Cross-host evolution of SARS coronavirus in palm civet and human. Proc. Natl. Acad. Sci. 102: 2430-2435
Alipaz, J. A., S. Fang and C.-I Wu 2005 Evolution of sexual isolation in laboratory populations: I. Genotypic vs. phenotypic changes during secondary contact. Amer. Natur. 165:420-428.
Alipaz, J. A., T. Karr and C.-I Wu 2005 Evolution of sexual isolation in laboratory populations: II. Fitness effects of mating traits and the associated hybrid incompatibilities Amer. Natur. 165:429-438.
Lu, Jian and C.-I Wu 2005 Weak selection revealed by the whole-genome comparison between the X and autosomes of human and chimpanzee. Proc. Natl. Acad. Sci. 102: 4063-4067.
Zhou, R., S. Shi and C-I Wu 2005 Molecular criteria for determining new hybrid species - an application to the Sonneratia hybrids. Molec. Phy. Evol. 35:595-601.
Osada, N. et al. C.-I Wu and K. Hashimoto. 2005. Substitution rate and structural divergence of 5’UTR evolution: Comparative analysis between human and cynomolgus monkey cDNAs. Molec. Biol. Evol. 22:1976-1982
Tang, H and C.-I Wu. 2006. A new method for estimating nonsynonymous substitutions and its applications to detecting positive selection. Molec. Biol. Evol. 23:372-379.
Greenberg, A. J., J. R. Moran, S. Fang and C.-I Wu. 2006. Adaptive loss of an old duplicate gene during incipient speciation. Mol. Biol. Evol., 23:401--410.
Greenberg, A. J., J. R. Moran and C.-I Wu 2006. Proper control of genetic background with precise allele substitution: A comment on Coyne and Elwyn. Evolution 60:623-625.
Greenberg, A. J. and C.-I Wu 2006. Molecular genetics of natural populations. Mol. Biol. Evol. 23:883-886.
Lu, J., T. Tang, J. Huang, S. Shi, and C.-I Wu. 2006. The accumulation of deleterious mutations in the rice genomes: a hypothesis on the cost of domestication. Trends in Genetics 22:126 - 131.
Liu, X., Y. Fu, (9 other authors), C.-I Wu, and A. Xu. 2006. An ancient balanced polymorphism in a regulatory region of human Major Histocompatibility Comlex is retained in Chinese minorities but lost worldwide. Amer. J. Hum. Genet.78: 393 - 400.
Osada, N., M. H. Kohn, and C.-I Wu. 2006.Genomic Inference of cis-regulatory nucleotide polymorphism underlying gene expression differences between Drosophila melanogaster mating races.Molec. Biol. Evol. 23(8):1585-1591.
Zeng, Kai, S.H.,Shi, Y.X. Fu, and C.-I Wu. 2006. Statistical Tests for Detecting Positive Selection by Utilizing High Frequency SNPs.Genetics 174:1431-1439.
Tang, T., J. Lu, J. Huang, J. He, Y. Shen, K. Zeng, S. R. McCouch, M. D. Purugganan, S. Shi and C.-I Wu. 2006. Genomic variation in rice - Genesis of highly polymorphic linkdage blocks during domestication. PlosGenetics 2:1824-1833.
Wang, Huan-Chieh Chien, Naoki Osada, Katsuyuki Hashimoto, Sumio Sugano, Takashi Gojobori, Chen-Kung Chou, Shih-Feng Tsai, Chung-I Wu, C.-K. James Shen. 2007. Rate of evolution in brain-expressed genes in humans. PlosBiology 5:335-342.
Shapiro, J. A. et al. C.-I Wu. 2007. Adaptive Genic evolution in the Drosophila genomes. Proc. Natl. Acad. Sci. 104:2271-2276.
Gojobori, J., H. Tang, J. Akey and C.-I Wu 2007. Negative correlation between the two phases of molecular evolution - constraint and adaptation in the human genomes. Proc. Natl. Acad. Sci. 104:3907-3912.
Zeng, K., S. Mano, S. Shi and C.-I Wu 2007. Comparisons of site- and Haplotype-Frequency methods for detecting positive selection. Molec. Biol. Evol. 24(7):1562-1574.
Zeng K., S. Shi and C.-I Wu 2007. Compound tests for the detection of hitchhiking under positive selection. Molec. Biol. Evol. 24(8):1898-1908.
Zhou, R, K. Zeng, W. Wu, X. Chen, Z. Yang, S. Shi and C.-I Wu. Population genetics of speciation in non-model organisms: I. Ancestral polymorphism in mangroves. Molec. Biol. Evol. (in press)
Zhou, R., X. Gong, D. Boufford, C.-I Wu and S. Shi Testing the Hypothesis on Unidirectional Hybridization in Plants: Observations on Sonneratia, Bruguiera and Ligularia. BMC Evolutionary Biology (in press)
Lu, J., Y. Fu, S. Kumar, Y. Shen, R. W. Carthew and C.-I Wu. Adaptive evolution of newly-ermged microRNA genes in Drosophila. Molec. Biol. Evol.(in review)
Lu, J.,Y. Shen, Q. Wu, S. Kumar,B. He, R. W. Carthew, S. Wang and C.-I Wu. The birth and death of microRNA genes in Drosophila. Nature Genetics(in revision)
Wang, H., Y Fu, M. McPeek, X. Lu, S. Nuzhdin, A. Xu, M. Wu and C.-I Wu. Genetic architecture of expression differences between Drosophila races - Analysis of chromosome substitutions. Proc. Natl. Acad. Sci. (submitted)