Browsing by Author "Putnam, Nicholas H."
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Item Comparative Genomics of Cephalochordates(2015-04-23) Yue, Jiaxing; Kohn, Michael H.; Nakhleh, Luay K.; Shamoo, Yousif; Guerra, Rudy; Putnam, Nicholas H.Cephalochordates, commonly known as lancelets or amphioxus, represent an ancient chordate lineage falling at the boundary between invertebrates and vertebrates. They are considered the best living proxy for the common ancestor of all chordate animals and hold the key for understanding chordate evolution. Despite such great importance, current studies on cephalochordates are generally limited to the Branchiostoma genus, leaving the other two genera, Asymmetron and Epigonichthys largely unexplored. In this dissertation, I set out to fill this gap by developing an array of genomic resources for the Bahama cephalochordate, Asymmetron lucayanum, by both RNA-Seq and whole-genome shotgun (WGS) sequencing. The transcriptome and genome of this representative cephalochordate species were assembled and characterized via the state-of-arts comparative genomics approach. By comparing its transcriptome and genome sequences with those of a distant related cephalochordate species, Branchiostoma floridae, as well as with several representative vertebrate species, many aspects of their genome biology were illuminated, which includes lineage-specific molecular evolution rate, fast-evolving genes, evolution time frame, conserved non-coding elements, and germline-related genes. The raw genomic resources, technical pipelines and biological results and insights generated by this dissertation work will benefit the whole cephalochordate research community by providing a powerful guide for formulating new hypotheses and designing new experiments towards a better understanding about the biology and evolution of cephalochordates, as well as the evolutionary transition from invertebrates to vertebrates.Item Conserved Noncoding Elements in the Most Distant Genera of Cephalochordates: The Goldilocks Principle(Oxford University Press, 2016) Yue, Jia-Xing; Kozmikova, Iryna; Ono, Hiroki; Nossa, Carlos W.; Kozmik, Zbynek; Putnam, Nicholas H.; Yu, Jr-Kai; Holland, Linda Z.Cephalochordates, the sister group of vertebrates + tunicates, are evolving particularly slowly. Therefore, genome comparisons between two congeners of Branchiostoma revealed so many conserved noncoding elements (CNEs), that it was not clear how many are functional regulatory elements. To more effectively identify CNEs with potential regulatory functions, we compared noncoding sequences of genomes of the most phylogenetically distant cephalochordate genera, Asymmetron andBranchiostoma, which diverged approximately 120–160 million years ago. We found 113,070 noncoding elements conserved between the two species, amounting to 3.3% of the genome. The genomic distribution, target gene ontology, and enriched motifs of these CNEs all suggest that many of them are probably cis-regulatory elements. More than 90% of previously verified amphioxus regulatory elements were re-captured in this study. A search of the cephalochordate CNEs around 50 developmental genes in several vertebrate genomes revealed eight CNEs conserved between cephalochordates and vertebrates, indicating sequence conservation over >500 million years of divergence. The function of five CNEs was tested in reporter assays in zebrafish, and one was also tested in amphioxus. All five CNEs proved to be tissue-specific enhancers. Taken together, these findings indicate that even though Branchiostoma andAsymmetron are distantly related, as they are evolving slowly, comparisons between them are likely optimal for identifying most of their tissue-specific cis-regulatory elements laying the foundation for functional characterizations and a better understanding of the evolution of developmental regulation in cephalochordates.Item Effects of Gene Interactions on Polymorphism and Divergence(2014-05-20) Shih, Ching-Hua; Kohn, Michael H.; Nakhleh, Luay K.; Putnam, Nicholas H.; Kimmel, MarekPatterns of interactions could influence the biological systems at various levels and potentially affect the evolutionary history. Gene interactions could affect the relation among genotypes and their phenotypes. Polymorphisms of genes potentially alter interactions among genes, and hence, affect the fitness of individuals. Certain combinations of polymorphisms among genes can be maintained by selection. The main question of this thesis regards the effects of interactions in biological systems. Reproductive isolation arises as a by-product of different combinations of substitutions between divergent populations. Bateson-Dobzhansky-Muller (BDM) model states fitness changes due to incompatible combinations of loci. Nonlinear rates of accumulation of incompatibilities have been proposed considering interactions among multiple loci. However, the effects of topologies of gene interaction networks (GINs) altering the rates of accumulation of incompatibilities have not been investigated. The third topic revolves around effects of gene interactions in hybridizing species. Gene flow homogenizes the gene pool of incipient species and impedes divergence. This process can take place because incipient species either remain in spatial contact or have secondary contact through range shifts. The porous intrinsic reproductive barriers between species for loci post various properties contributing to success to move between species. We utilized human GINs combined with single nucleotide polymorphisms (SNPs) from human HapMap to investigate the correlations between interactions and interlocus nonrandom associations of polymorphisms. To investigate the effects of gene interactions between species, we modified the “snowball effect” and simulated the rates of accumulation of incompatibilities by introducing the structure information of GINs. To profile the functional characteristics of introgressed genes, we adopted the maximum likelihood method for public genomic resources focusing on a primate hybrid zone of cynomolgus monkey (Macaca fascicularis) and rhesus monkey (M. mulatta). Our results suggest that GINs enable global scale studies and provide polygenic insight of complex traits between and within species. Application of gene interactions ranges from enhancement of genome-wide association studies, identification of interacting polymorphisms to biomedical researches. Gene interactions also provide a platform of understanding hybridization and the dynamics of speciation.Item Evolution of the ᅠPerlecan/HSPG2ᅠ Gene and Its Activation in Regeneratingᅠ Nematostella vectensis(Public Library of Science, 2015) Warren, Curtis R.; Kassir, Elias; Spurlin, James W.; Martinez, Jerahme R.; Putnam, Nicholas H.; Farach-Carson, Mary C.The heparan sulfate proteoglycan 2 (HSPG2)/perlecan gene is ancient and conserved in all triploblastic species. Its presence maintains critical cell boundaries in tissue and its large (up to ~900 kDa) modular structure has prompted speculation about the evolutionary origin of the gene. The gene’s conservation amongst basal metazoans is unclear. After the recent sequencing of their genomes, the cnidarian Nematostella vectensis and the placozoanTrichoplax adhaerens have become favorite models for studying tissue regeneration and the evolution of multicellularity. More ancient basal metazoan phyla include the poriferan and ctenophore, whose evolutionary relationship has been clarified recently. Our in silico and PCR-based methods indicate that the HSPG2 gene is conserved in both the placozoan and cnidarian genomes, but not in those of the ctenophores and only partly in poriferan genomes.HSPG2 also is absent from published ctenophore and Capsaspora owczarzaki genomes. The gene in T. adhaerens is encoded as two separate but genetically juxtaposed genes that house all of the constituent pieces of the mammalian HSPG2 gene in tandem. These genetic constituents are found in isolated genes of various poriferan species, indicating a possible intronic recombinatory mechanism for assembly of the HSPG2 gene. Perlecan’s expression during wound healing and boundary formation is conserved, as expression of the gene was activated during tissue regeneration and reformation of the basement membrane of N.vectensis. These data indicate that the complex HSPG2 gene evolved concurrently in a common ancestor of placozoans, cnidarians and bilaterians, likely along with the development of differentiated cell types separated by acellular matrices, and is activated to reestablish these tissue borders during wound healing.Item Evolutionary profiling reveals the heterogeneous origins of classes of human disease genes: implications for modeling disease genetics in animals(BioMed Central, 2014) Maxwell, Evan K.; Schnitzler, Christine E.; Havlak, Paul; Putnam, Nicholas H.; Nguyen, Anh-Dao; Moreland, R. TravisBackground: The recent expansion of whole-genome sequence data available from diverse animal lineages provides an opportunity to investigate the evolutionary origins of specific classes of human disease genes. Previous studies have observed that human disease genes are of particularly ancient origin. While this suggests that many animal species have the potential to serve as feasible models for research on genes responsible for human disease, it is unclear whether this pattern has meaningful implications and whether it prevails for every class of human disease. Results: We used a comparative genomics approach encompassing a broad phylogenetic range of animals with sequenced genomes to determine the evolutionary patterns exhibited by human genes associated with different classes of disease. Our results support previous claims that most human disease genes are of ancient origin but, more importantly, we also demonstrate that several specific disease classes have a significantly large proportion of genes that emerged relatively recently within the metazoans and/or vertebrates. An independent assessment of the synonymous to non-synonymous substitution rates of human disease genes found in mammals reveals that disease classes that arose more recently also display unexpected rates of purifying selection between their mammalian and human counterparts. Conclusions: Our results reveal the heterogeneity underlying the evolutionary origins of (and selective pressures on) different classes of human disease genes. For example, some disease gene classes appear to be of uncommonly recent (i.e., vertebrate-specific) origin and, as a whole, have been evolving at a faster rate within mammals than the majority of disease classes having more ancient origins. The novel patterns that we have identified may provide new insight into cases where studies using traditional animal models were unable to produce results that translated to humans. Conversely, we note that the larger set of disease classes do have ancient origins, suggesting that many non-traditional animal models have the potential to be useful for studying many human disease genes. Taken together, these findings emphasize why model organism selection should be done on a disease-by-disease basis, with evolutionary profiles in mind.Item The Genome of the Ctenophore Mnemiopsis leidyi and Its Implications for Cell Type Evolution(American Association for the Advancement of Science, 2013) Ryan, Joseph F.; Pang, Kevin; Schnitzler, Christine E.; Nguyen, Anh-Dao; Moreland, R. Travis; Simmons, David K.; Koch, Bernard J.; Francis, Warren R.; Havlak, Paul; NISC Comparative Sequencing Program; Smith, Stephen A.; Putnam, Nicholas H.; Haddock, Steven H.D.; Dunn, Casey W.; Wolfsberg, Tyra G.; Mullikin, James C.; Martindale, Mark Q.; Baxevanis, Andreas D.An understanding of ctenophore biology is critical for reconstructing events that occurred early in animal evolution. Toward this goal, we have sequenced, assembled, and annotated the genome of the ctenophore Mnemiopsis leidyi. Our phylogenomic analyses of both amino acid positions and gene content suggest that ctenophores rather than sponges are the sister lineage to all other animals. Mnemiopsis lacks many of the genes found in bilaterian mesodermal cell types, suggesting that these cell types evolved independently. The set of neural genes in Mnemiopsis is similar to that of sponges, indicating that sponges may have lost a nervous system. These results present a newly supported view of early animal evolution that accounts for major losses and/or gains of sophisticated cell types, including nerve and muscle cells.Item Hemichordate genomes and deuterostome origins(Macmillan Publishers Limited, 2015) Simakov, Oleg; Kawashima, Takeshi; Marlétaz, Ferdinand; Jenkins, Jerry; Koyanagi, Ryo; Mitros, Therese; Hisata, Kanako; Bredeson, Jessen; Shoguchi, Eiichi; Gyoja, Fuki; Yue, Jia-Xing; Chen, Yi-Chih; Freeman, Robert M.; Sasaki, Akane; Hikosaka-Katayama, Tomoe; Sato, Atsuko; Fujie, Manabu; Baughman, Kenneth W.; Levine, Judith; Gonzalez, Paul; Cameron, Christopher; Fritzenwanker, Jens H.; Pani, Ariel M.; Goto, Hiroki; Kanda, Miyuki; Arakaki, Nana; Yamasaki, Shinichi; Qu, Jiaxin; Cree, Andrew; Ding, Yan; Dinh, Huyen H.; Dugan, Shannon; Holder, Michael; Jhangiani, Shalini N.; Kovar, Christie L.; Lee, Sandra L.; Lewis, Lora R.; Morton, Donna; Nazareth, Lynne V.; Okwuonu, Geoffrey; Santibanez, Jireh; Chen, Rui; Richards, Stephen; Muzny, Donna M.; Gillis, Andrew; Peshkin, Leonid; Wu, Michael; Humphreys, Tom; Su, Yi-Hsien; Putnam, Nicholas H.; Schmutz, Jeremy; Fujiyama, Asao; Yu, Jr-Kai; Tagawa, Kunifumi; Worley, Kim C.; Gibbs, Richard A.; Kirschner, Marc W.; Lowe, Christopher J.; Satoh, Noriyuki; Rokhsar, Daniel S.; Gerhart, JohnAcorn worms, also known as enteropneust (literally, ‘gut-breathing’) hemichordates, are marine invertebrates that share features with echinoderms and chordates. Together, these three phyla comprise the deuterostomes. Here we report the draft genome sequenItem Joint assembly and genetic mapping of the Atlantic horseshoe crab genome reveals ancient whole genome duplication(BioMed Central, 2014) Nossa, Carlos W.; Havlak, Paul; Yue, Jia-Xing; Lv, Jie; Vincent, Kimberly Y.; Brockmann, H.Jane; Putnam, Nicholas H.Horseshoe crabs are marine arthropods with a fossil record extending back approximately 450 million years. They exhibit remarkable morphological stability over their long evolutionary history, retaining a number of ancestral arthropod traits, and are often cited as examples of “living fossils.” As arthropods, they belong to the Ecdysozoa, an ancient super-phylum whose sequenced genomes (including insects and nematodes) have thus far shown more divergence from the ancestral pattern of eumetazoan genome organization than cnidarians, deuterostomes and lophotrochozoans. However, much of ecdysozoan diversity remains unrepresented in comparative genomic analyses. Here we apply a new strategy of combined de novo assembly and genetic mapping to examine the chromosome-scale genome organization of the Atlantic horseshoe crab, Limulus polyphemus. We constructed a genetic linkage map of this 2.7 Gbp genome by sequencing the nuclear DNA of 34 wild-collected, full-sibling embryos and their parents at a mean redundancy of 1.1x per sample. The map includes 84,307 sequence markers grouped into 1,876 distinct genetic intervals and 5,775 candidate conserved protein coding genes. Comparison with other metazoan genomes shows that the L. polyphemus genome preserves ancestral bilaterian linkage groups, and that a common ancestor of modern horseshoe crabs underwent one or more ancient whole genome duplications 300 million years ago, followed by extensive chromosome fusion. These results provide a counter-example to the often noted correlation between whole genome duplication and evolutionary radiations. The new, low-cost genetic mapping method for obtaining a chromosome-scale view of non-model organism genomes that we demonstrate here does not require laboratory culture, and is potentially applicable to a broad range of other species.Item Low coverage sequencing of three echinoderm genomes: the brittle star Ophionereis fasciata, the sea star Patiriella regularis, and the sea cucumber Australostichopus mollis(BioMed Central, 2016) Long, Kyle A.; Nossa, Carlos W.; Sewell, Mary A.; Putnam, Nicholas H.; Ryan, Joseph F.BACKGROUND: There are five major extant groups of Echinodermata: Crinoidea (feather stars and sea lillies), Ophiuroidea (brittle stars and basket stars), Asteroidea (sea stars), Echinoidea (sea urchins, sea biscuits, and sand dollars), and Holothuroidea (sea cucumbers). These animals are known for their pentaradial symmetry as adults, unique water vascular system, mutable collagenous tissues, and endoskeletons of high magnesium calcite. To our knowledge, the only echinoderm species with a genome sequence available to date is Strongylocentrotus pupuratus (Echinoidea). The availability of additional echinoderm genome sequences is crucial for understanding the biology of these animals. FINDINGS: Here we present assembled draft genomes of the brittle star Ophionereis fasciata, the sea star Patiriella regularis, and the sea cucumber Australostichopus mollis from Illumina sequence data with coverages of 12.5x, 22.5x, and 21.4x, respectively. CONCLUSIONS: These data provide a resource for mining gene superfamilies, identifying non-coding RNAs, confirming gene losses, and designing experimental constructs. They will be important comparative resources for future genomic studies in echinoderms.Item Network-guided genome-wide studies reveal a complex genetic architecture of warfarin resistance in the Norway rat (Rattus norvegicus)(2013-09-16) Li, Shuwei; Kohn, Michael H.; Putnam, Nicholas H.; Nakhleh, Luay K.; Kimmel, Marek; Peng, BoA fundamental challenge in evolutionary biology and medical genetic research is to connect the phenotype (a disease in humans or an adaptive trait in animals or plants) with the genotype. Using a classical example of an adaptive trait with a strong Mendelian genetic basis - warfarin resistance in the Norway rat (Rattus norvegicus), my dissertation tests the main hypothesis that speculated ‘simple’ adaptive trait has a more complex genetic architecture. Warfarin is an anticoagulant rodenticide used since the 1950s, and also is a widely prescribed blood-thinning drug in human. As a rodenticide, warfarin has initially been very effective. However, resistant rodents have evolved quickly and Vkorc1 (vitamin K epoxide reductase complex subunit 1) is the known resistance gene. As a popular drug, warfarin has a narrow therapeutic window with several genes VKORC1, CYP2C9, CYP4F2 established as biomarkers predicting warfarin dose in humans, suggesting a complex genetic architecture of warfarin resistance in rodents. In my thesis I performed network-guided genomic association studies (NetGWAS) and gene expression analysis to identify candidate genes involved in warfarin resistance based on a sample of ~600 wild rats from 19 populations in Germany. My thesis work revealed that the resistance mutation in Vkorc1 likely is under balancing selection and was recently introduced to the rat population in our study area. A key innovation of my thesis is adopting a NetGWAS approach to prioritize true associations and conducting co-expression network analysis to detect expression changes related to warfarin. My work shows that additional candidate genes are connected to the vitamin K pathway of which Vkorc1 is an essential component. While the validation of identified genes remains a challenge, the value of my thesis for future investigation is shown: one candidate gene Calu (Calumenin) is associated with warfarin resistance in multiple populations and is an essential part of the vitamin K cycle. Finally, my thesis briefly examines the genetics underlying a newly postulated cost of resistance, arterial calcification. This dissertation provides us an innovative framework in which we learned the genetic architecture of an adaptive trait in multiple dimensions: nucleotide or expression variation, genomic distribution and gene-gene interactions.Item The Transcriptome of an Amphioxus, Asymmetron lucayanum, from the Bahamas: A Window into Chordate Evolution(Oxford University Press, 2014) Yue, Jia-Xing; Yu, Jr-Kai; Putnam, Nicholas H.; Holland, Linda Z.Item Timing and Scope of Genomic Expansion within Annelida: Evidence from Homeoboxes in the Genome of the Earthworm Eisenia fetida(Oxford University Press, 2015) Zwarycz, Allison S.; Nossa, Carlos W.; Putnam, Nicholas H.; Ryan, Joseph F.Annelida represents a large and morphologically diverse group of bilaterian organisms. The recently published polychaete and leech genome sequences revealed an equally dynamic range of diversity at the genomic level. The availability of more annelid genomes will allow for the identification of evolutionary genomic events that helped shape the annelid lineage and better understand the diversity within the group. We sequenced and assembled the genome of the common earthworm, Eisenia fetida. As a first pass at understanding the diversity within the group, we classified 363 earthworm homeoboxes and compared them with those of the leech Helobdella robusta and the polychaete Capitella teleta. We inferred many gene expansions occurring in the lineage connecting the most recent common ancestor (MRCA) of Capitella and Eisenia to the Eisenia/Helobdella MRCA. Likewise, the lineage leading from the Eisenia/Helobdella MRCA to the leech H. robusta has experienced substantial gains and losses. However, the lineage leading from Eisenia/Helobdella MRCA to E. fetida is characterized by extraordinary levels of homeobox gain. The evolutionary dynamics observed in the homeoboxes of these lineages are very likely to be generalizable to all genes. These genome expansions and losses have likely contributed to the remarkable biology exhibited in this group. These results provide a new perspective from which to understand the diversity within these lineages, show the utility of sub-draft genome assemblies for understanding genomic evolution, and provide a critical resource from which the biology of these animals can be studied.