Yong-Hwee Eddie Loh

Survey Sequencing and Comparative Analysis of the Elephant Shark (Callorhinchus milii) Genome

Owing to their phylogenetic position, cartilaginous fishes (sharks, rays, skates, and chimaeras) provide a critical reference for our understanding of vertebrate genome evolution. The relatively small genome of the elephant shark, Callorhinchus milii, a chimaera, makes it an attractive model cartilaginous fish genome for whole-genome sequencing and comparative analysis. Here, the authors describe survey sequencing (1.4× coverage) and comparative analysis of the elephant shark genome, one of the first cartilaginous fish genomes to be sequenced to this depth. Repetitive sequences, represented mainly by a novel family of short interspersed element–like and long interspersed element–like sequences, account for about 28% of the elephant shark genome. Fragments of approximately 15,000 elephant shark genes reveal specific examples of genes that have been lost differentially during the evolution of tetrapod and teleost fish lineages. Interestingly, the degree of conserved synteny and conserved sequences between the human and elephant shark genomes are higher than that between human and teleost fish genomes. Elephant shark contains putative four Hox clusters indicating that, unlike teleost fish genomes, the elephant shark genome has not experienced an additional whole-genome duplication. These findings underscore the importance of the elephant shark as a critical reference vertebrate genome for comparative analysis of the human and other vertebrate genomes. This study also demonstrates that a survey-sequencing approach can be applied productively for comparative analysis of distantly related vertebrate genomes.

Ancient noncoding elements conserved in the human genome.

Cartilaginous fishes represent the living group of jawed vertebrates that diverged from the common ancestor of human and teleost fish lineages about 530 million years ago. We generated ~1.4× genome sequence coverage for a cartilaginous fish, the elephant shark (Callorhinchus milii), and compared this genome with the human genome to identify conserved noncoding elements (CNEs). The elephant shark sequence revealed twice as many CNEs as were identified by whole-genome comparisons between teleost fishes and human. The ancient vertebrate-specific CNEs in the elephant shark and human genomes are likely to play key regulatory roles in vertebrate gene expression.

Comparative analysis reveals signatures of differentiation amid genomic polymorphism in Lake Malawi cichlids

BackgroundCichlid fish from East Africa are remarkable for phenotypic and behavioral diversity on a backdrop of genomic similarity. In 2006, the Joint Genome Institute completed low coverage survey sequencing of the genomes of five phenotypically and ecologically diverse Lake Malawi species. We report a computational and comparative analysis of these data that provides insight into the mechanisms that make closely related species different from one another.ResultsWe produced assemblies for the five species ranging in aggregate length from 68 to 79 megabase pairs, identified putative orthologs for more than 12,000 human genes, and predicted more than 32,000 cross-species single nucleotide polymorphisms (SNPs). Nucleotide diversity was lower than that found among laboratory strains of the zebrafish. We collected around 36,000 genotypes to validate a subset of SNPs within and among populations and across multiple individuals of about 75 Lake Malawi species. Notably, there were no fixed differences observed between focal species nor between major lineages. Roughly 3% to 5% of loci surveyed are statistical outliers for genetic differentiation (FST) within species, between species, and between major lineages. Outliers for FST are candidate genes that may have experienced a history of natural selection in the Malawi lineage.ConclusionWe present a novel genome sequencing strategy, which is useful when evolutionary diversity is the question of interest. Lake Malawi cichlids are phenotypically and behaviorally diverse, but they appear genetically like a subdivided population. The unique structure of Lake Malawl cichlid genomes should facilitate conceptually new experiments, employing SNPs to identity genotype-phenotype association, using the entire species flock as a mapping panel.

Genetic Adaptation and Neandertal Admixture Shaped the Immune System of Human Populations

Summary Humans differ in the outcome that follows exposure to life-threatening pathogens, yet the extent of population differences in immune responses and their genetic and evolutionary determinants remain undefined. Here, we characterized, using RNA sequencing, the transcriptional response of primary monocytes from Africans and Europeans to bacterial and viral stimuli—ligands activating Toll-like receptor pathways (TLR1/2, TLR4, and TLR7/8) and influenza virus—and mapped expression quantitative trait loci (eQTLs). We identify numerous cis-eQTLs that contribute to the marked differences in immune responses detected within and between populations and a strong trans-eQTL hotspot at TLR1 that decreases expression of pro-inflammatory genes in Europeans only. We find that immune-responsive regulatory variants are enriched in population-specific signals of natural selection and show that admixture with Neandertals introduced regulatory variants into European genomes, affecting preferentially responses to viral challenges. Together, our study uncovers evolutionarily important determinants of differences in host immune responsiveness between human populations.

Origins of Shared Genetic Variation in African Cichlids

Cichlid fishes have evolved tremendous morphological and behavioral diversity in the waters of East Africa. Within each of the Great Lakes Tanganyika, Malawi, and Victoria, the phenomena of hybridization and retention of ancestral polymorphism explain allele sharing across species. Here, we explore the sharing of single nucleotide polymorphisms (SNPs) between the major East African cichlid assemblages. A set of approximately 200 genic and nongenic SNPs was ascertained in five Lake Malawi species and genotyped in a diverse collection of ∼160 species from across Africa. We observed segregating polymorphism outside of the Malawi lineage for more than 50% of these loci; this holds similarly for genic versus nongenic SNPs, as well as for SNPs at putative CpG versus non-CpG sites. Bayesian and principal component analyses of genetic structure in the data demonstrate that the Lake Malawi endemic flock is not monophyletic and that river species have likely contributed significantly to Malawi genomes. Coalescent simulations support the hypothesis that river cichlids have transported polymorphism between lake assemblages. We observed strong genetic differentiation between Malawi lineages for approximately 8% of loci, with contributions from both genic and nongenic SNPs. Notably, more than half of these outlier loci between Malawi groups are polymorphic outside of the lake. Cichlid fishes have evolved diversity in Lake Malawi as new mutations combined with standing genetic variation shared across East Africa.

Evolution of MicroRNAs and the Diversification of Species

MicroRNAs (miRNAs) are ancient, short noncoding RNA molecules that regulate the transcriptome through post-transcriptional mechanisms. miRNA riboregulation is involved in a diverse range of biological processes, and misregulation is implicated in disease. It is generally thought that miRNAs function to canalize cellular outputs, for instance as “fail-safe” repressors of gene misexpression. Genomic surveys in humans have revealed reduced genetic polymorphism and the signature of negative selection for both miRNAs themselves and the target sequences to which they are predicted to bind. We investigated the evolution of miRNAs and their binding sites across cichlid fishes from Lake Malawi (East Africa), where hundreds of diverse species have evolved in the last million years. Using low-coverage genome sequence data, we identified 100 cichlid miRNA genes with mature regions that are highly conserved in other animal species. We computationally predicted target sites on the 3′-untranslated regions (3′-UTRs) of cichlid genes to which miRNAs may bind and found that these sites possessed elevated single nucleotide polymorphism (SNP) densities. Furthermore, polymorphic sites in predicted miRNA targets showed higher minor allele frequencies on average and greater genetic differentiation between Malawi lineages when compared with a neutral expectation and nontarget 3′-UTR SNPs. Our data suggest that divergent selection on miRNA riboregulation may have contributed to the diversification of cichlid species and may similarly play a role in rapid phenotypic evolution of other natural systems.

Early life stress confers lifelong stress susceptibility in mice via ventral tegmental area OTX2

An early window of stress susceptibility defines a mouse’s response to stress in adulthood. Early life stress in depression susceptibility The linkage between stress early in life and behavioral depression in adulthood is complex. Peña et al. were able to define a time period in early development when mice are especially susceptible to stress. Mice subjected to stress during this time period were less resilient to stress in adulthood. Genes regulated by the transcription factor orthodenticle homeobox 2 (OTX2) primed the response toward depression in adulthood. Although early stress could establish the groundwork for later depression, that priming could be undone by intervention at the right moment. Science, this issue p. 1185 Early life stress increases risk for depression. Here we establish a “two-hit” stress model in mice wherein stress at a specific postnatal period increases susceptibility to adult social defeat stress and causes long-lasting transcriptional alterations that prime the ventral tegmental area (VTA)—a brain reward region—to be in a depression-like state. We identify a role for the developmental transcription factor orthodenticle homeobox 2 (Otx2) as an upstream mediator of these enduring effects. Transient juvenile—but not adult—knockdown of Otx2 in VTA mimics early life stress by increasing stress susceptibility, whereas its overexpression reverses the effects of early life stress. This work establishes a mechanism by which early life stress encodes lifelong susceptibility to stress via long-lasting transcriptional programming in VTA mediated by Otx2.

Sex-specific transcriptional signatures in human depression

Major depressive disorder (MDD) is a leading cause of disease burden worldwide. While the incidence, symptoms and treatment of MDD all point toward major sex differences, the molecular mechanisms underlying this sexual dimorphism remain largely unknown. Here, combining differential expression and gene coexpression network analyses, we provide a comprehensive characterization of male and female transcriptional profiles associated with MDD across six brain regions. We overlap our human profiles with those from a mouse model, chronic variable stress, and capitalize on converging pathways to define molecular and physiological mechanisms underlying the expression of stress susceptibility in males and females. Our results show a major rearrangement of transcriptional patterns in MDD, with limited overlap between males and females, an effect seen in both depressed humans and stressed mice. We identify key regulators of sex-specific gene networks underlying MDD and confirm their sex-specific impact as mediators of stress susceptibility. For example, downregulation of the female-specific hub gene Dusp6 in mouse prefrontal cortex mimicked stress susceptibility in females, but not males, by increasing ERK signaling and pyramidal neuron excitability. Such Dusp6 downregulation also recapitulated the transcriptional remodeling that occurs in prefrontal cortex of depressed females. Together our findings reveal marked sexual dimorphism at the transcriptional level in MDD and highlight the importance of studying sex-specific treatments for this disorder.

Lake Malawi cichlid evolution along a benthic/limnetic axis

Divergence along a benthic to limnetic habitat axis is ubiquitous in aquatic systems. However, this type of habitat divergence has largely been examined in low diversity, high latitude lake systems. In this study, we examined the importance of benthic and limnetic divergence within the incredibly species-rich radiation of Lake Malawi cichlid fishes. Using novel phylogenetic reconstructions, we provided a series of hypotheses regarding the evolutionary relationships among 24 benthic and limnetic species that suggests divergence along this axis has occurred multiple times within Lake Malawi cichlids. Because pectoral fin morphology is often associated with divergence along this habitat axis in other fish groups, we investigated divergence in pectoral fin muscles in these benthic and limnetic cichlid species. We showed that the eight pectoral fin muscles and fin area generally tended to evolve in a tightly correlated manner in the Lake Malawi cichlids. Additionally, we found that larger pectoral fin muscles are strongly associated with the independent evolution of the benthic feeding habit across this group of fish. Evolutionary specialization along a benthic/limnetic axis has occurred multiple times within this tropical lake radiation and has produced repeated convergent matching between exploitation of water column habitats and locomotory morphology.

ENL links histone acetylation to oncogenic gene expression in acute myeloid leukaemia

Cancer cells are characterized by aberrant epigenetic landscapes and often exploit chromatin machinery to activate oncogenic gene expression programs. Recognition of modified histones by ‘reader’ proteins constitutes a key mechanism underlying these processes; therefore, targeting such pathways holds clinical promise, as exemplified by the development of bromodomain and extra-terminal (BET) inhibitors. We recently identified the YEATS domain as an acetyl-lysine-binding module, but its functional importance in human cancer remains unknown. Here we show that the YEATS domain-containing protein ENL, but not its paralogue AF9, is required for disease maintenance in acute myeloid leukaemia. CRISPR–Cas9-mediated depletion of ENL led to anti-leukaemic effects, including increased terminal myeloid differentiation and suppression of leukaemia growth in vitro and in vivo. Biochemical and crystal structural studies and chromatin-immunoprecipitation followed by sequencing analyses revealed that ENL binds to acetylated histone H3, and co-localizes with H3K27ac and H3K9ac on the promoters of actively transcribed genes that are essential for leukaemia. Disrupting the interaction between the YEATS domain and histone acetylation via structure-based mutagenesis reduced the recruitment of RNA polymerase II to ENL-target genes, leading to the suppression of oncogenic gene expression programs. Notably, disrupting the functionality of ENL further sensitized leukaemia cells to BET inhibitors. Together, our data identify ENL as a histone acetylation reader that regulates oncogenic transcriptional programs in acute myeloid leukaemia, and suggest that displacement of ENL from chromatin may be a promising epigenetic therapy, alone or in combination with BET inhibitors, for aggressive leukaemia.