Publications by authors named "Igor Antoshechkin"

42 Publications

Suppressing mosquito populations with precision guided sterile males.

Nat Commun 2021 Sep 10;12(1):5374. Epub 2021 Sep 10.

Division of Biological Sciences, Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, CA, USA.

The mosquito Aedes aegypti is the principal vector for arboviruses including dengue/yellow fever, chikungunya, and Zika virus, infecting hundreds of millions of people annually. Unfortunately, traditional control methodologies are insufficient, so innovative control methods are needed. To complement existing measures, here we develop a molecular genetic control system termed precision-guided sterile insect technique (pgSIT) in Aedes aegypti. PgSIT uses a simple CRISPR-based approach to generate flightless females and sterile males that are deployable at any life stage. Supported by mathematical models, we empirically demonstrate that released pgSIT males can compete, suppress, and even eliminate mosquito populations. This platform technology could be used in the field, and adapted to many vectors, for controlling wild populations to curtail disease in a safe, confinable, and reversible manner.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-021-25421-wDOI Listing
September 2021

Engineered reproductively isolated species drive reversible population replacement.

Nat Commun 2021 06 2;12(1):3281. Epub 2021 Jun 2.

Division of Biological Sciences, Section of Cell and Developmental Biology, University of California San Diego, San Diego, CA, USA.

Engineered reproductive species barriers are useful for impeding gene flow and driving desirable genes into wild populations in a reversible threshold-dependent manner. However, methods to generate synthetic barriers are lacking in advanced eukaryotes. Here, to overcome this challenge, we engineer SPECIES (Synthetic Postzygotic barriers Exploiting CRISPR-based Incompatibilities for Engineering Species), an engineered genetic incompatibility approach, to generate postzygotic reproductive barriers. Using this approach, we create multiple reproductively isolated SPECIES and demonstrate their reproductive isolation and threshold-dependent gene drive capabilities in D. melanogaster. Given the near-universal functionality of CRISPR tools, this approach should be portable to many species, including insect disease vectors in which confinable gene drives could be of great practical utility.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-021-23531-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8173020PMC
June 2021

A Sensitive, Rapid, and Portable CasRx-based Diagnostic Assay for SARS-CoV-2.

medRxiv 2020 Oct 20. Epub 2020 Oct 20.

Since its first emergence from China in late 2019, the SARS-CoV-2 virus has spread globally despite unprecedented containment efforts, resulting in a catastrophic worldwide pandemic. Successful identification and isolation of infected individuals can drastically curtail virus spread and limit outbreaks. However, during the early stages of global transmission, point-of-care diagnostics were largely unavailable and continue to remain difficult to procure, greatly inhibiting public health efforts to mitigate spread. Furthermore, the most prevalent testing kits rely on reagent- and time-intensive protocols to detect viral RNA, preventing rapid and cost-effective diagnosis. Therefore the development of an extensive toolkit for point-of-care diagnostics that is expeditiously adaptable to new emerging pathogens is of critical public health importance. Recently, a number of novel CRISPR-based diagnostics have been developed to detect COVID-19. Herein, we outline the development of a CRISPR-based nucleic acid molecular diagnostic utilizing a Cas13d ribonuclease derived from (CasRx) to detect SARS-CoV-2, an approach we term SENSR (Sensitive Enzymatic Nucleic-acid Sequence Reporter). We demonstrate SENSR robustly detects SARS-CoV-2 sequences in both synthetic and patient-derived samples by lateral flow and fluorescence, thus expanding the available point-of-care diagnostics to combat current and future pandemics.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/2020.10.14.20212795DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7587836PMC
October 2020

Improved reference genome of the arboviral vector Aedes albopictus.

Genome Biol 2020 08 26;21(1):215. Epub 2020 Aug 26.

Genome Informatics Section, Computational and Statistical Genomics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, 20892-2152, MD, USA.

Background: The Asian tiger mosquito Aedes albopictus is globally expanding and has become the main vector for human arboviruses in Europe. With limited antiviral drugs and vaccines available, vector control is the primary approach to prevent mosquito-borne diseases. A reliable and accurate DNA sequence of the Ae. albopictus genome is essential to develop new approaches that involve genetic manipulation of mosquitoes.

Results: We use long-read sequencing methods and modern scaffolding techniques (PacBio, 10X, and Hi-C) to produce AalbF2, a dramatically improved assembly of the Ae. albopictus genome. AalbF2 reveals widespread viral insertions, novel microRNAs and piRNA clusters, the sex-determining locus, and new immunity genes, and enables genome-wide studies of geographically diverse Ae. albopictus populations and analyses of the developmental and stage-dependent network of expression data. Additionally, we build the first physical map for this species with 75% of the assembled genome anchored to the chromosomes.

Conclusion: The AalbF2 genome assembly represents the most up-to-date collective knowledge of the Ae. albopictus genome. These resources represent a foundation to improve understanding of the adaptation potential and the epidemiological relevance of this species and foster the development of innovative control measures.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13059-020-02141-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7448346PMC
August 2020

Genome elimination mediated by gene expression from a selfish chromosome.

Sci Adv 2020 04 3;6(14):eaaz9808. Epub 2020 Apr 3.

Division of Biological Sciences, Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, CA 92093, USA.

Numerous plants and animals harbor selfish B chromosomes that "drive" or transmit themselves at super-Mendelian frequencies, despite long-term fitness costs to the organism. Currently, it is unknown how B chromosome drive is mediated, and whether B-gene expression plays a role. We used modern sequencing technologies to analyze the fine-scale sequence composition and expression of paternal sex ratio (PSR), a B chromosome in the jewel wasp . PSR causes female-to-male conversion by destroying the sperm's hereditary material in young embryos to drive. Using RNA interference, we demonstrate that testis-specific expression of a PSR-linked gene, named , facilitates this genome elimination-and-sex conversion effect. encodes a putative protein with a DNA binding domain, suggesting a functional link with the sperm-derived chromatin.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/sciadv.aaz9808DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7124933PMC
April 2020

The Developmental Transcriptome of , a Major Worldwide Human Disease Vector.

G3 (Bethesda) 2020 03 5;10(3):1051-1062. Epub 2020 Mar 5.

Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, California,

mosquitoes are important vectors for a number of human pathogens including the Zika, dengue, and chikungunya viruses. Capable of displacing populations, this mosquito adapts to cooler environments which increases its geographical range and transmission potential. There are limited control strategies for mosquitoes which is likely attributed to the lack of comprehensive biological studies on this emerging vector. To fill this void, here using RNAseq we characterized mRNA expression profiles at 34 distinct time points throughout development providing the first high-resolution comprehensive view of the developmental transcriptome of this worldwide human disease vector. This enabled us to identify several patterns of shared gene expression among tissues as well as sex-specific expression patterns. To illuminate the similarities and differences with , a related human disease vector, we also performed a comparative analysis between the two developmental transcriptomes, identifying life stages where the two species exhibit similar and distinct gene expression patterns. These findings provide insights into the similarities and differences between and mosquito biology. In summary, the results generated from this study should form the basis for future investigations on the biology of and provide a gold mine resource for the development of transgene-based vector control strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1534/g3.119.401006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7056973PMC
March 2020

Broad dengue neutralization in mosquitoes expressing an engineered antibody.

PLoS Pathog 2020 01 16;16(1):e1008103. Epub 2020 Jan 16.

Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, California, United States of America.

With dengue virus (DENV) becoming endemic in tropical and subtropical regions worldwide, there is a pressing global demand for effective strategies to control the mosquitoes that spread this disease. Recent advances in genetic engineering technologies have made it possible to create mosquitoes with reduced vector competence, limiting their ability to acquire and transmit pathogens. Here we describe the development of Aedes aegypti mosquitoes synthetically engineered to impede vector competence to DENV. These mosquitoes express a gene encoding an engineered single-chain variable fragment derived from a broadly neutralizing DENV human monoclonal antibody and have significantly reduced viral infection, dissemination, and transmission rates for all four major antigenically distinct DENV serotypes. Importantly, this is the first engineered approach that targets all DENV serotypes, which is crucial for effective disease suppression. These results provide a compelling route for developing effective genetic-based DENV control strategies, which could be extended to curtail other arboviruses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.ppat.1008103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6964813PMC
January 2020

Engineered resistance to Zika virus in transgenic expressing a polycistronic cluster of synthetic small RNAs.

Proc Natl Acad Sci U S A 2019 02 5;116(9):3656-3661. Epub 2019 Feb 5.

Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, CA 92093;

Recent Zika virus (ZIKV) outbreaks have highlighted the necessity for development of novel vector control strategies to combat arboviral transmission, including genetic versions of the sterile insect technique, artificial infection with to reduce population size and/or vectoring competency, and gene drive-based methods. Here, we describe the development of mosquitoes synthetically engineered to impede vector competence to ZIKV. We demonstrate that a polycistronic cluster of engineered synthetic small RNAs targeting ZIKV is expressed and fully processed in , ensuring the formation of mature synthetic small RNAs in the midgut where ZIKV resides in the early stages of infection. Critically, we demonstrate that engineered mosquitoes harboring the anti-ZIKV transgene have significantly reduced viral infection, dissemination, and transmission rates of ZIKV. Taken together, these compelling results provide a promising path forward for development of effective genetic-based ZIKV control strategies, which could potentially be extended to curtail other arboviruses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1810771116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6397566PMC
February 2019

Improved reference genome of Aedes aegypti informs arbovirus vector control.

Nature 2018 11 14;563(7732):501-507. Epub 2018 Nov 14.

Vector Biology Department, Liverpool School of Tropical Medicine, Liverpool, UK.

Female Aedes aegypti mosquitoes infect more than 400 million people each year with dangerous viral pathogens including dengue, yellow fever, Zika and chikungunya. Progress in understanding the biology of mosquitoes and developing the tools to fight them has been slowed by the lack of a high-quality genome assembly. Here we combine diverse technologies to produce the markedly improved, fully re-annotated AaegL5 genome assembly, and demonstrate how it accelerates mosquito science. We anchored physical and cytogenetic maps, doubled the number of known chemosensory ionotropic receptors that guide mosquitoes to human hosts and egg-laying sites, provided further insight into the size and composition of the sex-determining M locus, and revealed copy-number variation among glutathione S-transferase genes that are important for insecticide resistance. Using high-resolution quantitative trait locus and population genomic analyses, we mapped new candidates for dengue vector competence and insecticide resistance. AaegL5 will catalyse new biological insights and intervention strategies to fight this deadly disease vector.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41586-018-0692-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6421076PMC
November 2018

Integrated measurement of intracellular proteins and transcripts in single cells.

Lab Chip 2018 10;18(21):3251-3262

California Institute of Techology, Division of Chemistry and Chemical Engineering, Pasadena, CA, USA.

Biological function arises from the interplay of proteins, transcripts, and metabolites. An ongoing revolution in miniaturization technologies has created tools to analyze any one of these species in single cells, thus resolving the heterogeneity of tissues previously invisible to bulk measurements. An emerging frontier is single cell multi-omics, which is the measurement of multiple classes of analytes from single cells. Here, we combine bead-based transcriptomics with microchip-based proteomics to measure intracellular proteins and transcripts from single cells and defined small numbers of cells. The transcripts and proteins are independently measured by sequencing and fluorescent immunoassays respectively, to preserve their optimal measurement modes, and linked by encoding the physical address locations of the cells into digital sequencing space using spatially patterned DNA barcodes. We resolve cell-type-specific protein and transcript signatures and present a path forward to scaling the platform to high-throughput.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/c8lc00639cDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6752714PMC
October 2018

The Female-Like State: Decoupling the Transcriptomic Effects of Aging and Sperm Status.

G3 (Bethesda) 2017 09 7;7(9):2969-2977. Epub 2017 Sep 7.

Department of Biology and Biological Engineering, and Howard Hughes Medical Institute, Caltech, Pasadena, California 91125

Understanding genome and gene function in a whole organism requires us to fully comprehend the life cycle and the physiology of the organism in question. XX animals are hermaphrodites that exhaust their sperm after 3 d of egg-laying. Even though can live for many days after cessation of egg-laying, the molecular physiology of this state has not been as intensely studied as other parts of the life cycle, despite documented changes in behavior and metabolism. To study the effects of sperm depletion and aging of during the first 6 d of adulthood, we measured the transcriptomes of first-day adult hermaphrodites and sixth-day sperm-depleted adults, and, at the same time points, mutant worms that have a feminized germline phenotype. We found that we could separate the effects of biological aging from sperm depletion. For a large subset of genes, young adult animals had the same gene expression changes as sperm-depleted sixth-day wild-type hermaphrodites, and these genes did not change expression when females reached the sixth day of adulthood. Taken together, this indicates that changing sperm status causes a change in the internal state of the worm, which we call the female-like state. Our data provide a high-quality picture of the changes that happen in global gene expression throughout the period of early aging in the worm.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1534/g3.117.300080DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5592924PMC
September 2017

Stepwise metamorphosis of the tubeworm Hydroides elegans is mediated by a bacterial inducer and MAPK signaling.

Proc Natl Acad Sci U S A 2016 09 22;113(36):10097-102. Epub 2016 Aug 22.

Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125; Howard Hughes Medical Institute, California Institute of Technology, Pasadena, CA 91125.

Diverse animal taxa metamorphose between larval and juvenile phases in response to bacteria. Although bacteria-induced metamorphosis is widespread among metazoans, little is known about the molecular changes that occur in the animal upon stimulation by bacteria. Larvae of the tubeworm Hydroides elegans metamorphose in response to surface-bound Pseudoalteromonas luteoviolacea bacteria, producing ordered arrays of phage tail-like metamorphosis-associated contractile structures (MACs). Sequencing the Hydroides genome and transcripts during five developmental stages revealed that MACs induce the regulation of groups of genes important for tissue remodeling, innate immunity, and mitogen-activated protein kinase (MAPK) signaling. Using two MAC mutations that block P. luteoviolacea from inducing settlement or metamorphosis and three MAPK inhibitors, we established a sequence of bacteria-induced metamorphic events: MACs induce larval settlement; then, particular properties of MACs encoded by a specific locus in P. luteoviolacea initiate cilia loss and activate metamorphosis-associated transcription; finally, signaling through p38 and c-Jun N-terminal kinase (JNK) MAPK pathways alters gene expression and leads to morphological changes upon initiation of metamorphosis. Our results reveal that the intricate interaction between Hydroides and P. luteoviolacea can be dissected using genomic, genetic, and pharmacological tools. Hydroides' dependency on bacteria for metamorphosis highlights the importance of external stimuli to orchestrate animal development. The conservation of Hydroides genome content with distantly related deuterostomes (urchins, sea squirts, and humans) suggests that mechanisms of bacteria-induced metamorphosis in Hydroides may have conserved features in diverse animals. As a major biofouling agent, insight into the triggers of Hydroides metamorphosis might lead to practical strategies for fouling control.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1603142113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5018781PMC
September 2016

Comparative genomics of Steinernema reveals deeply conserved gene regulatory networks.

Genome Biol 2015 Sep 21;16:200. Epub 2015 Sep 21.

Department of Developmental and Cell Biology, University of California, Irvine, CA, 92697, USA.

Background: Parasitism is a major ecological niche for a variety of nematodes. Multiple nematode lineages have specialized as pathogens, including deadly parasites of insects that are used in biological control. We have sequenced and analyzed the draft genomes and transcriptomes of the entomopathogenic nematode Steinernema carpocapsae and four congeners (S. scapterisci, S. monticolum, S. feltiae, and S. glaseri).

Results: We used these genomes to establish phylogenetic relationships, explore gene conservation across species, and identify genes uniquely expanded in insect parasites. Protein domain analysis in Steinernema revealed a striking expansion of numerous putative parasitism genes, including certain protease and protease inhibitor families, as well as fatty acid- and retinol-binding proteins. Stage-specific gene expression of some of these expanded families further supports the notion that they are involved in insect parasitism by Steinernema. We show that sets of novel conserved non-coding regulatory motifs are associated with orthologous genes in Steinernema and Caenorhabditis.

Conclusions: We have identified a set of expanded gene families that are likely to be involved in parasitism. We have also identified a set of non-coding motifs associated with groups of orthologous genes in Steinernema and Caenorhabditis involved in neurogenesis and embryonic development that are likely part of conserved protein-DNA relationships shared between these two genera.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13059-015-0746-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4578762PMC
September 2015

The genome and transcriptome of the zoonotic hookworm Ancylostoma ceylanicum identify infection-specific gene families.

Nat Genet 2015 Apr 2;47(4):416-22. Epub 2015 Mar 2.

1] Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts, USA. [2] Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, California, USA.

Hookworms infect over 400 million people, stunting and impoverishing them. Sequencing hookworm genomes and finding which genes they express during infection should help in devising new drugs or vaccines against hookworms. Unlike other hookworms, Ancylostoma ceylanicum infects both humans and other mammals, providing a laboratory model for hookworm disease. We determined an A. ceylanicum genome sequence of 313 Mb, with transcriptomic data throughout infection showing expression of 30,738 genes. Approximately 900 genes were upregulated during early infection in vivo, including ASPRs, a cryptic subfamily of activation-associated secreted proteins (ASPs). Genes downregulated during early infection included ion channels and G protein-coupled receptors; this downregulation was observed in both parasitic and free-living nematodes. Later, at the onset of heavy blood feeding, C-lectin genes were upregulated along with genes for secreted clade V proteins (SCVPs), encoding a previously undescribed protein family. These findings provide new drug and vaccine targets and should help elucidate hookworm pathogenesis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/ng.3237DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4617383PMC
April 2015

A comparative encyclopedia of DNA elements in the mouse genome.

Nature 2014 Nov;515(7527):355-64

Bioinformatics and Genomics, Centre for Genomic Regulation (CRG) and UPF, Doctor Aiguader, 88, 08003 Barcelona, Catalonia, Spain.

The laboratory mouse shares the majority of its protein-coding genes with humans, making it the premier model organism in biomedical research, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature13992DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4266106PMC
November 2014

Pol II docking and pausing at growth and stress genes in C. elegans.

Cell Rep 2014 Feb 30;6(3):455-66. Epub 2014 Jan 30.

Department of Biology, Duke Center for Systems Biology, Duke University, Durham, NC 27708, USA. Electronic address:

Fluctuations in nutrient availability profoundly impact gene expression. Previous work revealed postrecruitment regulation of RNA polymerase II (Pol II) during starvation and recovery in Caenorhabditis elegans, suggesting that promoter-proximal pausing promotes rapid response to feeding. To test this hypothesis, we measured Pol II elongation genome wide by two complementary approaches and analyzed elongation in conjunction with Pol II binding and expression. We confirmed bona fide pausing during starvation and also discovered Pol II docking. Pausing occurs at active stress-response genes that become downregulated in response to feeding. In contrast, "docked" Pol II accumulates without initiating upstream of inactive growth genes that become rapidly upregulated upon feeding. Beyond differences in function and expression, these two sets of genes have different core promoter motifs, suggesting alternative transcriptional machinery. Our work suggests that growth and stress genes are both regulated postrecruitment during starvation but at initiation and elongation, respectively, coordinating gene expression with nutrient availability.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2014.01.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4026043PMC
February 2014

Transcriptome analysis reveals novel players in the cranial neural crest gene regulatory network.

Genome Res 2014 Feb 3;24(2):281-90. Epub 2014 Jan 3.

Division of Biology, California Institute of Technology, Pasadena, California 91125, USA;

The neural crest is an embryonic stem cell population that gives rise to a multitude of derivatives. In particular, the cranial neural crest (CNC) is unique in its ability to contribute to both facial skeleton and peripheral ganglia. To gain further insight into the molecular underpinnings that distinguish the CNC from other embryonic tissues, we have utilized a CNC-specific enhancer as a tool to isolate a pure, region-specific NC subpopulation for transcriptional profiling. The resulting data set reveals previously unknown transcription factors and signaling pathways that may influence the CNC's ability to migrate and/or differentiate into unique derivatives. To elaborate on the CNC gene regulatory network, we evaluated the effects of knocking down known neural plate border genes and early neural crest specifier genes on selected neural crest-enriched transcripts. The results suggest that ETS1 and SOX9 may act as pan-neural crest regulators of the migratory CNC. Taken together, our analysis provides unprecedented characterization of the migratory CNC transcriptome and identifies new links in the gene regulatory network responsible for development of this critical cell population.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gr.161182.113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3912418PMC
February 2014

The genome and developmental transcriptome of the strongylid nematode Haemonchus contortus.

Genome Biol 2013 Aug 28;14(8):R89. Epub 2013 Aug 28.

Background: The barber's pole worm, Haemonchus contortus, is one of the most economically important parasites of small ruminants worldwide. Although this parasite can be controlled using anthelmintic drugs, resistance against most drugs in common use has become a widespread problem. We provide a draft of the genome and the transcriptomes of all key developmental stages of H. contortus to support biological and biotechnological research areas of this and related parasites.

Results: The draft genome of H. contortus is 320 Mb in size and encodes 23,610 protein-coding genes. On a fundamental level, we elucidate transcriptional alterations taking place throughout the life cycle, characterize the parasite's gene silencing machinery, and explore molecules involved in development, reproduction, host-parasite interactions, immunity, and disease. The secretome of H. contortus is particularly rich in peptidases linked to blood-feeding activity and interactions with host tissues, and a diverse array of molecules is involved in complex immune responses. On an applied level, we predict drug targets and identify vaccine molecules.

Conclusions: The draft genome and developmental transcriptome of H. contortus provide a major resource to the scientific community for a wide range of genomic, genetic, proteomic, metabolomic, evolutionary, biological, ecological, and epidemiological investigations, and a solid foundation for biotechnological outcomes, including new anthelmintics, vaccines and diagnostic tests. This first draft genome of any strongylid nematode paves the way for a rapid acceleration in our understanding of a wide range of socioeconomically important parasites of one of the largest nematode orders.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/gb-2013-14-8-r89DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4053716PMC
August 2013

Transgene-free genome editing in Caenorhabditis elegans using CRISPR-Cas.

Genetics 2013 Nov 26;195(3):1167-71. Epub 2013 Aug 26.

Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125.

CRISPR-Cas is an efficient method for genome editing in organisms from bacteria to human cells. We describe a transgene-free method for CRISPR-Cas-mediated cleavage in nematodes, enabling RNA-homology-targeted deletions that cause loss of gene function; analysis of whole-genome sequencing indicates that the nuclease activity is highly specific.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1534/genetics.113.155879DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3813845PMC
November 2013

Transcriptome profiling of Nasonia vitripennis testis reveals novel transcripts expressed from the selfish B chromosome, paternal sex ratio.

G3 (Bethesda) 2013 Sep 4;3(9):1597-605. Epub 2013 Sep 4.

Division of Biology, MC156-29, California Institute of Technology, Pasadena, California 91125.

A widespread phenomenon in nature is sex ratio distortion of arthropod populations caused by microbial and genetic parasites. Currently little is known about how these agents alter host developmental processes to favor one sex or the other. The paternal sex ratio (PSR) chromosome is a nonessential, paternally transmitted centric fragment that segregates in natural populations of the jewel wasp, Nasonia vitripennis. To persist, PSR is thought to modify the hereditary material of the developing sperm, with the result that all nuclear DNA other than the PSR chromosome is destroyed shortly after fertilization. This results in the conversion of a fertilized embryo--normally a female--into a male, thereby insuring transmission of the "selfish" PSR chromosome, and simultaneously leading to wasp populations that are male-biased. To begin to understand this system at the mechanistic level, we carried out transcriptional profiling of testis from WT and PSR-carrying males. We identified a number of transcripts that are differentially expressed between these conditions. We also discovered nine transcripts that are uniquely expressed from the PSR chromosome. Four of these PSR-specific transcripts encode putative proteins, whereas the others have very short open reading frames and no homology to known proteins, suggesting that they are long noncoding RNAs. We propose several different models for how these transcripts could facilitate PSR-dependent effects. Our analyses also revealed 15.71 MB of novel transcribed regions in the N. vitripennis genome, thus increasing the current annotation of total transcribed regions by 53.4%. Finally, we detected expression of multiple meiosis-related genes in the wasp testis, despite the lack of conventional meiosis in the male sex.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1534/g3.113.007583DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3755920PMC
September 2013

The developmental transcriptome of the mosquito Aedes aegypti, an invasive species and major arbovirus vector.

G3 (Bethesda) 2013 Sep 4;3(9):1493-509. Epub 2013 Sep 4.

Division of Biology, MC 156-29, California Institute of Technology, Pasadena, California 91125.

Mosquitoes are vectors of a number of important human and animal diseases. The development of novel vector control strategies requires a thorough understanding of mosquito biology. To facilitate this, we used RNA-seq to identify novel genes and provide the first high-resolution view of the transcriptome throughout development and in response to blood feeding in a mosquito vector of human disease, Aedes aegypti, the primary vector for Dengue and yellow fever. We characterized mRNA expression at 34 distinct time points throughout Aedes development, including adult somatic and germline tissues, by using polyA+ RNA-seq. We identify a total of 14,238 novel new transcribed regions corresponding to 12,597 new loci, as well as many novel transcript isoforms of previously annotated genes. Altogether these results increase the annotated fraction of the transcribed genome into long polyA+ RNAs by more than twofold. We also identified a number of patterns of shared gene expression, as well as genes and/or exons expressed sex-specifically or sex-differentially. Expression profiles of small RNAs in ovaries, early embryos, testes, and adult male and female somatic tissues also were determined, resulting in the identification of 38 new Aedes-specific miRNAs, and ~291,000 small RNA new transcribed regions, many of which are likely to be endogenous small-interfering RNAs and Piwi-interacting RNAs. Genes of potential interest for transgene-based vector control strategies also are highlighted. Our data have been incorporated into a user-friendly genome browser located at www.Aedes.caltech.edu, with relevant links to Vectorbase (www.vectorbase.org).
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1534/g3.113.006742DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3755910PMC
September 2013

Novel synthetic Medea selfish genetic elements drive population replacement in Drosophila; a theoretical exploration of Medea-dependent population suppression.

ACS Synth Biol 2014 Dec 28;3(12):915-28. Epub 2012 Dec 28.

Division of Biology, MC 156-29, California Institute of Technology , Pasadena, California 91125, United States.

Insects act as vectors for diseases of plants, animals, and humans. Replacement of wild insect populations with genetically modified individuals unable to transmit disease provides a potentially self-perpetuating method of disease prevention. Population replacement requires a gene drive mechanism in order to spread linked genes mediating disease refractoriness through wild populations. We previously reported the creation of synthetic Medea selfish genetic elements able to drive population replacement in Drosophila. These elements use microRNA-mediated silencing of myd88, a maternally expressed gene required for embryonic dorso-ventral pattern formation, coupled with early zygotic expression of a rescuing transgene, to bring about gene drive. Medea elements that work through additional mechanisms are needed in order to be able to carry out cycles of population replacement and/or remove existing transgenes from the population, using second-generation elements that spread while driving first-generation elements out of the population. Here we report the synthesis and population genetic behavior of two new synthetic Medea elements that drive population replacement through manipulation of signaling pathways involved in cellular blastoderm formation or Notch signaling, demonstrating that in Drosophila Medea elements can be generated through manipulation of diverse signaling pathways. We also describe the mRNA and small RNA changes in ovaries and early embryos associated from Medea-bearing females. Finally, we use modeling to illustrate how Medea elements carrying genes that result in diapause-dependent female lethality could be used to bring about population suppression.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/sb300079hDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3742681PMC
December 2014

Divergent mitochondrial biogenesis responses in human cardiomyopathy.

Circulation 2013 May 15;127(19):1957-67. Epub 2013 Apr 15.

Department of Anesthesiology, Division of Molecular Medicine, David Geffen School of Medicine at UCLA, BH-569 CHS, BOX 957115, Los Angeles, CA 90095, USA.

Background: Mitochondria are key players in the development and progression of heart failure (HF). Mitochondrial (mt) dysfunction leads to diminished energy production and increased cell death contributing to the progression of left ventricular failure. The fundamental mechanisms that underlie mt dysfunction in HF have not been fully elucidated.

Methods And Results: To characterize mt morphology, biogenesis, and genomic integrity in human HF, we investigated left ventricular tissue from nonfailing hearts and end-stage ischemic (ICM) or dilated (DCM) cardiomyopathic hearts. Although mt dysfunction was present in both types of cardiomyopathy, mt were smaller and increased in number in DCM compared with ICM or nonfailing hearts. mt volume density and mtDNA copy number was increased by ≈2-fold (P<0.001) in DCM hearts in comparison with ICM hearts. These changes were accompanied by an increase in the expression of mtDNA-encoded genes in DCM versus no change in ICM. mtDNA repair and antioxidant genes were reduced in failing hearts, suggestive of a defective repair and protection system, which may account for the 4.1-fold increase in mtDNA deletion mutations in DCM (P<0.05 versus nonfailing hearts, P<0.05 versus ICM).

Conclusions: In DCM, mt dysfunction is associated with mtDNA damage and deletions, which could be a consequence of mutating stress coupled with a peroxisome proliferator-activated receptor γ coactivator 1α-dependent stimulus for mt biogenesis. However, this maladaptive compensatory response contributes to additional oxidative damage. Thus, our findings support further investigations into novel mechanisms and therapeutic strategies for mt dysfunction in DCM.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1161/CIRCULATIONAHA.112.001219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4236313PMC
May 2013

The draft genome and transcriptome of Panagrellus redivivus are shaped by the harsh demands of a free-living lifestyle.

Genetics 2013 Apr 14;193(4):1279-95. Epub 2013 Feb 14.

Division of Biology, California Institute of Technology, Pasadena, CA 91125, USA.

Nematodes compose an abundant and diverse invertebrate phylum with members inhabiting nearly every ecological niche. Panagrellus redivivus (the "microworm") is a free-living nematode frequently used to understand the evolution of developmental and behavioral processes given its phylogenetic distance to Caenorhabditis elegans. Here we report the de novo sequencing of the genome, transcriptome, and small RNAs of P. redivivus. Using a combination of automated gene finders and RNA-seq data, we predict 24,249 genes and 32,676 transcripts. Small RNA analysis revealed 248 microRNA (miRNA) hairpins, of which 63 had orthologs in other species. Fourteen miRNA clusters containing 42 miRNA precursors were found. The RNA interference, dauer development, and programmed cell death pathways are largely conserved. Analysis of protein family domain abundance revealed that P. redivivus has experienced a striking expansion of BTB domain-containing proteins and an unprecedented expansion of the cullin scaffold family of proteins involved in multi-subunit ubiquitin ligases, suggesting proteolytic plasticity and/or tighter regulation of protein turnover. The eukaryotic release factor protein family has also been dramatically expanded and suggests an ongoing evolutionary arms race with viruses and transposons. The P. redivivus genome provides a resource to advance our understanding of nematode evolution and biology and to further elucidate the genomic architecture leading to free-living lineages, taking advantage of the many fascinating features of this worm revealed by comparative studies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1534/genetics.112.148809DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3606103PMC
April 2013

A large-scale in vivo analysis reveals that TALENs are significantly more mutagenic than ZFNs generated using context-dependent assembly.

Nucleic Acids Res 2013 Feb 8;41(4):2769-78. Epub 2013 Jan 8.

Division of Biology, California Institute of Technology, Pasadena, CA 91125, USA.

Zinc-finger nucleases (ZFNs) and TAL effector nucleases (TALENs) have been shown to induce targeted mutations, but they have not been extensively tested in any animal model. Here, we describe a large-scale comparison of ZFN and TALEN mutagenicity in zebrafish. Using deep sequencing, we found that TALENs are significantly more likely to be mutagenic and induce an average of 10-fold more mutations than ZFNs. We observed a strong correlation between somatic and germ-line mutagenicity, and identified germ line mutations using ZFNs whose somatic mutations rates are well below the commonly used threshold of 1%. Guidelines that have previously been proposed to predict optimal ZFN and TALEN target sites did not predict mutagenicity in vivo. However, we observed a significant negative correlation between TALEN mutagenicity and the number of CpG repeats in TALEN target sites, suggesting that target site methylation may explain the poor mutagenicity of some TALENs in vivo. The higher mutation rates and ability to target essentially any sequence make TALENs the superior technology for targeted mutagenesis in zebrafish, and likely other animal models.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/nar/gks1356DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3575824PMC
February 2013

Landscape of transcription in human cells.

Nature 2012 Sep;489(7414):101-8

Centre for Genomic Regulation and UPF, Doctor Aiguader 88, Barcelona 08003, Catalonia, Spain.

Eukaryotic cells make many types of primary and processed RNAs that are found either in specific subcellular compartments or throughout the cells. A complete catalogue of these RNAs is not yet available and their characteristic subcellular localizations are also poorly understood. Because RNA represents the direct output of the genetic information encoded by genomes and a significant proportion of a cell's regulatory capabilities are focused on its synthesis, processing, transport, modification and translation, the generation of such a catalogue is crucial for understanding genome function. Here we report evidence that three-quarters of the human genome is capable of being transcribed, as well as observations about the range and levels of expression, localization, processing fates, regulatory regions and modifications of almost all currently annotated and thousands of previously unannotated RNAs. These observations, taken together, prompt a redefinition of the concept of a gene.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature11233DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3684276PMC
September 2012

Nutritional control of mRNA isoform expression during developmental arrest and recovery in C. elegans.

Genome Res 2012 Oct 26;22(10):1920-9. Epub 2012 Apr 26.

Department of Biology, Duke Center for Systems Biology, Duke University, Durham, North Carolina 27708, USA.

Nutrient availability profoundly influences gene expression. Many animal genes encode multiple transcript isoforms, yet the effect of nutrient availability on transcript isoform expression has not been studied in genome-wide fashion. When Caenorhabditis elegans larvae hatch without food, they arrest development in the first larval stage (L1 arrest). Starved larvae can survive L1 arrest for weeks, but growth and post-embryonic development are rapidly initiated in response to feeding. We used RNA-seq to characterize the transcriptome during L1 arrest and over time after feeding. Twenty-seven percent of detectable protein-coding genes were differentially expressed during recovery from L1 arrest, with the majority of changes initiating within the first hour, demonstrating widespread, acute effects of nutrient availability on gene expression. We used two independent approaches to track expression of individual exons and mRNA isoforms, and we connected changes in expression to functional consequences by mining a variety of databases. These two approaches identified an overlapping set of genes with alternative isoform expression, and they converged on common functional patterns. Genes affecting mRNA splicing and translation are regulated by alternative isoform expression, revealing post-transcriptional consequences of nutrient availability on gene regulation. We also found that phosphorylation sites are often alternatively expressed, revealing a common mode by which alternative isoform expression modifies protein function and signal transduction. Our results detail rich changes in C. elegans gene expression as larvae initiate growth and post-embryonic development, and they provide an excellent resource for ongoing investigation of transcriptional regulation and developmental physiology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gr.133587.111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3460187PMC
October 2012
-->