Suchergebnisse

Regulation of male sexual differentiation by a Y chromosome-linked male determining factor (M-factor) is one of a diverse array of sex determination mechanisms found in insects. By deep sequencing of small RNAs from Bactrocera dorsalis early embryos, we identified an autosomal-derived microRNA, miR-1-3p, that has predicted target sites in the transformer gene (Bdtra) required for female sex determination. We further demonstrate by both in vitro and in vivo tests that miR-1-3p suppresses Bdtra expression. Injection of a miR-1-3p mimic in early embryos results in 87-92% phenotypic males, whereas knockdown of miR-1-3p by an inhibitor results in 67-77% phenotypic females. Finally, CRISPR/Cas9-mediated knockout of miR-1-3p results in the expression of female-specific splice variants of Bdtra and doublesex (Bddsx), and induced sex reversal of XY individuals into phenotypic females. These results indicate that miR-1-3p is required for male sex determination in early embryogenesis in B. dorsalis as an intermediate male determiner. ; National Key R&D Program of China [2017YFD0202000]; earmarked fund for the China Agricultural Research System [CARS-26]; International Atomic Energy Agency's Coordinated Research Project [D42016]; National Natural Science Foundation of ChinaNational Natural Science Foundation of China (NSFC) [31872931, 31572008]; Fundamental Research Funds for the Central UniversitiesFundamental Research Funds for the Central Universities [2662019PY055]; USDA-NIFA-AFRI Foundational Program [2016-67013-25087]; FFABR2017; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [AI123338, AI121284] ; This research was supported by National Key R&D Program of China (No. 2017YFD0202000), the earmarked fund for the China Agricultural Research System (No. CARS-26), International Atomic Energy Agency's Coordinated Research Project (No. D42016), the National Natural Science Foundation of China (Nos. 31872931 and 31572008) and the Fundamental Research Funds for the Central Universities (No. 2662019PY055) to H.Z. Support is also acknowledged from the USDA-NIFA-AFRI Foundational Program (No. 2016-67013-25087) to AMH, grant FFABR2017 to G.S., and NIH grants AI123338 and AI121284 to Z.J.T. We thank Dr. Eric Jang from USDA-ARS, Pacific West Area, Tropical Crop and Commodity Protection Research Unit, for providing the B. dorsalis white/brown pupae sexing strain. ; Public domain authored by a U.S. government employee

Complementing genome sequence with deep transcriptome and proteome data could enable more accurate assembly and annotation of newly sequenced genomes. Here, we provide a proof-of-concept of an integrated approach for analysis of the genome and proteome of Anopheles stephensi, which is one of the most important vectors of the malaria parasite. To achieve broad coverage of genes, we carried out transcriptome sequencing and deep proteome profiling of multiple anatomically distinct sites. Based on transcriptomic data alone, we identified and corrected 535 events of incomplete genome assembly involving 1196 scaffolds and 868 protein-coding gene models. This proteogenomic approach enabled us to add 365 genes that were missed during genome annotation and identify 917 gene correction events through discovery of 151 novel exons, 297 protein extensions, 231 exon extensions,192 novel protein start sites,19 novel translational frames, 28 events of joining of exons, and 76 events of joining of adjacent genes as a single gene. Incorporation of proteomic evidence allowed us to change the designation of more than 87 predicted "noncoding RNAs" to conventional mRNAs coded by protein-coding genes. Importantly, extension of the newly corrected genome assemblies and gene models to 15 other newly assembled Anopheline genomes led to the discovery of a large number of apparent discrepancies in assembly and annotation of these genomes. Our data provide a framework for how future genome sequencing efforts should incorporate transcriptomic and proteomic analysis in combination with simultaneous manual curation to achieve near complete assembly and accurate annotation of genomes. ; Science and Engineering Research Board (SERB), Department of Science and Technology, Government of India [EMR/2014/000444]; DBT Program Support grant on "Development of infrastructure and a computational framework for analysis of proteomic data" [BT/01/COE/08/05]; Infosys Foundation; pilot grant from the Johns Hopkins Malaria Research Institute; Council of Scientific and Industrial Research, Department of Biotechnology, University Grants Commission; Department of Science and Technology, Government of India; Indian Council of Medical Research ; This paper is funded by the joint research project to NIMR and IOB entitled "Characterization of Malaria vector Anopheles stephensi Proteome and Transcriptome" (EMR/2014/000444) from the Science and Engineering Research Board (SERB), Department of Science and Technology, Government of India. T.S.K.P. is also supported by the DBT Program Support grant on "Development of infrastructure and a computational framework for analysis of proteomic data" (BT/01/COE/08/05). We also thank Infosys Foundation for financial support to IOB. A.P. and P.S. were funded by a pilot grant from the Johns Hopkins Malaria Research Institute. This paper bears the NIMR publication screening committee approval No. 009/2015. H.G. is a Wellcome Trust-DBT India Alliance Early Career Fellow. We thank the Council of Scientific and Industrial Research, Department of Biotechnology, University Grants Commission, Indian Council of Medical Research and Department of Science and Technology, Government of India for research fellowships to M.K., S.K.S., G.D., R.S.N., S.M.P., A.K. M. (IOB), S.S.M., M.K.G., S.B.D., D.S.K., P.R., N.S., S.D.Y., K.K.D., R.R., A.A.K., A.R., G.J.S., S.C., and R.V. M.D. is funded by the Faculty Improvement Program of Siddaganga Institute of Technology, Tumkur.

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. ; National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services [U19AI110818]; USDA [2017-05741]; NIH Intramural Research Program; National Library of Medicine; National Human Genome Research Institute; NSF [PHY-1427654]; NIH [R01AI101112, R35GM118336, R21AI121853, R01AI123338, T32GM007739, NIH/NCATS UL1TR000043, DP2OD008540, U01AI088647, 1R01AI121211, D43TW001130-08, U01HL130010, UM1HG009375, 5K22AI113060, 1R21AI123937, R00DC012069]; Defence Advanced Research Project Agency [HR0011-17-2-0047]; Jane Coffin Childs Memorial Fund; Center for Theoretical Biological Physics postdoctoral fellowship; Robertson Foundation; McNair Foundation; Welch Foundation [Q-1866]; French Government's Investissement d'Avenir program, Laboratoire d'Excellence Integrative Biology of Emerging Infectious Diseases [ANR-10-LABX-62-IBEID]; Agence Nationale de la Recherche [ANR-17-ERC2-0016-01]; European Union [734584]; Pew and Searle Scholars Programs; Klingenstein-Simons Fellowship in the Neurosciences; Verily Life Sciences ; We thank R. Andino; S. Emrich and D. Lawson (Vectorbase); A. A. James, M. Kunitomi, C. Nusbaum, D. Severson, N. Whiteman; T. Dickinson, M. Hartley and B. Rice (Dovetail Genomics) for early participation in the AGWG; C. Bargmann, D. Botstein, E. Jarvis and E. Lander for encouragement and facilitation. N. Keivanfar, D. Jaffe and D. M. Church (10X Genomics) prepared DNA for structural-variant analysis. We thank A. Harmon of the New York Times and acknowledge generous pro bono data and analysis from our corporate collaborators. This research was supported in part by federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under grant number U19AI110818 to the Broad Institute (S.N.R. and D.E.N.); USDA 2017-05741 (E.L.A.); NSF PHY-1427654 Center for Theoretical Biological Physics (E.L.A.); NIH Intramural Research Program, National Library of Medicine and National Human Genome Research Institute (A.M.P. and S.K.) and the following extramural NIH grants: R01AI101112 (J.R.P.), R35GM118336 (R.S.M. and W.J.G.), R21AI121853 (M.V.S., I.V.S. and A. S.), R01AI123338 (Z.T.), T32GM007739 (M.H.), NIH/NCATS UL1TR000043 (Rockefeller University), DP2OD008540 (E.L.A.), U01AI088647, 1R01AI121211 (W.C.B. IV), Fogarty Training Grant D43TW001130-08, U01HL130010 (E.L.A.), UM1HG009375 (E.L.A), 5K22AI113060 (O.S.A.), 1R21AI123937 (O.S.A.), and R00DC012069 (C.S.M.); Defence Advanced Research Project Agency: HR0011-17-2-0047 (O.S.A.). Other support was provided by Jane Coffin Childs Memorial Fund (B.J.M.), Center for Theoretical Biological Physics postdoctoral fellowship (O.D.), Robertson Foundation (L.Z.), and McNair & Welch (Q-1866) Foundations (E.L.A.), French Government's Investissement d'Avenir program, Laboratoire d'Excellence Integrative Biology of Emerging Infectious Diseases (grant ANR-10-LABX-62-IBEID to L.L.), Agence Nationale de la Recherche grant ANR-17-ERC2-0016-01 (L.L.), European Union's Horizon 2020 research and innovation program under ZikaPLAN grant agreement no. 734584 (L.L.), Pew and Searle Scholars Programs (C.S.M.), Klingenstein-Simons Fellowship in the Neurosciences (C.S.M.). A.M.W., B.J.W., J.E.C. and S.N.M. were supported by Verily Life Sciences. L.B.V. is an investigator of the Howard Hughes Medical Institute.