title: High-throughput cis-regulatory element discovery in the vector mosquito Aedes aegypti.
abstract: Despite substantial progress in mosquito genomic and genetic research, few cis-regulatory elements (CREs), DNA sequences that control gene expression, have been identified in mosquitoes or other non-model insects. Formaldehyde-assisted isolation of regulatory elements paired with DNA sequencing, FAIRE-seq, is emerging as a powerful new high-throughput tool for global CRE discovery. FAIRE results in the preferential recovery of open chromatin DNA fragments that are not bound by nucleosomes, an evolutionarily conserved indicator of regulatory activity, which are then sequenced. Despite the power of the approach, FAIRE-seq has not yet been applied to the study of non-model insects. In this investigation, we utilized FAIRE-seq to profile open chromatin and identify likely regulatory elements throughout the genome of the human disease vector mosquito Aedes aegypti. We then assessed genetic variation in the regulatory elements of dengue virus susceptible (Moyo-S) and refractory (Moyo-R) mosquito strains.
  
  
  
  Analysis of sequence data obtained through next generation sequencing of FAIRE DNA isolated from A. aegypti embryos revealed >121,000 FAIRE peaks (FPs), many of which clustered in the 1 kb 5' upstream flanking regions of genes known to be expressed at this stage. As expected, known transcription factor consensus binding sites were enriched in the FPs, and of these FoxA1, Hunchback, Gfi, Klf4, MYB/ph3 and Sox9 are most predominant. All of the elements tested in vivo were confirmed to drive gene expression in transgenic Drosophila reporter assays. Of the >13,000 single nucleotide polymorphisms (SNPs) recently identified in dengue virus-susceptible and refractory mosquito strains, 3365 were found to map to FPs.
  
  
  
  FAIRE-seq analysis of open chromatin in A. aegypti permitted genome-wide discovery of CREs. The results of this investigation indicate that FAIRE-seq is a powerful tool for identification of regulatory DNA in the genomes of non-model organisms, including human disease vector mosquitoes.
authors.affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN, 46556, USA.|Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA.
authors.forename: Susanta K
authors.initials: SK
authors.lastname: Behura
authors.affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN, 46556, USA.|Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA.|Department of Medical and Molecular Genetics, Indiana University School of Medicine, 1234 Notre Dame Ave., South Bend, IN, 46617, USA.
authors.forename: Joseph
authors.initials: J
authors.lastname: Sarro
authors.affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN, 46556, USA.|Department of Medical and Molecular Genetics, Indiana University School of Medicine, 1234 Notre Dame Ave., South Bend, IN, 46617, USA.
authors.forename: Ping
authors.initials: P
authors.lastname: Li
authors.affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN, 46556, USA.|Department of Medical and Molecular Genetics, Indiana University School of Medicine, 1234 Notre Dame Ave., South Bend, IN, 46617, USA.
authors.forename: Keshava
authors.initials: K
authors.lastname: Mysore
authors.affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN, 46556, USA.|Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA.|Department of Medical and Molecular Genetics, Indiana University School of Medicine, 1234 Notre Dame Ave., South Bend, IN, 46617, USA.
authors.forename: David W
authors.initials: DW
authors.lastname: Severson
authors.affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN, 46556, USA. semrich@nd.edu.|Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA. semrich@nd.edu.|Department of Computer Science and Engineering, University of Notre Dame, Notre Dame, IN, 46556, USA. semrich@nd.edu.
authors.forename: Scott J
authors.initials: SJ
authors.lastname: Emrich
authors.affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN, 46556, USA. mscheel@nd.edu.|Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA. mscheel@nd.edu.|Department of Medical and Molecular Genetics, Indiana University School of Medicine, 1234 Notre Dame Ave., South Bend, IN, 46617, USA. mscheel@nd.edu.
authors.forename: Molly
authors.initials: M
authors.lastname: Duman-Scheel
mesh_terms: D000330:Aedes / Q000235:genetics* / Q000821:virology; D000818:Animals; D001665:Binding Sites; D002874:Chromosome Mapping; D015870:Gene Expression; D017930:Genes, Reporter; D014644:Genetic Variation; D049750:Genome, Insect; D023281:Genomics / Q000379:methods; D059014:High-Throughput Nucleotide Sequencing; D007303:Insect Vectors / Q000235:genetics / Q000821:virology; D000090062:Kruppel-Like Factor 4; D012045:Regulatory Sequences, Nucleic Acid; D014157:Transcription Factors / Q000378:metabolism; D020506:Untranslated Regions
chemical_list: C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D014157:Transcription Factors; D020506:Untranslated Regions
keywords: Dengue virus; Drosophila; FAIRE-seq; Genome; Next generation sequencing; Zika