Analysis of the transcriptomes downstream of Eyeless and the Hedgehog, Decapentaplegic and Notch signaling pathways in Drosophila melanogaster

PLoS One. 2012;7(8):e44583. doi: 10.1371/journal.pone.0044583. Epub 2012 Aug 31.

Abstract

Tissue-specific transcription factors are thought to cooperate with signaling pathways to promote patterned tissue specification, in part by co-regulating transcription. The Drosophila melanogaster Pax6 homolog Eyeless forms a complex, incompletely understood regulatory network with the Hedgehog, Decapentaplegic and Notch signaling pathways to control eye-specific gene expression. We report a combinatorial approach, including mRNAseq and microarray analyses, to identify targets co-regulated by Eyeless and Hedgehog, Decapentaplegic or Notch. Multiple analyses suggest that the transcriptomes resulting from co-misexpression of Eyeless+signaling factors provide a more complete picture of eye development compared to previous efforts involving Eyeless alone: (1) Principal components analysis and two-way hierarchical clustering revealed that the Eyeless+signaling factor transcriptomes are closer to the eye control transcriptome than when Eyeless is misexpressed alone; (2) more genes are upregulated at least three-fold in response to Eyeless+signaling factors compared to Eyeless alone; (3) based on gene ontology analysis, the genes upregulated in response to Eyeless+signaling factors had a greater diversity of functions compared to Eyeless alone. Through a secondary screen that utilized RNA interference, we show that the predicted gene CG4721 has a role in eye development. CG4721 encodes a neprilysin family metalloprotease that is highly up-regulated in response to Eyeless+Notch, confirming the validity of our approach. Given the similarity between D. melanogaster and vertebrate eye development, the large number of novel genes identified as potential targets of Ey+signaling factors will provide novel insights to our understanding of eye development in D. melanogaster and humans.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Arthropod Antennae / metabolism
  • DNA-Binding Proteins / metabolism
  • Down-Regulation / genetics
  • Drosophila Proteins / chemistry
  • Drosophila Proteins / genetics*
  • Drosophila Proteins / metabolism*
  • Drosophila melanogaster / cytology
  • Drosophila melanogaster / genetics*
  • Drosophila melanogaster / growth & development
  • Eye / cytology
  • Eye / metabolism
  • Gene Expression Profiling*
  • Gene Expression Regulation, Developmental
  • Genome, Insect / genetics
  • Hedgehog Proteins / metabolism
  • Molecular Sequence Data
  • Neprilysin / chemistry
  • Neprilysin / metabolism
  • Organ Specificity / genetics
  • Photoreceptor Cells, Invertebrate / metabolism
  • RNA Interference
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Receptors, Notch / metabolism
  • Signal Transduction / genetics*
  • Transcription, Genetic
  • Transcriptome / genetics*
  • Up-Regulation / genetics
  • Wings, Animal / cytology
  • Wings, Animal / metabolism

Substances

  • DNA-Binding Proteins
  • Drosophila Proteins
  • Hedgehog Proteins
  • N protein, Drosophila
  • RNA, Messenger
  • Receptors, Notch
  • dpp protein, Drosophila
  • ey protein, Drosophila
  • hh protein, Drosophila
  • Neprilysin