Functional dissection of eyes absent reveals new modes of regulation within the retinal determination gene network.
The retinal determination (RD) gene network encodes a group of transcription factors and cofactors necessary for eye development. Transcriptional and posttranslational regulation of RD family members is achieved through interactions within the network and with extracellular signaling pathways, including epidermal growth factor receptor/RAS/mitogen-activated protein kinase (MAPK), transforming growth factor beta/DPP, Wingless, Hedgehog, and Notch. Here we present the results of structure-function analyses that reveal novel aspects of Eyes absent (EYA) function and regulation. We find that the conserved C-terminal EYA domain negatively regulates EYA transactivation potential, and that GROUCHO-SINE OCULIS (SO) interactions provide another mechanism for negative regulation of EYA-SO target genes. We have mapped the transactivation potential of EYA to an internal proline-, serine-, and threonine-rich region that includes the EYA domain 2 (ED2) and two MAPK phosphorylation consensus sites and demonstrate that activation of the RAS/MAPK pathway potentiates transcriptional output of EYA and the EYA-SO complex in certain contexts. Drosophila S2 cell two-hybrid assays were used to describe a novel homotypic interaction that is mediated by EYA's N terminus. Our data suggest that EYA requires homo- and heterotypic interactions and RAS/MAPK signaling responsiveness to ensure context-appropriate RD gene network activity.
Silver
Serena J
SJ
Whitehead Institute for Biomedical Research. MIT Department of Biology, Cambridge, Massachusetts 02142, USA.
Davies
Erin L
EL
Doyon
Laura
L
Rebay
Ilaria
I
D000818:Animals; D030801:Animals, Genetically Modified; D051792:Basic Helix-Loop-Helix Proteins; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004330:Drosophila / Q000235:genetics; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism*; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009154:Mutation; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D012045:Regulatory Sequences, Nucleic Acid; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D012160:Retina / Q000502:physiology*; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D020798:Two-Hybrid System Techniques; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism
D051792:Basic Helix-Loop-Helix Proteins; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D005136:Eye Proteins; D018398:Homeodomain Proteins; D020928:Mitogen-Activated Protein Kinases; D012097:Repressor Proteins; D014157:Transcription Factors; D018631:ras Proteins; C087345:So protein, Drosophila; C092694:eya protein, Drosophila; C091023:gro protein, Drosophila