A global survey identifies novel upstream components of the Ath5 neurogenic network.
BACKGROUND
Investigating the architecture of gene regulatory networks (GRNs) is essential to decipher the logic of developmental programs during embryogenesis. In this study we present an upstream survey approach, termed trans-regulation screen, to comprehensively identify the regulatory input converging on endogenous regulatory sequences.


RESULTS
Our dual luciferase-based screen queries transcriptome-scale collections of cDNAs. Using this approach we study the regulation of Ath5, the central node in the GRN controlling retinal ganglion cell (RGC) specification in vertebrates. The Ath5 promoter integrates the input of upstream regulators to enable the transient activation of the gene, which is an essential step for RGC differentiation. We efficiently identified potential Ath5 regulators that were further filtered for true positives by an in situ hybridization screen. Their regulatory activity was validated in vivo by functional assays in medakafish embryos.


CONCLUSIONS
Our analysis establishes functional groups of genes controlling different regulatory phases, including the onset of Ath5 expression at cell-cycle exit and its down-regulation prior to terminal RGC differentiation. These results extent the current model of the GRN controlling retinal neurogenesis in vertebrates.
Souren
Marcel
M
Developmental Biology Unit, EMBL-Heidelberg, Meyerhofstrasse, Heidelberg, 69117, Germany. Marcel.Souren@embl.de
Martinez-Morales
Juan Ramon
JR
Makri
Panagiota
P
Wittbrodt
Beate
B
Wittbrodt
Joachim
J
D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D005123:Eye / Q000166:cytology / Q000294:innervation / Q000378:metabolism; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism; D029941:Fish Proteins / Q000235:genetics* / Q000378:metabolism; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D053263:Gene Regulatory Networks; D017404:In Situ Hybridization, Fluorescence; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D009990:Oryzias / Q000196:embryology / Q000235:genetics / Q000254:growth & development; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012165:Retinal Ganglion Cells / Q000166:cytology / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection
D051792:Basic Helix-Loop-Helix Proteins; D005136:Eye Proteins; D029941:Fish Proteins; D008156:Luciferases; D009419:Nerve Tissue Proteins; D011993:Recombinant Fusion Proteins