Cis-regulatory analysis of the sea urchin pigment cell gene polyketide synthase.

The Strongylocentrotus purpuratus polyketide synthase gene (SpPks) encodes an enzyme required for the biosynthesis of the larval pigment echinochrome. SpPks is expressed exclusively in pigment cells and their precursors starting at blastula stage. The 7th-9th cleavage Delta-Notch signaling, required for pigment cell development, positively regulates SpPks. In previous studies, the transcription factors glial cell missing (SpGcm), SpGatae and kruppel-like (SpKrl/z13) have been shown to positively regulate SpPks. To uncover the structure of the Gene Regulatory Network (GRN) regulating the specification and differentiation processes of pigment cells, we experimentally analyzed the putative SpPks cis-regulatory region. We established that the -1.5kb region is sufficient to recapitulate the correct spatial and temporal expression of SpPks. Predicted DNA-binding sites for SpGcm, SpGataE and SpKrl are located within this region. The mutagenesis of these DNA-binding sites indicated that SpGcm, SpGataE and SpKrl are direct positive regulators of SpPks. These results demonstrate that the sea urchin GRN for pigment cell development is quite shallow, which is typical of type I embryo development.