The CyIIIa actin gene of Strongylocentrotus purpuratus is
transcribed exclusively in the embryonic aboral ectoderm,
under the control of 2.3 kb cis-regulatory domain that
contains a proximal module that controls expression in
early embryogenesis, and a middle module that controls
expression in later embryogenesis. Previous studies demonstrated that the SpRunt-1 target site within the middle
module is required for the sharp increase in CyIIIa transcription which accompanies differentiation of the aboral ectoderm, and that a negative regulatory region near the SpRunt-1 target site is required to prevent ectopic transcription in the oral ectoderm and skeletogenic mesenchyme. This negative regulatory region contains a
consensus binding site for the myb family of transcription
factors. In vitro DNA-binding experiments reveal that a
protein in blastula-stage nuclei interacts specifically with
the myb target site. Gene transfer experiments utilizing
CyIIIa reporter constructs containing oligonucleotide substitutions indicate that this site is both necessary and sufficient to prevent ectopic expression of CyIIIa. Synthetic oligonucleotides containing the myb target site were used to purify a protein from sea urchin embryo nuclear extracts by affinity chromatography. This protein is immunoprecipitated by antibodies specific to the evolutionarily conserved myb domain, and amino acid sequences obtained from the purified protein were found to be identical to sequences within the myb domain. Sequence information was used to obtain cDNA clones of SpMyb, the S. purpuratus member of the myb family of transcription factors. Through interactions within the middle module, SpMyb functions to repress activation of CyIIIa in the oral
ectoderm and skeletogenic mesenchyme