Strongylocentrotus purpuratus transcription factor GATA-E binds to and represses transcription at an Otx-Goosecoid cis-regulatory element within the aboral ectoderm-specific spec2a enhancer.
During Strongylocentrotus purpuratus embryogenesis, aboral ectoderm-specific expression of spec2a relies on an upstream enhancer that confers its spatial specificity largely through repression. The purpose of this study was to determine how spec2a expression is repressed in endoderm and oral ectoderm territories. A 78-base pair DNA sequence within the enhancer contains five tightly spaced cis-regulatory elements including proximal (TAATCT) and distal (TAATCC) elements that bind to both SpOtx, a broadly distributed transcriptional activator, and SpGoosecoid (SpGsc), an oral ectoderm-restricted transcriptional repressor. We show here that these two seemingly redundant Otx/Gsc elements have distinct functions. The proximal element bound to SpGATA-E, an endomesoderm-specific transcription factor. Treatment with SpGATA-E and SpGsc morpholino antisense oligonucleotides (MASOs) resulted in enhanced transcriptional activity from the proximal element, suggesting that both factors functioned as repressors at this site. SpGATA-E MASO-treated embryos failed to express ectoderm markers, indicating a role for SpGATA-E in ectoderm differentiation. The spec2a proximal element was distinct from the corresponding element in the related spec1 enhancer, and swaps between spec1 and spec2a cis-regulatory elements indicated, that for optimal repression, the proximal element had to interact with a nearby CCAAT-binding factor element. Our results show that the recently evolved proximal element contributes to the repression of spec2a in endomesoderm and oral ectoderm territories.
Kiyama
Takae
T
Department of Biochemistry and Molecular Biology, The University of Texas M. D. Anderson Cancer Center, Houston, 77030, USA.
Zhang
Ning
N
Dayal
Sandeep
S
Yun Lee
Pei
P
Liang
Shuguang
S
Villinski
Jeffrey T
JT
Klein
William H
WH
D000595:Amino Acid Sequence; D000818:Animals; D036703:Blastula / Q000378:metabolism; D023081:CCAAT-Binding Factor / Q000378:metabolism / Q000502:physiology; D002454:Cell Differentiation; D002467:Cell Nucleus / Q000378:metabolism; D015536:Down-Regulation; D004475:Ectoderm / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D005982:Glutathione Transferase / Q000378:metabolism; D051956:Goosecoid Protein; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D015151:Immunoblotting; D008156:Luciferases / Q000378:metabolism; D008648:Mesoderm / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000737:chemistry; D051857:Otx Transcription Factors; D010957:Plasmids / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D012097:Repressor Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D047270:Strongylocentrotus purpuratus / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D016335:Zinc Fingers
D023081:CCAAT-Binding Factor; D051956:Goosecoid Protein; D018398:Homeodomain Proteins; D016376:Oligonucleotides, Antisense; D051857:Otx Transcription Factors; D012333:RNA, Messenger; D011994:Recombinant Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D008156:Luciferases; D005982:Glutathione Transferase