Temporal activation of the sea urchin late H1 gene requires stage- specific phosphorylation of the embryonic transcription factor SSAP

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Abstract

Stage-specific activator protein (SSAP) is a 41-kDa polypeptide that hinds to embryonic enhancer elements of the sea urchin late H1 gene. These enhancer elements mediate the transcriptional activation of the late H1 gene in a temporally specific manner at the mid-blastula stage of embryogenesis. Although SSAP can transactivate the late HI gene only at late stages of the development, it resides in the sea urchin nucleus and maintains DNA binding activity throughout early embryogenesis. In addition, it has been shown that SSAP undergoes a conversion from a 41-kDa monomer to a ~80- to 100-kDa dimer when the late H1 gene is activated. We have demonstrated that SSAP is differentially phosphorylated during embryogenesis. Serine 87, a cyclic AMP- dependent protein kinase consensus site located in the N-terminal DNA binding domain, is constitutively phosphorylated. At the mid-blastula stage of embryogenesis, temporally correlated with SSAP dimer formation and late H1 gene activation, a threonine residue in the C-terminal transactivation domain is phosphorylated. This phosphorylation can be catalyzed by a break-ended double-stranded DNA-activated protein kinase activity from the sea urchin nucleus in vitro. Microinjection of synthetic SSAP mRNAs encoding either serine or threonine phosphorylation mutants results in the failure to transactivate reporter genes that contain the enhancer element, suggesting that both serine and threonine phosphorylation of SSAP are required for the activation of the late H1 gene. Furthermore, SSAP can undergo blastula- stage-specific homodimerization through its GQ-rich transactivation domain. The late-specific threonine phosphorylation in this domain is essential for the dimer assembly. These observations indicate that temporally regulated SSAP activation is promoted by threonine phosphorylation on its transactivation domain, which triggers the formation of a transcriptionally active SSAP homodimer.

Original languageEnglish (US)
Pages (from-to)3684-3695
Number of pages12
JournalMolecular and Cellular Biology
Volume19
Issue number5
StatePublished - May 1999

Fingerprint

Sea Urchins
Transcription Factors
Phosphorylation
Threonine
Transcriptional Activation
Genes
Proteins
Blastula
Embryonic Development
Serine
DNA-Activated Protein Kinase
DNA
Microinjections
Cyclic AMP-Dependent Protein Kinases
Reporter Genes
Messenger RNA
Peptides

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

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title = "Temporal activation of the sea urchin late H1 gene requires stage- specific phosphorylation of the embryonic transcription factor SSAP",
abstract = "Stage-specific activator protein (SSAP) is a 41-kDa polypeptide that hinds to embryonic enhancer elements of the sea urchin late H1 gene. These enhancer elements mediate the transcriptional activation of the late H1 gene in a temporally specific manner at the mid-blastula stage of embryogenesis. Although SSAP can transactivate the late HI gene only at late stages of the development, it resides in the sea urchin nucleus and maintains DNA binding activity throughout early embryogenesis. In addition, it has been shown that SSAP undergoes a conversion from a 41-kDa monomer to a ~80- to 100-kDa dimer when the late H1 gene is activated. We have demonstrated that SSAP is differentially phosphorylated during embryogenesis. Serine 87, a cyclic AMP- dependent protein kinase consensus site located in the N-terminal DNA binding domain, is constitutively phosphorylated. At the mid-blastula stage of embryogenesis, temporally correlated with SSAP dimer formation and late H1 gene activation, a threonine residue in the C-terminal transactivation domain is phosphorylated. This phosphorylation can be catalyzed by a break-ended double-stranded DNA-activated protein kinase activity from the sea urchin nucleus in vitro. Microinjection of synthetic SSAP mRNAs encoding either serine or threonine phosphorylation mutants results in the failure to transactivate reporter genes that contain the enhancer element, suggesting that both serine and threonine phosphorylation of SSAP are required for the activation of the late H1 gene. Furthermore, SSAP can undergo blastula- stage-specific homodimerization through its GQ-rich transactivation domain. The late-specific threonine phosphorylation in this domain is essential for the dimer assembly. These observations indicate that temporally regulated SSAP activation is promoted by threonine phosphorylation on its transactivation domain, which triggers the formation of a transcriptionally active SSAP homodimer.",
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T1 - Temporal activation of the sea urchin late H1 gene requires stage- specific phosphorylation of the embryonic transcription factor SSAP

AU - Zhe, L. I.

AU - Childs, Geoffrey J.

PY - 1999/5

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N2 - Stage-specific activator protein (SSAP) is a 41-kDa polypeptide that hinds to embryonic enhancer elements of the sea urchin late H1 gene. These enhancer elements mediate the transcriptional activation of the late H1 gene in a temporally specific manner at the mid-blastula stage of embryogenesis. Although SSAP can transactivate the late HI gene only at late stages of the development, it resides in the sea urchin nucleus and maintains DNA binding activity throughout early embryogenesis. In addition, it has been shown that SSAP undergoes a conversion from a 41-kDa monomer to a ~80- to 100-kDa dimer when the late H1 gene is activated. We have demonstrated that SSAP is differentially phosphorylated during embryogenesis. Serine 87, a cyclic AMP- dependent protein kinase consensus site located in the N-terminal DNA binding domain, is constitutively phosphorylated. At the mid-blastula stage of embryogenesis, temporally correlated with SSAP dimer formation and late H1 gene activation, a threonine residue in the C-terminal transactivation domain is phosphorylated. This phosphorylation can be catalyzed by a break-ended double-stranded DNA-activated protein kinase activity from the sea urchin nucleus in vitro. Microinjection of synthetic SSAP mRNAs encoding either serine or threonine phosphorylation mutants results in the failure to transactivate reporter genes that contain the enhancer element, suggesting that both serine and threonine phosphorylation of SSAP are required for the activation of the late H1 gene. Furthermore, SSAP can undergo blastula- stage-specific homodimerization through its GQ-rich transactivation domain. The late-specific threonine phosphorylation in this domain is essential for the dimer assembly. These observations indicate that temporally regulated SSAP activation is promoted by threonine phosphorylation on its transactivation domain, which triggers the formation of a transcriptionally active SSAP homodimer.

AB - Stage-specific activator protein (SSAP) is a 41-kDa polypeptide that hinds to embryonic enhancer elements of the sea urchin late H1 gene. These enhancer elements mediate the transcriptional activation of the late H1 gene in a temporally specific manner at the mid-blastula stage of embryogenesis. Although SSAP can transactivate the late HI gene only at late stages of the development, it resides in the sea urchin nucleus and maintains DNA binding activity throughout early embryogenesis. In addition, it has been shown that SSAP undergoes a conversion from a 41-kDa monomer to a ~80- to 100-kDa dimer when the late H1 gene is activated. We have demonstrated that SSAP is differentially phosphorylated during embryogenesis. Serine 87, a cyclic AMP- dependent protein kinase consensus site located in the N-terminal DNA binding domain, is constitutively phosphorylated. At the mid-blastula stage of embryogenesis, temporally correlated with SSAP dimer formation and late H1 gene activation, a threonine residue in the C-terminal transactivation domain is phosphorylated. This phosphorylation can be catalyzed by a break-ended double-stranded DNA-activated protein kinase activity from the sea urchin nucleus in vitro. Microinjection of synthetic SSAP mRNAs encoding either serine or threonine phosphorylation mutants results in the failure to transactivate reporter genes that contain the enhancer element, suggesting that both serine and threonine phosphorylation of SSAP are required for the activation of the late H1 gene. Furthermore, SSAP can undergo blastula- stage-specific homodimerization through its GQ-rich transactivation domain. The late-specific threonine phosphorylation in this domain is essential for the dimer assembly. These observations indicate that temporally regulated SSAP activation is promoted by threonine phosphorylation on its transactivation domain, which triggers the formation of a transcriptionally active SSAP homodimer.

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