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Developmental changes in phosphorylation of the transcription factor CREB in the embryonic murine palate
1Department of Pathology, Anatomy, and Cell Biology, Jefferson Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
Insights
Cyclic AMP-response element binding protein (CREB) is present in developing palates. Its phosphorylation increases during palate development, suggesting it regulates gene expression via phosphorylation, not expression levels.
Area of Science:
- Developmental Biology
- Molecular Biology
- Cellular Biology
Background:
- Cyclic AMP (cAMP) and cAMP-dependent protein kinase (PKA) are crucial for mammalian palate development.
- CREB is a transcription factor regulated by PKA and controls cAMP-inducible gene expression.
Purpose of the Study:
- To investigate the expression, distribution, and phosphorylation of CREB in the developing murine palate.
- To determine if CREB levels or phosphorylation regulate its activity during palate development.
Main Methods:
- Immunofluorescent staining using anti-CREB antibodies to determine CREB localization.
- Western blot analysis using phospho-specific antibodies to quantify CREB phosphorylation at serine 133.
- Analysis of staged palatal shelves during development.
Main Results:
- CREB is constitutively expressed and ubiquitously distributed in palatal tissue, localized to cell nuclei.
- Phosphorylation of CREB at serine 133 significantly increases during palate development.
- Increased CREB phosphorylation correlates with cAMP-regulated gene expression patterns.
Conclusions:
- CREB is present and active throughout palate development.
- CREB activity is regulated primarily by protein phosphorylation, not changes in protein expression levels.
- Phosphorylation of CREB is a key mechanism in regulating gene expression during mammalian palate development.
Abstract:
Cyclic AMP, via activation of cAMP-dependent protein kinase (PKA) and subsequent protein phosphorylation, regulates a number of cellular and tissue responses that are critical to normal development of the mammalian palate. The present study examines the expression, distribution, and phosphorylation in the developing murine palate of a substrate for PKA known as the cAMP-response element binding protein (CREB). This 43 x 10(3) M(r) protein functions as a regulator of cAMP-inducible gene expression. CREB is expressed constituitively throughout the palatal morphogenetic period and is ubiquitously distributed throughout palatal tissue. Immunofluorescent staining of palatal cells and tissues with an anti-CREB antibody revealed CREB to be localized to cell nuclei. Western blot analysis of extracts of staged palatal shelves with an antibody specific for phospho-ser 133-CREB demonstrated a steady increase in CREB phosphorylation at this residue during palate development. These observations show a temporal correlation with expression levels of cAMP-regulated genes in palate cells. The data indicate that CREB activity in the developing palate is most likely to be regulated at the level of protein phosphorylation as opposed to changes in levels of CREB protein expression.