scholarly journals A Positive Regulatory Domain in CCAAT/Enhancer Binding Protein β (C/EBPβ) Is Required for the Glucocorticoid-Mediated Displacement of Histone Deacetylase 1 (HDAC1) from the C/ebpα Promoter and Maximum Adipogenesis

Endocrinology ◽  
2013 ◽  
Vol 154 (4) ◽  
pp. 1454-1464 ◽  
Author(s):  
Houssein-Salem Abdou ◽  
Ella Atlas ◽  
Robert J. G. Haché
Keyword(s):  
Histone Deacetylase ◽  
Transcriptional Activity ◽  
Cellular Proliferation ◽  
Binding Protein ◽  
Regulatory Domain ◽  
Glucocorticoid Treatment ◽  
Histone Deacetylase 1 ◽  
The Metabolic Syndrome ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein

Abstract Glucocorticoids promote adipogenesis and contribute to the metabolic syndrome through a number of mechanisms. One of the effectors of glucocorticoid action is the CCAAT/enhancer binding protein β (C/EBPβ). C/EBPβ is a basic leucine-zipper transcription factor involved in diverse processes including differentiation, cellular proliferation, and inflammation. C/EBPβ transcriptional activity is regulated, in part, by its acetylation profile resulting from its dynamic interaction with either acetylases general control nonrepressed protein 5/p300/CBP associated factor (GCN5/PCAF) or deacetylase complexes (mSin3A/histone deacetylase 1 [HDAC1]). Glucocorticoid treatment of preadipocytes promotes C/EBPβ acetylation, leading to mSin3A/HDAC1 dissociation from C/EBPβ and resulting in C/ebpα promoter activation at the onset of adipogenesis, thus increasing the differentiation rate. We recently showed that the regulatory domain 1 (RD1) of C/EBPβ contains four residues (153–156) required for its interaction with HDAC1, therefore supporting RD1 proposed inhibitory role. In an attempt to further elucidate the intrinsic regulatory property of RD1, we sought to characterize the regulatory potential of the N terminus region of RD1 (residues 141–149). In this study, we show that C/EBPβΔ141–149 transcriptional activity was compromised on the C/ebpα, but not on the Pparγ, promoter. Additionally, the ability of C/EBPβΔ141–149 to induce adipogenesis in NIH 3T3 cells was compromised when compared with C/EBPβwt owing to a delayed expression of C/ebpα at the onset of differentiation. Furthermore, the data suggest that the reduced expression of C/ebpα in cells expressing C/EBPβΔ141–149 was due to a persistent recruitment of HDAC1 to the C/ebpα promoter after glucocorticoid treatment. Together, these results suggest that amino acids 141–149 of C/EBPβ act as a positive regulatory domain required for maximum transcriptional activity.

Post-Translational Modification and Differential Protein Binding Partners of CCAAT Enhancer Binding Protein Alpha (C/EBPα) p42 Versus p30 May Contribute to Aberrant C/EBPα Activity in Acute Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3577-3577
Author(s):  
Matthew Silver ◽  
Nirmalee Abayasekara ◽  
Dylan Perry ◽  
Hong Sun ◽  
Nancy Berliner ◽  
...  
Keyword(s):  
Protein Binding ◽  
Transcriptional Activity ◽  
Binding Protein ◽  
Myeloid Cells ◽  
Leukemic Cells ◽  
Myeloid Differentiation ◽  
Differential Protein ◽  
Binding Partners ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein

Abstract CCAAT enhancer binding protein alpha (C/EBPα) is the founding member of a family of basic region/leucine zipper (bzip) transcription factors and has been shown to be a master regulator of granulopoiesis It is expressed at high levels throughout myeloid differentiation and has been shown to bind to the promoters of multiple myeloid- specific gene promoters at different stages of myeloid maturation. Profound hematopoietic abnormalities have been reported for mice nullizygous for including a selective early block in the differentiation of C/EBPα, granulocytes. Mutations in C/EBPα have been demonstrated in a subset of patients with AML presenting with a normal karyotype. These mutations can result in the expression of a 30kD dominant negative C/EBPα isoform which contributes to loss of C/EBPα function. We have sought to understand the molecular basis for this observation. We and others have demonstrated that C/EBPα is post-translationally modified by small ubiquitin-related modifier (SUMO) at a lysine residue (K159) that lies within a region of the C/EBPα protein that can negatively affect transcriptional activity. We have demonstrated that the levels of sumoylated p42C/EBPα decrease upon normal neutrophil maturation and that transactivation of the myeloid-specific lactoferrin (LF) promoter reporter is significantly enhanced by a p42 sumoylation mutant of C/EBPα (K159A). Additionally, in oligonucleotide pull down assays, we show that sumoylated p42C/EBPα binds to the C/EBP site in the LF promoter in immature myeloid cells (which do not express LF) while loss binding and LF of sumoylation correlates with loss of p42C/EBPα expression in more mature cells. Based on these observations we is associated with the negative conclude that sumoylated p42C/EBPα regulation of LF in early myeloid cells. We further demonstrate that sumoylated p42C/EBPα remains bound to the LF promoter following ATRA induction of the leukemic NB4 cells, which do not express LF despite induction of morphologic maturation. Based on these observations we conclude that during normal myeloid differentiation, sumoylated p42C/EBPα is associated with the negative regulation of LF in early myeloid cells, and that LF expression upon maturation is associated with loss of binding of sumoylated p42 C/EBPα In leukemic cells induced toward mature neutrophils, sumoylated p42C/EBPα remains bound to the LF promoter, contributing to the lack of expression of LF in these cells. We show in addition, that p30 C/EBPα can also be sumoylated. In transactivation assays, however, sumoylated p42C/EBPα suppresses LF promoter activity more efficiently than p30C/EBPα in 293 cells. In order to identify differential protein binding partners of p30 and p42C/EBPα that could account for the differential transcriptional activity of the two isoforms, we have used a one step purification method that allows isolation of biotinylated C/EBPα p30 and p42- containing complexes using magnetic-streptavidin beads. The K562 myelomonocytic cell line stably expressing a biotin ligase (BirA) plasmid was transfected with p30C/EBPα or p42C/EBPα each containing a 23 amino acid tag at the N-terminus that allows for in vivo biotinylation. Proteins complexed with the two C/EBP isofoms have been isolated and are currently being identified by LC- MS MS analysis. Their differential association with the two isofoms of C/EBPα will be confimed by coimmunoprecipitation assays in normal myeloid and in leukemic cells. The identification of differentially bound proteins to p30 and p42 C/EBPα may identify molecular targets for future drug development.

scholarly journals Cyclin C regulates adipogenesis by stimulating transcriptional activity of CCAAT/enhancer-binding protein α

2017 ◽  
Vol 292 (21) ◽  
pp. 8918-8932 ◽  
Author(s):  
Ziyi Song ◽  
Alus M. Xiaoli ◽  
Quanwei Zhang ◽  
Yi Zhang ◽  
Ellen S. T. Yang ◽  
...  
Keyword(s):  
Transcriptional Activity ◽  
Binding Protein ◽  
Enhancer Binding Protein ◽  
Cyclin C ◽  
Ccaat Enhancer Binding Protein

scholarly journals Repression of Transcriptional Activity of C/EBPα by E2F-Dimerization Partner Complexes

2010 ◽  
Vol 30 (9) ◽  
pp. 2293-2304 ◽  
Author(s):  
Katrin Zaragoza ◽  
Valérie Bégay ◽  
Anja Schuetz ◽  
Udo Heinemann ◽  
Achim Leutz
Keyword(s):  
Transcription Factor ◽  
Transcriptional Activity ◽  
Target Genes ◽  
Binding Protein ◽  
Pocket Proteins ◽  
Specific Differentiation ◽  
Enhancer Binding Protein ◽  
Transactivation Potential ◽  
Ccaat Enhancer Binding Protein ◽  
Acute Myeloid

ABSTRACT The transcription factor CCAAT/enhancer-binding protein α (C/EBPα) coordinates proliferation arrest and the differentiation of myeloid progenitors, adipocytes, hepatocytes, keratinocytes, and cells of the lung and placenta. C/EBPα transactivates lineage-specific differentiation genes and inhibits proliferation by repressing E2F-regulated genes. The myeloproliferative C/EBPα BRM2 mutant serves as a paradigm for recurrent human C-terminal bZIP C/EBPα mutations that are involved in acute myeloid leukemogenesis. BRM2 fails to repress E2F and to induce adipogenesis and granulopoiesis. The data presented here show that, independently of pocket proteins, C/EBPα interacts with the dimerization partner (DP) of E2F and that C/EBPα-E2F/DP interaction prevents both binding of C/EBPα to its cognate sites on DNA and transactivation of C/EBP target genes. The BRM2 mutant, in addition, exhibits enhanced interaction with E2F-DP and reduced affinity toward DNA and yet retains transactivation potential and differentiation competence that becomes exposed when E2F/DP levels are low. Our data suggest a tripartite balance between C/EBPα, E2F/DP, and pocket proteins in the control of proliferation, differentiation, and tumorigenesis.

scholarly journals Glucocorticoid Regulation of the Promoter of 11β-Hydroxysteroid Dehydrogenase Type 1 Is Indirect and Requires CCAAT/Enhancer-Binding Protein-β

2008 ◽  
Vol 22 (9) ◽  
pp. 2049-2060 ◽  
Author(s):  
Shuji Sai ◽  
Cristina L. Esteves ◽  
Val Kelly ◽  
Zoi Michailidou ◽  
Karen Anderson ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Binding Protein ◽  
A549 Cells ◽  
Hydroxysteroid Dehydrogenase ◽  
Glucocorticoid Treatment ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein ◽  
Glucocorticoid Induction ◽  
Glucocorticoid Action

Abstract 11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) converts inert 11keto-glucocorticoids to active 11β-hydroxy forms, thereby amplifying intracellular glucocorticoid action. Up-regulation of 11β-HSD1 in adipose tissue and liver is of pathogenic importance in metabolic syndrome. However, the mechanisms controlling 11β-HSD1 transcription are poorly understood. Glucocorticoids themselves potently increase 11β-HSD1 expression in many cells, providing a potential feed-forward system to pathology. We have investigated the molecular mechanisms by which glucocorticoids regulate transcription of 11β-HSD1, exploiting an A549 cell model system in which endogenous 11β-HSD1 is expressed and is induced by dexamethasone. We show that glucocorticoid induction of 11β-HSD1 is indirect and requires new protein synthesis. A glucocorticoid-responsive region maps to between −196 and −88 with respect to the transcription start site. This region contains two binding sites for CCAAT/enhancer-binding protein (C/EBP) that together are essential for the glucocorticoid response and that bind predominantly C/EBPβ, with C/EBPδ present in a minority of the complexes. Both C/EBPβ and C/EBPδ are rapidly induced by glucocorticoids in A549 cells, but small interfering RNA-mediated knockdown shows that only C/EBPβ reduction attenuates the glucocorticoid induction of 11β-HSD1. Chromatin immunoprecipitation studies demonstrated increased binding of C/EBPβ to the 11β-HSD1 promoter in A549 cells after glucocorticoid treatment. A similar mechanism may apply in adipose tissue in vivo where increased C/EBPβ mRNA levels after glucocorticoid treatment were associated with increased 11β-HSD1 expression. C/EBPβ is a key mediator of metabolic and inflammatory signaling. Positive regulation of 11β-HSD1 by C/EBPβ may link amplification of glucocorticoid action with metabolic and inflammatory pathways and may represent an endogenous innate host-defense mechanism.

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