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Importance of cAMP-response element-binding protein in regulation of expression of the murine cyclic nucleotide phosphodiesterase 3B (Pde3b) gene in differentiating 3T3-L1 preadipocytes

Liu, Hanguan ; Tang, Jing Rong ; Choi, Young Hun ; Napolitano, Maria ; Hockman, Steven ; Taira, Masato ; Degerman, Eva LU orcid and Manganiello, Vincent C. (2006) In Journal of Biological Chemistry 281(30). p.21096-21113
Abstract
Incubation of 3T3-L1 preadipocytes with isobutylmethylxanthine (IBMX), dexamethasone, and insulin, alone or in combination, demonstrated that IBMX, which increased cAMP-response element-binding protein (CREB) phosphorylation, was the predominant regulator of Pde3b expression. Real time PCR and immunoblotting indicated that in 3T3-L1 preadipocytes, IBMX-stimulated induction of Pde3b mRNA and protein was markedly inhibited by dominant-negative CREB proteins. By transfecting preadipocytes, differentiating preadipocytes, and HEK293A cells with luciferase reporter vectors containing different fragments of the 5'- flanking region of the Pde3b gene, we identified a distal promoter that contained canonical cis-acting cAMP-response elements (CRE)... (More)
Incubation of 3T3-L1 preadipocytes with isobutylmethylxanthine (IBMX), dexamethasone, and insulin, alone or in combination, demonstrated that IBMX, which increased cAMP-response element-binding protein (CREB) phosphorylation, was the predominant regulator of Pde3b expression. Real time PCR and immunoblotting indicated that in 3T3-L1 preadipocytes, IBMX-stimulated induction of Pde3b mRNA and protein was markedly inhibited by dominant-negative CREB proteins. By transfecting preadipocytes, differentiating preadipocytes, and HEK293A cells with luciferase reporter vectors containing different fragments of the 5'- flanking region of the Pde3b gene, we identified a distal promoter that contained canonical cis-acting cAMP-response elements (CRE) and a proximal, GC-rich promoter region, which contained atypical CRE. Mutation of the CRE sequences dramatically reduced distal promoter activity; H89 inhibited IBMX-stimulated CREB phosphorylation and proximal and distal promoter activities. Distal promoter activity was stimulated by IBMX and phorbol ester (PMA) in Raw264.7 monocytes, but only by IBMX in 3T3-L1 preadipocytes. Chromatin immunoprecipitation analyses with specific antibodies against CREB, phospho-CREB, and CBP/p300 (CREB-binding protein) showed that these proteins associated with both distal and proximal promoters and that interaction of phospho-CREB, the active form of CREB, with both Pde3b promoter regions was increased in IBMX-treated preadipocytes. These results indicate that CRE in distal and proximal promoter regions and activation of CREB proteins play a crucial role in transcriptional regulation of Pde3b expression during preadipocyte differentiation. (Less)
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author
; ; ; ; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of Biological Chemistry
volume
281
issue
30
pages
21096 - 21113
publisher
American Society for Biochemistry and Molecular Biology
external identifiers
  • wos:000239187300046
  • scopus:33746374753
  • pmid:16702214
ISSN
1083-351X
DOI
10.1074/jbc.M601307200
language
English
LU publication?
yes
id
22b14732-23a7-49da-acf7-b7dd55095f70 (old id 401346)
date added to LUP
2016-04-01 12:17:37
date last changed
2022-01-27 01:39:32
@article{22b14732-23a7-49da-acf7-b7dd55095f70,
  abstract     = {{Incubation of 3T3-L1 preadipocytes with isobutylmethylxanthine (IBMX), dexamethasone, and insulin, alone or in combination, demonstrated that IBMX, which increased cAMP-response element-binding protein (CREB) phosphorylation, was the predominant regulator of Pde3b expression. Real time PCR and immunoblotting indicated that in 3T3-L1 preadipocytes, IBMX-stimulated induction of Pde3b mRNA and protein was markedly inhibited by dominant-negative CREB proteins. By transfecting preadipocytes, differentiating preadipocytes, and HEK293A cells with luciferase reporter vectors containing different fragments of the 5'- flanking region of the Pde3b gene, we identified a distal promoter that contained canonical cis-acting cAMP-response elements (CRE) and a proximal, GC-rich promoter region, which contained atypical CRE. Mutation of the CRE sequences dramatically reduced distal promoter activity; H89 inhibited IBMX-stimulated CREB phosphorylation and proximal and distal promoter activities. Distal promoter activity was stimulated by IBMX and phorbol ester (PMA) in Raw264.7 monocytes, but only by IBMX in 3T3-L1 preadipocytes. Chromatin immunoprecipitation analyses with specific antibodies against CREB, phospho-CREB, and CBP/p300 (CREB-binding protein) showed that these proteins associated with both distal and proximal promoters and that interaction of phospho-CREB, the active form of CREB, with both Pde3b promoter regions was increased in IBMX-treated preadipocytes. These results indicate that CRE in distal and proximal promoter regions and activation of CREB proteins play a crucial role in transcriptional regulation of Pde3b expression during preadipocyte differentiation.}},
  author       = {{Liu, Hanguan and Tang, Jing Rong and Choi, Young Hun and Napolitano, Maria and Hockman, Steven and Taira, Masato and Degerman, Eva and Manganiello, Vincent C.}},
  issn         = {{1083-351X}},
  language     = {{eng}},
  number       = {{30}},
  pages        = {{21096--21113}},
  publisher    = {{American Society for Biochemistry and Molecular Biology}},
  series       = {{Journal of Biological Chemistry}},
  title        = {{Importance of cAMP-response element-binding protein in regulation of expression of the murine cyclic nucleotide phosphodiesterase 3B (Pde3b) gene in differentiating 3T3-L1 preadipocytes}},
  url          = {{http://dx.doi.org/10.1074/jbc.M601307200}},
  doi          = {{10.1074/jbc.M601307200}},
  volume       = {{281}},
  year         = {{2006}},
}