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Molecular and Cellular Biology, July 2001, p. 4636-4646, Vol. 21, No. 14
0270-7306/01/$04.00+0   DOI: 10.1128/MCB.21.14.4636-4646.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

CREB Is One Component of the Binding Complex of the Ces-2/E2A-HLF Binding Element and Is an Integral Part of the Interleukin-3 Survival Signal

Wannhsin Chen,1,2 Yung-Luen Yu,1,2 Shern-Fwu Lee,2 Yun-Jung Chiang,2 Jyh-Rong Chao,3 Jin-Hwa Huang,2 Jiao-How Chiong,2 Chang-Jen Huang,4 Ming-Zong Lai,3 Hsin-Fang Yang-Yen,3 and Jeffrey J.-Y. Yen2,*

Graduate Institute of Life Sciences, National Defense Medical Center1 and Institute of Biomedical Sciences,2 Institute of Molecular Biology,3 and Institute of Biological Chemistry,4 Academia Sinica, Taipei, Taiwan

Received 2 March 2001/Returned for modification 4 April 2001/Accepted 19 April 2001

The Ces-2/E2A-HLF binding element (CBE) is recognized by Caenorhabditis elegans death specification gene product Ces-2 and human acute lymphocytic leukemia oncoprotein E2A-HLF. In an attempt to identify a cellular CBE-binding protein(s) that may be involved in apoptosis regulation in mammals, multiple nuclear binding complexes of CBE were identified in various mammalian cell lines and tissues by electrophoretic mobility shift assay. Cyclic AMP (cAMP)-responsive element (CRE)-binding protein (CREB) was present in one major CBE complex of Ba/F3 and TF-1 cells, and both in vitro-translated and Escherichia coli-synthesized CREB bound to CBE. Activation of CREB by cAMP-elevating chemicals or the catalytic subunit of protein kinase A (PKAc) resulted in induction of the CBE-driven reporter gene. Stimulation of Ba/F3 cells with interleukin-3 (IL-3) promptly induced phosphorylation of CREB at serine133 partially via a PKA-dependent pathway. Consistently, Ba/F3 cell survival in the absence of IL-3 was prolonged by activation of PKA. Conversely, treatment of cells with a PKA inhibitor or expression of the dominant negative forms of the regulatory subunit type I of PKA and CREB overrode the survival activity of IL-3. Last, the bcl-2 gene was demonstrated to be one candidate cellular target of the CREB-containing CBE complex, as mutations in the CRE and CBE sites significantly reduced the IL-3 inducibility of the bcl-2 promoter. Together, our results suggest that CREB is one cellular counterpart of Ces-2/E2A-HLF and is part of IL-3 dependent apoptosis regulation in hematopoietic cells.

* Corresponding author. Mailing address: Institute of Biomedical Sciences, Academia Sinica, No. 128, Sec. 2, Yen-Jiou-Yuan Rd., Taipei 11529, Taiwan. Phone: 886-2-2652-3077. Fax: 886-2-2782-9142. E-mail: bmjyen{at}novell.ibms.sinica.edu.tw.

Molecular and Cellular Biology, July 2001, p. 4636-4646, Vol. 21, No. 14
0270-7306/01/$04.00+0   DOI: 10.1128/MCB.21.14.4636-4646.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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