Identification of a silencer module which selectively represses cyclic AMP-responsive element-dependent gene expression

Kwang Chul Chung, Delai Huang, Yuexian Chen, Sabine Short, Marc L. Short, Zhiping Zhang, Richard A. Jungmann

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Abstract

The cyclic AMP (cAMP)-inducible promoter from the rat lactate dehydrogenase A subunit gene (LDHA) is associated with a distal negative regulatory element (LDH-NRE) that represses inherent basal and cAMP-inducible promoter activity. The element is of dyad symmetry, consisting of a palindromic sequence with two half-sites, 5'-TCTTG-3'. It represses the expression of an LDH A/chloramphenicol acetyltransferase (CAT) reporter gene in a dose-dependent, orientation- and position-independent fashion, suggesting that it is a true silencer element. Uniquely, it selectively represses cAMP-responsive element (CRE)-dependent transcription but has no effect on promoters lacking a CRE sequence. The repressing action of LDH-NRE could be overcome by cotransfection with LDh A/CAT vector oligonucleotides containing either the LDH-NRE or CRE sequence. This suggests that the reversal of repression was caused by the removal of functional active, limiting transacting factors which associate with LDH-NRE as well as with CRE. Gel mobility shift, footprinting, and Southwestern blotting assays demonstrated the presence of a 69-kDa protein with specific binding activity for LDH-NRE. Additionally, gel supershift assays with anti-CREB and anti-Fos antibodies indicate the presence of CREB and Fos or antigenically closely related proteins with the LDH-NRE/protein complex. We suggest that the LDH- NRE and CRE modules functionally interact to achieve negative modulation of cAMP-responsive LDH A transcriptional activity.

Original languageEnglish
Pages (from-to)6139-6149
Number of pages11
JournalMolecular and Cellular Biology
Volume15
Issue number11
DOIs
Publication statusPublished - 1995 Jan 1

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Cyclic AMP
Gene Expression
Chloramphenicol O-Acetyltransferase
Southwestern Blotting
Gels
Transcriptional Silencer Elements
Proteins
Reporter Genes
Oligonucleotides
Anti-Idiotypic Antibodies
Genes
lactate dehydrogenase 5

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology

Cite this

Chung, Kwang Chul ; Huang, Delai ; Chen, Yuexian ; Short, Sabine ; Short, Marc L. ; Zhang, Zhiping ; Jungmann, Richard A. / Identification of a silencer module which selectively represses cyclic AMP-responsive element-dependent gene expression. In: Molecular and Cellular Biology. 1995 ; Vol. 15, No. 11. pp. 6139-6149.
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Identification of a silencer module which selectively represses cyclic AMP-responsive element-dependent gene expression. / Chung, Kwang Chul; Huang, Delai; Chen, Yuexian; Short, Sabine; Short, Marc L.; Zhang, Zhiping; Jungmann, Richard A.

In: Molecular and Cellular Biology, Vol. 15, No. 11, 01.01.1995, p. 6139-6149.

Research output: Contribution to journalArticle

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AU - Chung, Kwang Chul

AU - Huang, Delai

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