Activation of CMV promoter-controlled glycosyltransferase and β-galactosidase glycogenes by butyrate, tricostatin A, and 5-aza-2′-deoxycytidine

Hyun Choi Kyung, Hesham E Basma, Jaswant Singh, Pi-Wan Cheng

Research output: Contribution to journalArticle

57 Citations (Scopus)

Abstract

Cytomegalovirus (CMV) immediate early promoter is a powerful promoter frequently used for driving the expression of transgenes in mammalian cells. However, this promoter gradually becomes silenced in stably transfected cells. We employed Chinese Hamster Ovary (CHO) and human pancreatic cancer (Panc 1) cells stably tansfected with three glycogenes driven by a CMV promoter to study the activation of silenced glycogenes. We found that butyrate, tricostatin A (TSA), and 5-aza-2′-deoxycytidine (5-Aza-dC) can activate these CMV-driven glycogenes. The increase in mRNA and protein of a glycogene occurred 8-10 h after butyrate treatment, suggesting an indirect effect of butyrate in the activation of the transgene. The enhanced expression of the trangenes by butyrate and TSA, known inhibitors of histone deacetylase, was independent of the transgene or cell type. However, the transgene can be activated by these two agents in only a fraction of the cells derived from a single clone, suggesting that inactivation of histone deacetylase can only partially explain silencing of the transgenes. Combination treatment of one or both agents with 5-Aza-dC, a known inhibitor of DNA methylase, resulted in a synergistic activation of the transgene, suggesting a cross-talk between histone acetylation and DNA demethylation. Understanding the mechanisms of the inactivation and reactivation of CMV promoter-controlled transgenes should help develop an effective strategy to fully activate the CMV promoter-controlled therapeutic genes silenced by the host cells.

Original languageEnglish (US)
Pages (from-to)63-69
Number of pages7
JournalGlycoconjugate Journal
Volume22
Issue number1-2
DOIs
StatePublished - Feb 1 2005

Fingerprint

decitabine
trichostatin A
Galactosidases
Glycosyltransferases
Butyrates
Cytomegalovirus
Transgenes
Chemical activation
Acetylation
Histone Deacetylase Inhibitors
Histone Deacetylases
DNA
Histones
Genes
Cells
Messenger RNA
Cricetulus
Pancreatic Neoplasms
Ovary

Keywords

  • 5-Aza-2′-deoxycytidine
  • Butyrate
  • CMV promoter
  • Glycosyltransferases
  • Tricostatin A
  • β-galactosidase

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Activation of CMV promoter-controlled glycosyltransferase and β-galactosidase glycogenes by butyrate, tricostatin A, and 5-aza-2′-deoxycytidine. / Kyung, Hyun Choi; Basma, Hesham E; Singh, Jaswant; Cheng, Pi-Wan.

In: Glycoconjugate Journal, Vol. 22, No. 1-2, 01.02.2005, p. 63-69.

Research output: Contribution to journalArticle

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AB - Cytomegalovirus (CMV) immediate early promoter is a powerful promoter frequently used for driving the expression of transgenes in mammalian cells. However, this promoter gradually becomes silenced in stably transfected cells. We employed Chinese Hamster Ovary (CHO) and human pancreatic cancer (Panc 1) cells stably tansfected with three glycogenes driven by a CMV promoter to study the activation of silenced glycogenes. We found that butyrate, tricostatin A (TSA), and 5-aza-2′-deoxycytidine (5-Aza-dC) can activate these CMV-driven glycogenes. The increase in mRNA and protein of a glycogene occurred 8-10 h after butyrate treatment, suggesting an indirect effect of butyrate in the activation of the transgene. The enhanced expression of the trangenes by butyrate and TSA, known inhibitors of histone deacetylase, was independent of the transgene or cell type. However, the transgene can be activated by these two agents in only a fraction of the cells derived from a single clone, suggesting that inactivation of histone deacetylase can only partially explain silencing of the transgenes. Combination treatment of one or both agents with 5-Aza-dC, a known inhibitor of DNA methylase, resulted in a synergistic activation of the transgene, suggesting a cross-talk between histone acetylation and DNA demethylation. Understanding the mechanisms of the inactivation and reactivation of CMV promoter-controlled transgenes should help develop an effective strategy to fully activate the CMV promoter-controlled therapeutic genes silenced by the host cells.

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