Activation of CMV promoter-controlled glycosyltransferase and beta -galactosidase glycogenes by butyrate, tricostatin A, and 5-aza-2'-deoxycytidine.
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.
Choi
Kyung Hyun
KH
Department of Biochemistry and Molecular Biology, College of Medicine, University of Nebraska Medical Center, Omaha, 68198-5870, USA.
Basma
Hesham
H
Singh
Jaswant
J
Cheng
Pi-Wan
PW
D000818:Animals; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002087:Butyrates / Q000494:pharmacology*; D045744:Cell Line, Tumor; D006224:Cricetinae; D003412:Cricetulus; D003587:Cytomegalovirus / Q000235:genetics*; D015254:DNA Modification Methylases / Q000037:antagonists & inhibitors; D000077209:Decitabine; D004357:Drug Synergism; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D020868:Gene Silencing; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism*; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism*
D002087:Butyrates; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D012333:RNA, Messenger; C012589:trichostatin A; D000077209:Decitabine; D015254:DNA Modification Methylases; D016695:Glycosyltransferases; D001616:beta-Galactosidase; D001374:Azacitidine