Requirements for enhanced transgene expression by untranslated sequences from the human cytomegalovirus immediate-early gene

Robert D. Simari

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

Background: The cytomegalovirus immediate early (CMV IE) promoter has been widely used for heterologous expression. Further enhancements of gene expression from this potent promoter may allow for the development of improved gene transfer strategies. We aimed to determine whether inclusion of the first exon (5' untranslated) and first intron of the CMV IE gene would increase heterologous transgene expression in primary target cells and to determine the sequences required for any observed increases. Materials and Methods: Comparisons of reporter gene expression were made following transient transfection of vascular smooth muscle cells (VSMCs) with plasmids containing the first exon and intron from the CMV IE gene or deletional mutations. Comparisons were also made using a heterologous promoter (RSV). Results: Gene expression from the CMV IE promoter was increased 5.7-fold in VSMC with the inclusion of the first exon and intron. Similar increases were seen with other target cells and from the heterologous RSV promoter. This increase was associated with an increase in steady-state mRNA. Deletion analyses demonstrated that the enhancement was dependent on the presence of the 5' portion of the first exon while deletion of large segments within the intron was associated with similar levels of expression compared with the parental plasmid. Conclusions: Inclusion of the first exon and intron from the CMV IE gene increases expression from the CMV IE promoter. This enhancement is seen with the heterologous RSV promoter and is associated with an increase in steady-state mRNA. Deletion analyses suggest that this enhancement is associated with inclusion of sequences within the 5' portion of the first exon and inclusion of an intron.

Original languageEnglish (US)
Pages (from-to)700-706
Number of pages7
JournalMolecular Medicine
Volume4
Issue number11
StatePublished - 1998

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Immediate-Early Genes
Cytomegalovirus
Transgenes
Introns
Exons
Gene Expression
Vascular Smooth Muscle
Smooth Muscle Myocytes
Plasmids
Messenger RNA
Reporter Genes
Transfection
Mutation
Genes

ASJC Scopus subject areas

  • Genetics

Cite this

Requirements for enhanced transgene expression by untranslated sequences from the human cytomegalovirus immediate-early gene. / Simari, Robert D.

In: Molecular Medicine, Vol. 4, No. 11, 1998, p. 700-706.

Research output: Contribution to journalArticle

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