Effect of intron A from human cytomegalovirus (Towne) immediate-early gene on heterologous expression in mammalian cells

Barbara S. Chapman, Richard M. Thayer, Karen A. Vincent, Nancy Haigwood

Research output: Contribution to journalArticle

261 Citations (Scopus)

Abstract

A 2.4 kb fragment of hCMV (Towne strain), containing the 5′ end of the major immediate-early gene, has been cloned, sequenced, and used to construct a series of mammalian cell expression plasmids. The effects of regulatory regions present on this fragment were assessed using human glycoproteins as reporter molecules. We compared secreted levels of Factor VIII, t-PA, and HIV-1 envelope glycoproteins in cells transfected with plasmids in which intron A of the immediate-early gene was present or absent. Secretion of several glycoproteins was significantly higher when cells were transfected with intron A-containing plasmids. Mutation of three basepairs in the strong nuclear factor 1 (NF1) binding site in intron A led to reduced transient expression levels, but not to the level observed in the absence of intron A. Reduced expression from NF1 mutant plasmids was roughly correlated with reduced binding in vitro of NF1 proteins to a synthetic oligonucleotide containing the mutation. The evidence indicates that sequences in intron A positively regulate expression from the hCMV immediate-early enhancer/promoter in transformed monkey kidney cells.

Original languageEnglish (US)
Pages (from-to)3979-3986
Number of pages8
JournalNucleic Acids Research
Volume19
Issue number14
StatePublished - Jul 25 1991
Externally publishedYes

Fingerprint

Glycoproteins
Immediate-Early Genes
Cytomegalovirus
Introns
Glycoprotein
NFI Transcription Factors
Genes
Cells
Gene
Plasmids
Cell
Fragment
Mutation
Oligonucleotides
Binding sites
Secretion
Kidney
Promoter
Mutant
Envelope

ASJC Scopus subject areas

  • Genetics
  • Statistics, Probability and Uncertainty
  • Applied Mathematics
  • Health, Toxicology and Mutagenesis
  • Toxicology
  • Genetics(clinical)

Cite this

Effect of intron A from human cytomegalovirus (Towne) immediate-early gene on heterologous expression in mammalian cells. / Chapman, Barbara S.; Thayer, Richard M.; Vincent, Karen A.; Haigwood, Nancy.

In: Nucleic Acids Research, Vol. 19, No. 14, 25.07.1991, p. 3979-3986.

Research output: Contribution to journalArticle

Chapman, Barbara S. ; Thayer, Richard M. ; Vincent, Karen A. ; Haigwood, Nancy. / Effect of intron A from human cytomegalovirus (Towne) immediate-early gene on heterologous expression in mammalian cells. In: Nucleic Acids Research. 1991 ; Vol. 19, No. 14. pp. 3979-3986.
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