Manipulation of the cytoplasmic and transmembrane domains alters cell surface levels of the coxsackie-adenovirus receptor and changes the efficiency of adenovirus infection. Academic Article uri icon

Overview

MeSH

  • Animals
  • CHO Cells
  • COS Cells
  • Coxsackie and Adenovirus Receptor-Like Membrane Protein
  • Cricetinae
  • DNA Primers
  • Fluorescent Dyes
  • Gene Transfer Techniques
  • Genetic Vectors
  • Glycosylphosphatidylinositols
  • Humans
  • Oligopeptides
  • Peptides
  • Phosphatidylinositol Diacylglycerol-Lyase
  • RNA, Messenger
  • Type C Phospholipases
  • beta-Galactosidase

MeSH Major

  • Adenoviridae
  • Adenoviridae Infections
  • Enterovirus
  • Receptors, Virus

abstract

  • Expression of the coxsackie-adenovirus receptor (CAR) is a critical determinant in cellular susceptibility to infection with adenovirus-based gene transfer vectors. This study is focused on the hypothesis that manipulation of the cytoplasmic tail and transmembrane regions of CAR can be used to change cell surface levels of CAR and, consequently, to alter the efficiency of Ad-mediated gene transfer. To accomplish this, Flag-tagged ([F]) human CAR ([F]CAR), [F]tailless-CAR (lacking the cytoplasmic tail), and [F]GPI-CAR (containing a GPI lipid anchor instead of the transmembrane and cytoplasmic regions) were exogenously expressed in CHO cells. Analysis of (125)I-labeled anti-Flag antibody binding to transfected cells revealed that [F]tailless-CAR and [F]GPI-CAR were expressed on the cell surface in 1.8- to 2.5-fold higher amounts than [F]CAR, while the total expression levels were similar. Infection with replication-deficient adenovirus encoding beta-galactosidase (Ad-betagal) demonstrated 1.5- to 2-fold higher levels of transgene expression in CHO cells expressing [F]tailless-CAR or [F]GPI-CAR, respectively, compared with cells containing [F]CAR. The form of CAR expressed did not affect the transport of fluorescent Cy3-Ad particles from the cell surface to the nuclear region. These observations indicate that transduction of target cells by Ad vectors can be optimized by increasing cell surface levels of CAR through functional deletion of the tail and membrane protein domains.

publication date

  • January 1, 2001

has subject area

  • Adenoviridae
  • Adenoviridae Infections
  • Animals
  • CHO Cells
  • COS Cells
  • Coxsackie and Adenovirus Receptor-Like Membrane Protein
  • Cricetinae
  • DNA Primers
  • Enterovirus
  • Fluorescent Dyes
  • Gene Transfer Techniques
  • Genetic Vectors
  • Glycosylphosphatidylinositols
  • Humans
  • Oligopeptides
  • Peptides
  • Phosphatidylinositol Diacylglycerol-Lyase
  • RNA, Messenger
  • Receptors, Virus
  • Type C Phospholipases
  • beta-Galactosidase

Research

keywords

  • Journal Article

Identity

Language

  • eng

Digital Object Identifier (DOI)

  • 10.1089/104303401450933

PubMed ID

  • 11177539

Additional Document Info

start page

  • 25

end page

  • 34

volume

  • 12

number

  • 1