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Journal of Virology, August 2001, p. 7703-7711, Vol. 75, No. 16
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.16.7703-7711.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Apical Localization of the Coxsackie-Adenovirus Receptor by Glycosyl-Phosphatidylinositol Modification Is Sufficient for Adenovirus-Mediated Gene Transfer through the Apical Surface of Human Airway Epithelia

Robert W. Walters,1,2,3 Wouter van't Hof,4,5 Su Min P. Yi,1,6 Mary K. Schroth,7 Joseph Zabner,2 Ronald G. Crystal,4,5 and Michael J. Welsh1,2,3,*

Howard Hughes Medical Institute1 and Departments of Internal Medicine,2 Physiology and Biophysics,3 and Otolaryngology,6 University of Iowa College of Medicine, Iowa City, Iowa 52242; Division of Pulmonary and Critical Care Medicine,4 and Institute of Genetic Medicine,5 Weill Medical College of Cornell University, New York, New York 10021; and Department of Pediatrics, University of Wisconsin Medical School, Madison, Wisconsin 537927

Received 21 March 2001/Accepted 12 May 2001

In well-differentiated human airway epithelia, the coxsackie B and adenovirus type 2 and 5 receptor (CAR) resides primarily on the basolateral membrane. This location may explain the observation that gene transfer is inefficient when adenovirus vectors are applied to the apical surface. To further test this hypothesis and to investigate requirements and barriers to apical gene transfer to differentiated human airway epithelia, we expressed CAR in which the transmembrane and cytoplasmic tail were replaced by a glycosyl-phosphatidylinositol (GPI) anchor (GPI-CAR). As controls, we expressed wild-type CAR and CAR lacking the cytoplasmic domain (Tailless-CAR). All three constructs enhanced gene transfer with similar efficiencies in fibroblasts. In airway epithelia, GPI-CAR localized specifically to the apical membrane, where it bound adenovirus and enhanced gene transfer to levels obtained when vector was applied to the basolateral membrane. Moreover, GPI-CAR facilitated gene transfer of the cystic fibrosis transmembrane conductance regulator to cystic fibrosis airway epithelia, correcting the Cl- transport defect. In contrast, when we expressed wild-type CAR it localized to the basolateral membrane and failed to increase apical gene transfer. Only a small amount of Tailless-CAR resided in the apical membrane, and the effects on apical virus binding and gene transfer were minimal. These data indicate that binding of adenovirus to an apical membrane receptor is sufficient to mediate effective gene transfer to human airway epithelia and that the cytoplasmic domain of CAR is not required for this process. The results suggest that targeting apical receptors in differentiated airway epithelia may be sufficient for gene transfer in the genetic disease cystic fibrosis.

* Corresponding Author. Mailing address: Howard Hughes Medical Institute, University of Iowa College of Medicine, 500 EMRB, Iowa City, IA 52242. Phone: (319) 335-7619. Fax: (319) 335-7623. E-mail: mjwelsh{at}blue.weeg.uiowa.edu.

Journal of Virology, August 2001, p. 7703-7711, Vol. 75, No. 16
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.16.7703-7711.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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