Sorting of Two Polytopic Proteins, the γ-Aminobutyric Acid and Betaine Transporters, in Polarized Epithelial Cells

doi:10.1074/jbc.272.10.6584

Sorting of Two Polytopic Proteins, the γ-Aminobutyric Acid and Betaine Transporters, in Polarized Epithelial Cells*

From the ^‡ Consiglio Nazionale delle Ricerche Cellular and Molecular Pharmacology Center, Department of Pharmacology, University of Milan, Milan 20129, the
^§ Consiglio Nazionale delle Ricerche Institute of Hormone Chemistry, Milan 20129, the
^¶ DIBIT-San Raffaele Scientific Institute and Ceccarelli Center, Milan 20132, Italy, and the
^∥ Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06510

** To whom correspondence should be addressed:
CNR Cellular and Molecular Pharmacology Center, Via Vanvitelli, 32, 20129 Milano, Italy.
Tel.: 39-2-701-46358; Fax: 39-2-749-0574; E-mail: Grazia{at}Farma1.csfic.mi.cnr.it

Abstract

The γ-aminobutyric acid transporter (GAT-1) isoform of the γ-aminobutyric acid and the betaine (BGT) transporters exhibit distinct apical and basolateral distributions when introduced into Madin-Darby canine kidney cells (Pietrini, G., Suh, Y. J., Edelman, L., Rudnick, G., and Caplan, M. J. (1994) J. Biol. Chem. 269, 4668-4674). We have investigated the presence of sorting signals in their COOH-terminal cytosolic domains by expression in Madin-Darby canine kidney cells of mutated and chimeric transporters. Whereas truncated GAT-1 (ΔC-GAT) maintained the original functional activity and apical localization, either the removal (ΔC-myc BGT) or the substitution (BGS chimera) of the cytosolic tail of BGT generated proteins that accumulated in the endoplasmic reticulum. Moreover, we have found that the cytosolic tail of BGT redirected apical proteins, the polytopic GAT-1 (GBS chimera) and the monotopic human nerve growth factor receptor, to the basolateral surface. These results suggest the presence of basolateral sorting information in the cytosolic tail of BGT. We have further shown that information necessary for the exit of BGT from the endoplasmic reticulum and for the basolateral localization of the GBS chimera is contained in a short segment, rich in basic residues, within the cytosolic tail of BGT.

Footnotes

↵* This work was supported in part by Consiglio Nazionale delle Ricerche Grant 94.00377.CT14.115.28212B/0004 (to G. P.). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

MDCK

Madin-Darby canine kidney

GABA

γ-aminobutyric acid

GAT-1

GABA transporter

BGT

betaine transporter

hNGFR

human nerve growth factor receptor

PCR

polymerase chain reaction

KLH

keyhole limpet hemocyanin

ER

endoplasmic reticulum

mAb

monoclonal antibody.
↵²M. J. Caplan, unpublished data.
- Received June 25, 1996.
- Revision received October 28, 1996.