Calmodulin Binds to the Basolateral Targeting Signal of the Polymeric Immunoglobulin Receptor

doi:10.1074/jbc.271.3.1336

Calmodulin Binds to the Basolateral Targeting Signal of the Polymeric Immunoglobulin Receptor (*)

From the (1)Departments of Anatomy and Biochemistry and Biophysics
(2)Cardiovascular Research Institute, and
(3)Department of Medicine, University of California, San Francisco, California 94143 and the
(4)Departamento de Biologia Celular, Facultad de Medicina, Universidad de Barcelona, 08028 Barcelona, Spain

^§ To whom correspondence should be addressed:
Box 0452, Dept. of Anatomy, University of California, San Francisco, CA 94143-0452.
Tel.: 415-476-6048; Fax: 415-476-4845.

↵^§§ Present address: Dept. of Cell and Animal Biology, Institute of Life Sciences, Hebrew University of Jerusalem, Jerusalem 91904, Israel.

Abstract

We have identified a major calmodulin (CaM)-binding protein in rat liver endosomes using I-CaM overlays from two-dimensional protein blots. Immunostaining of blots demonstrates that this protein is the polymeric immunoglobulin receptor (pIgR). We further investigated the interaction between pIgR and CaM using Madin-Darby canine kidney cells stably expressing cloned wild-type and mutant pIgR. We found that detergent-solubilized pIgR binds to CaM-agarose in a Ca-dependent fashion, and binding is inhibited by the addition of excess free CaM or the CaM antagonist W-13 (N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide), suggesting that pIgR binding to CaM is specific. Furthermore, pIgR is the most prominent S-labeled CaM-binding protein in the detergent phase of Triton X-114-solubilized, metabolically labeled pIgR-expressing Madin-Darby canine kidney cells. CaM can be chemically cross-linked to both solubilized and membrane-associated pIgR, suggesting that binding can occur while the pIgR is in intact membranes. The CaM binding site is located in the membrane-proximal 17-amino acid segment of the pIgR cytoplasmic tail. This region of pIgR constitutes an autonomous basolateral targeting signal. However, binding of CaM to various pIgR mutants suggests that CaM binding is not necessary for basolateral targeting. We suggest that CaM may be involved in regulation of pIgR transcytosis and/or signaling by pIgR.

Footnotes

↵^¶ These authors made equal contributions to this work.
↵** Supported by fellowships from Ministerio de Educación y Ciencia and CIRIT 93-22.
↵* This work was supported in part by National Institutes of Health Grants HL-14237 (Arteriosclerosis SCOR) (to R. H.) and RO1 AI 25144 and R01 AI 36953 (to K. M.), American Cancer Society Grant IM 763 (to K. M.), an Established Investigator Award from the American Heart Association-Wyeth-Ayerst (to K. M.), a grant from the Lucille P. Markey Charitable Trust (to the UCSF Cell Biology Program), an American Heart Association California Division Fellowship (to B. A.), and American Cancer Society Fellowship PF3666 (to S. C.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used in this paper are:

TGN

trans-Golgi network

pIgR

polymeric immunoglobulin receptor

CaM

calmodulin

d/pIgA

dimeric/polymeric immunoglobulin A

VSVG

vesicular stomatitis virus protein G

BS³

bis(sulfosuccinimidyl) suberate

[Ca]

cytosolic free calcium

RRC

receptor recycling compartment

MVB

multivesicular body

CURL

compartment of uncoupling of receptor and ligand

SC

secretory component

PAGE

polyacrylamide gel electrophoresis.
↵²M. Cardone, F. Luton, and K. E. Mostov, unpublished data.
↵³M. Cardone and K. E. Mostov, unpublished data.
- Received August 23, 1995.