Calmodulin binds to the basolateral targeting signal of the polymeric immunoglobulin receptor.
We have identified a major calmodulin (CaM)-binding protein in rat liver endosomes using 125I-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(2+)-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 35S-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.
Chapin
S J
SJ
Department of Anatomy, University of California, San Francisco 94143, USA.
Enrich
C
C
Aroeti
B
B
Havel
R J
RJ
Mostov
K E
KE
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002147:Calmodulin / Q000378:metabolism*; D002148:Calmodulin-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D002460:Cell Line; D002846:Chromatography, Affinity; D004285:Dogs; D015180:Electrophoresis, Gel, Two-Dimensional; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism; D004958:Estradiol / Q000494:pharmacology; D007668:Kidney; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D051381:Rats; D019056:Receptors, Polymeric Immunoglobulin / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity; D014162:Transfection
D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D019056:Receptors, Polymeric Immunoglobulin; D011994:Recombinant Proteins; D004958:Estradiol