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Targeting mucosal sites by polymeric immunoglobulin receptor-directed peptides.

White KD, Capra JD - J. Exp. Med. (2002)

Bottom Line: Here by binding human secretory component to overlapping decapeptides of Calpha3, we confirm these residues and also uncover an additional site.Some transcytosis-selected peptides map to the same 402-410 pIgR-binding Calpha3 site.Further in vivo studies document that at least one of these peptides is transported in a rat model measuring hepatic bile transport.

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunogenetics Program, Oklahoma Medical Research Foundation, 825 NE 13th Street, Oklahoma City, OK 73104, USA.

ABSTRACT
Polymeric immunoglobulins provide first line humoral defense at mucosal surfaces to which they are specifically transported by the polymeric immunoglobulin receptor (pIgR) on mucosal and glandular epithelial cells. Previous studies from our laboratory suggested that amino acids 402-410 of the Calpha3 domain of dimeric IgA (dIgA) represented a potential binding site for the pIgR. Here by binding human secretory component to overlapping decapeptides of Calpha3, we confirm these residues and also uncover an additional site. Furthermore, we show that the Calpha3 motif appears to be sufficient to direct transport of green fluorescent protein through the pIgR-specific cellular transcytosis system. An alternative approach identified phage peptides, selected from a library by the in vitro Madin Darby Canine Kidney transcytosis assay, for pIgR-mediated transport through epithelial cells. Some transcytosis-selected peptides map to the same 402-410 pIgR-binding Calpha3 site. Further in vivo studies document that at least one of these peptides is transported in a rat model measuring hepatic bile transport. In addition to identifying small peptides that are both bound and transported by the pIgR, this study provides evidence that the pIgR-mediated mucosal secretion system may represent a means of targeting small molecule therapeutics and genes to mucosal epithelial cells.

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Reactivity of human pIgR with Cα3 decapeptides overlapping by eight amino acids. (a) Background reactivity of the decapeptides with pIgR detection reagents sheep anti-pIgR and donkey anti–sheep IgG alone. (b) Interaction of the decapeptides with human SC detected with sheep anti-pIgR and donkey anti–sheep IgG alkaline phosphatase. Results are means from three assays.
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fig1: Reactivity of human pIgR with Cα3 decapeptides overlapping by eight amino acids. (a) Background reactivity of the decapeptides with pIgR detection reagents sheep anti-pIgR and donkey anti–sheep IgG alone. (b) Interaction of the decapeptides with human SC detected with sheep anti-pIgR and donkey anti–sheep IgG alkaline phosphatase. Results are means from three assays.

Mentions: We previously reported the localization of an important pIgR-binding site on dimeric human IgA to a predicted loop structure in the Cα3 domain (10). To further localize this interaction, the binding of pIgR to overlapping decapeptides comprising Cα3 was evaluated. When the Cα3 decapeptides bound to pins were screened for reactivity with the polyclonal anti-SC serum and anti–sheep IgG conjugate, no significant background was demonstrated (Fig. 1 a). However, screening the peptides with SC identified two pIgR-reactive regions within the sequence of Cα3, peptide numbers 35–36 and 48–50 (Fig. 1 b). Decapeptide numbers 35–36 correspond to amino acids 404–415 of Cα3 and decapeptides 48–50 correspond to amino acids 430–443. Previous phage display selection of random peptides with cell-expressed pIgR coupled with subsequent alanine substitution provided evidence for amino acids 402–410 of Cα3 in dIgA being involved in the interaction with pIgR (10). The current results for decapeptides 35–36 correspond to the same region in Cα3 and thus provide further evidence for the importance of this specific interaction. When the region of Cα3 identified by decapeptides 48–50 (Cα3 amino acids 430–443) are compared with the corresponding region of Cγ3 (amino acids 424–437), part of the region would be predicted to form a loop structure and thus be accessible for ligand binding based on the crystal structure of IgG Fc (14). Therefore, amino acids 430–443 of Cα3 may represent another part of dIgA that cooperates with the 402–410 amino acid region for the initial high affinity interaction of dIgA with pIgR.


Targeting mucosal sites by polymeric immunoglobulin receptor-directed peptides.

White KD, Capra JD - J. Exp. Med. (2002)

Reactivity of human pIgR with Cα3 decapeptides overlapping by eight amino acids. (a) Background reactivity of the decapeptides with pIgR detection reagents sheep anti-pIgR and donkey anti–sheep IgG alone. (b) Interaction of the decapeptides with human SC detected with sheep anti-pIgR and donkey anti–sheep IgG alkaline phosphatase. Results are means from three assays.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196054&req=5

fig1: Reactivity of human pIgR with Cα3 decapeptides overlapping by eight amino acids. (a) Background reactivity of the decapeptides with pIgR detection reagents sheep anti-pIgR and donkey anti–sheep IgG alone. (b) Interaction of the decapeptides with human SC detected with sheep anti-pIgR and donkey anti–sheep IgG alkaline phosphatase. Results are means from three assays.
Mentions: We previously reported the localization of an important pIgR-binding site on dimeric human IgA to a predicted loop structure in the Cα3 domain (10). To further localize this interaction, the binding of pIgR to overlapping decapeptides comprising Cα3 was evaluated. When the Cα3 decapeptides bound to pins were screened for reactivity with the polyclonal anti-SC serum and anti–sheep IgG conjugate, no significant background was demonstrated (Fig. 1 a). However, screening the peptides with SC identified two pIgR-reactive regions within the sequence of Cα3, peptide numbers 35–36 and 48–50 (Fig. 1 b). Decapeptide numbers 35–36 correspond to amino acids 404–415 of Cα3 and decapeptides 48–50 correspond to amino acids 430–443. Previous phage display selection of random peptides with cell-expressed pIgR coupled with subsequent alanine substitution provided evidence for amino acids 402–410 of Cα3 in dIgA being involved in the interaction with pIgR (10). The current results for decapeptides 35–36 correspond to the same region in Cα3 and thus provide further evidence for the importance of this specific interaction. When the region of Cα3 identified by decapeptides 48–50 (Cα3 amino acids 430–443) are compared with the corresponding region of Cγ3 (amino acids 424–437), part of the region would be predicted to form a loop structure and thus be accessible for ligand binding based on the crystal structure of IgG Fc (14). Therefore, amino acids 430–443 of Cα3 may represent another part of dIgA that cooperates with the 402–410 amino acid region for the initial high affinity interaction of dIgA with pIgR.

Bottom Line: Here by binding human secretory component to overlapping decapeptides of Calpha3, we confirm these residues and also uncover an additional site.Some transcytosis-selected peptides map to the same 402-410 pIgR-binding Calpha3 site.Further in vivo studies document that at least one of these peptides is transported in a rat model measuring hepatic bile transport.

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunogenetics Program, Oklahoma Medical Research Foundation, 825 NE 13th Street, Oklahoma City, OK 73104, USA.

ABSTRACT
Polymeric immunoglobulins provide first line humoral defense at mucosal surfaces to which they are specifically transported by the polymeric immunoglobulin receptor (pIgR) on mucosal and glandular epithelial cells. Previous studies from our laboratory suggested that amino acids 402-410 of the Calpha3 domain of dimeric IgA (dIgA) represented a potential binding site for the pIgR. Here by binding human secretory component to overlapping decapeptides of Calpha3, we confirm these residues and also uncover an additional site. Furthermore, we show that the Calpha3 motif appears to be sufficient to direct transport of green fluorescent protein through the pIgR-specific cellular transcytosis system. An alternative approach identified phage peptides, selected from a library by the in vitro Madin Darby Canine Kidney transcytosis assay, for pIgR-mediated transport through epithelial cells. Some transcytosis-selected peptides map to the same 402-410 pIgR-binding Calpha3 site. Further in vivo studies document that at least one of these peptides is transported in a rat model measuring hepatic bile transport. In addition to identifying small peptides that are both bound and transported by the pIgR, this study provides evidence that the pIgR-mediated mucosal secretion system may represent a means of targeting small molecule therapeutics and genes to mucosal epithelial cells.

Show MeSH