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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.

Show MeSH
Phage peptides selected by transcytosis and frequency of selection. The in vitro MDCK-pIgR transcytosis system was used to identify phage peptides that were transported from the basolateral medium to the apical medium. Two experiments used a 40-mer random peptide library (experiments no. 1 and no. 2) and one experiment used a 20-mer random peptide library (experiment no. 3). Phage peptides are referred to by the first three amino acids of their sequence.
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fig3: Phage peptides selected by transcytosis and frequency of selection. The in vitro MDCK-pIgR transcytosis system was used to identify phage peptides that were transported from the basolateral medium to the apical medium. Two experiments used a 40-mer random peptide library (experiments no. 1 and no. 2) and one experiment used a 20-mer random peptide library (experiment no. 3). Phage peptides are referred to by the first three amino acids of their sequence.

Mentions: It is conceivable that other structures, including smaller peptides, could substitute for polymeric immunoglobulins in the pIgR-mediated transcytosis process. To identify such molecules, the MDCK transcytosis system was used to select random phage-displayed peptides transcytosed in vitro. Three independent MDCK-pIgR transcytosis system phage display selection experiments were performed. Two experiments used a 40-mer random peptide library (experiments no. 1 and no. 2) and one experiment used a 20-mer random peptide library (experiment no. 3). Both phage libraries were constructed as pIII peptide fusions as this results in low number of peptides per phage being expressed and thus should allow selection of high affinity interactions. After eight rounds of selection, 20 clones from experiment no. 1 and 18 clones from experiments no. 2 and no. 3 were chosen at random and sequenced. Shown in Fig. 3 are the eight phage peptides selected from the three experiments and the frequency of selection of each phage clone.


Targeting mucosal sites by polymeric immunoglobulin receptor-directed peptides.

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

Phage peptides selected by transcytosis and frequency of selection. The in vitro MDCK-pIgR transcytosis system was used to identify phage peptides that were transported from the basolateral medium to the apical medium. Two experiments used a 40-mer random peptide library (experiments no. 1 and no. 2) and one experiment used a 20-mer random peptide library (experiment no. 3). Phage peptides are referred to by the first three amino acids of their sequence.
© Copyright Policy
Related In: Results  -  Collection

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

fig3: Phage peptides selected by transcytosis and frequency of selection. The in vitro MDCK-pIgR transcytosis system was used to identify phage peptides that were transported from the basolateral medium to the apical medium. Two experiments used a 40-mer random peptide library (experiments no. 1 and no. 2) and one experiment used a 20-mer random peptide library (experiment no. 3). Phage peptides are referred to by the first three amino acids of their sequence.
Mentions: It is conceivable that other structures, including smaller peptides, could substitute for polymeric immunoglobulins in the pIgR-mediated transcytosis process. To identify such molecules, the MDCK transcytosis system was used to select random phage-displayed peptides transcytosed in vitro. Three independent MDCK-pIgR transcytosis system phage display selection experiments were performed. Two experiments used a 40-mer random peptide library (experiments no. 1 and no. 2) and one experiment used a 20-mer random peptide library (experiment no. 3). Both phage libraries were constructed as pIII peptide fusions as this results in low number of peptides per phage being expressed and thus should allow selection of high affinity interactions. After eight rounds of selection, 20 clones from experiment no. 1 and 18 clones from experiments no. 2 and no. 3 were chosen at random and sequenced. Shown in Fig. 3 are the eight phage peptides selected from the three experiments and the frequency of selection of each phage clone.

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