Identification of receptor ligands by screening phage-display peptide libraries ex vivo on microdissected kidney tubules.
A novel method to identify receptor ligands for defined renal tubular segments has been developed. Ex vivo screening of phage-display peptide libraries on isolated intact rat proximal convoluted tubules (PCT) and cortical collecting ducts (CCD) allowed the direct access of phage to the basolateral surface of tubular epithelial cells. Two distinct peptide motifs were selected for CCD and PCT, indicating differential expression of some membrane receptors on the basolateral surface of defined kidney tubule segments. Using the linear peptide motif ELRGD(R/M)AX(W/L), recovered from freshly isolated rat CCD, mediated 16-fold selectivity of phage binding to CCD compared with PCT. Binding to CCD was 39-fold higher than that of a random control phage. Binding and subsequent internalization of phage, most likely by an integrin-mediated endocytosis pathway, was abolished by the addition of the corresponding synthetic peptide. Furthermore, the results demonstrate that presentation and flanking amino acids determine the specific binding properties of RGD ligands to their putative integrin receptors. The results emphasize the need of a native cell system for the identification of renal epithelial cell surface ligands. Such ligands are of potential relevance for the analysis of interactions between extracellular matrix and kidney tubules or for the development of improved vectors for kidney-specific drug delivery or gene transfer.
Odermatt
Alex
A
Division of Nephrology and Hypertension, Department of Clinical Research, Berne, Switzerland.
Audigé
Annette
A
Division of Nephrology and Hypertension, Department of Clinical Research, Berne, Switzerland.
Frick
Christoph
C
Division of Nephrology and Hypertension, Department of Clinical Research, Berne, Switzerland.
Vogt
Bruno
B
Division of Nephrology and Hypertension, Department of Clinical Research, Berne, Switzerland.
Frey
Brigitte M
BM
Division of Nephrology and Hypertension, Department of Clinical Research, Berne, Switzerland.
Frey
Felix J
FJ
Division of Nephrology and Hypertension, Department of Clinical Research, Berne, Switzerland.
Mazzucchelli
Luca
L
Institute of Pathology, University of Berne, Berne, Switzerland.
D000818:Animals; D003110:Colonic Neoplasms / Q000378:metabolism; D004210:Dissection; D007684:Kidney Tubules / Q000378:metabolism*; D008024:Ligands; D008297:Male; D019151:Peptide Library; D051381:Rats; D017207:Rats, Sprague-Dawley; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011971:Receptors, Immunologic / Q000378:metabolism; D018000:Receptors, Peptide / Q000378:metabolism
D008024:Ligands; D019151:Peptide Library; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D018000:Receptors, Peptide; C051382:arginyl-glycyl-aspartic acid directed cell adhesion receptor