The carboxyl terminus of Neph family members binds to the PDZ domain protein zonula occludens-1.
The PSD95/Dlg/ZO-1 (PDZ) domain-containing protein zonula occludens-1 (ZO-1) selectively localizes to the cytoplasmic basis of the slit diaphragm, a specialized cell-cell contact in between glomerular podocytes necessary to prevent the loss of protein in the urine. However, the function of ZO-1 at the slit diaphragm has remained elusive. Deletion of Neph1, a slit diaphragm protein of the immunoglobulin superfamily with a cytoplasmic PDZ binding site, causes proteinuria in mice. We demonstrate now that Neph1 binds ZO-1. This interaction was mediated by the first PDZ domain of ZO-1 and involved the conserved PDZ domain binding motif present in the carboxyl terminus of the three known Neph family members. Furthermore, Neph1 co-immunoprecipitates with ZO-1 from lysates of mouse kidneys, demonstrating that this interaction occurs in vivo. Both deletion of the PDZ binding motif of Neph1 as well as threonine-to-glutamate mutation of the threonine within the binding motif abrogated binding of ZO-1, suggesting that phosphorylation may regulate this interaction. ZO-1 binding was associated with a strong increase in tyrosine phosphorylation of the cytoplasmic tail of Neph1 and dramatically accelerated the ability of Neph1 to induce signal transduction. Thus, our data suggest that ZO-1 may organize Neph proteins and recruit signal transduction components to the slit diaphragm of podocytes.
Huber
Tobias B
TB
Renal Division, University Hospital Freiburg, Hugstetter Str. 55, 79106 Freiburg, Germany.
Schmidts
Miriam
M
Gerke
Peter
P
Schermer
Bernhard
B
Zahn
Anne
A
Hartleben
Björn
B
Sellin
Lorenz
L
Walz
Gerd
G
Benzing
Thomas
T
D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D017353:Gene Deletion; D006801:Humans; D007668:Kidney / Q000502:physiology; D008565:Membrane Proteins / Q000737:chemistry* / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016297:Mutagenesis, Site-Directed; D010750:Phosphoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011507:Proteinuria / Q000235:genetics; D017384:Sequence Deletion; D015398:Signal Transduction; D062826:Zonula Occludens-1 Protein
C431137:KIRREL1 protein, human; C486606:Kirrel1 protein, mouse; D008565:Membrane Proteins; D010750:Phosphoproteins; C569042:TJP1 protein, human; C569043:Tjp1 protein, mouse; D062826:Zonula Occludens-1 Protein