Polycystin-1 Interacts with Intermediate Filaments

Abstract

Polycystin-1, the protein defective in a majority of patients with autosomal dominant polycystic kidney disease, is a ubiquitously expressed multi-span transmembrane protein of unknown function. Subcellular localization studies found this protein to be a component of various cell junctional complexes and to be associated with the cytoskeleton, but the specificity and nature of such associations are not known. To identify proteins that interact with the polycystin-1 C-tail (P1CT), this segment was used as bait in a yeast two-hybrid screening of a kidney epithelial cell library. The intermediate filament (IF) protein vimentin was identified as a strong polycystin-1-interacting partner. Cytokeratins K8 and K18 and desmin were also found to interact with P1CT. These interactions were mediated by coiled-coil motifs in polycystin-1 and IF proteins. Vimentin, cytokeratins K8 and K18, and desmin also bound directly to P1CT in GST pull-down and in in vitro filament assembly assays. Two observations confirmed these interactions in vivo: (i) a cell membrane-anchored form of recombinant P1CT decorated the IF network and was found to associate with the cytoskeleton in detergent-solubilized cells and (ii) endogenous polycystin-1 distributed with IF at desmosomal junctions. Polycystin-1 may utilize this association for structural, storage, or signaling functions. Autosomal dominant polycystic kidney disease (ADPKD)1 is one of the most common genetic diseases in humans, affecting ϳ1 in 400 individuals and accounting for 8 -10% of end stage renal disease (1). ADPKD is characterized by the formation and progressive expansion of cysts in ductal organs including both kidneys as well as by cardiovascular abnormalities. About 85% of ADPKD cases are caused by defects in polycystin-1 (2, 3), a broadly expressed and developmentally regulated multi-span transmembrane protein with a large extracellular region rich in protein-protein and protein-carbohydrate interaction modules. Although this composition is suggestive of a role in cellcell and/or cell-matrix adhesion, the precise function(s) mediated by polycystin-1 remains unknown.Genetic, immunochemical, and biochemical studies have suggested that polycystin-1 is involved in maintaining the structural integrity of cell-cell junctions in vascular and epithelial structures. In mouse models of ADPKD, mouse embryos lacking a functional polycystin-1 are uniformly edematous (4, 5) and die in utero of hemorrhages preceded by vascular leaks (4). Consistent with these data, polycystin-1 has been shown to distribute at apicolateral cell junctions in both epithelial and endothelial cells (6 -9). Immunolocalization and/or cell fractionation studies found polycystin-1 at tight (10), adherens (7), and desmosomal (9) junctions, at focal contacts (11), and in the cytoplasm. A fraction has also been found in the Triton X-100-insoluble cell fraction, reflecting a strong association with cytoskeleton (12). Recently polycystin-1 was shown to be transported through a tuberin-dependent pathway to the lateral ...

Pkd1 Haploinsufficiency Increases Renal Damage and Induces Microcyst Formation following Ischemia/Reperfusion

“…The involvement of collecting tubules in the IR injury of Pkd1-haploinsufficient mice is not surprising, because PC1 has been shown to be particularly expressed in the distal nephron and collecting tubules. 32,33 Pkd1 haploinsufficiency, therefore, magnifies the morphologic injury determined by IR up to 14 d after the insult. This conclusion is also supported by the immunoelectron microscopy analysis, which revealed more severe proximal tubular and endothelial injuries in HTs.…”

Intermediate filaments: A role in epithelial polarity

“…Although an equivalent mechanism for keratin IFs has not been described, it remains a potential mechanism to be tested in future experiments. Another possibility is that membrane proteins may bind directly to IFs, which may be the case for polycystin-1 [90], a protein that, in addition to its well-known apical cilium localization, also concentrates around desmosomes [91]. Other examples are the interaction between aquaporin and filensin [92] and a 334-aa cancerassociated protein with the structure of a membrane protein that colocalizes with IFs [93].…”

Ciliary dysfunction in polycystic kidney disease: an emerging model with polarizing potential

“…The trafficking of polycystin-1 to the lateral membranes may be due, in part, to a dependence on tuberin (88,93). Furthermore, polycystin-1 trafficking to the plasma membrane is dependent upon the cytoskeleton as the COOH terminal-tail directly interacts with intermediate filaments (94). Polycystin-1 also has the capacity to bind to the extracellular matrix components such as collagen I, laminin, fibronectin, and integrin (85,95,96).…”