Cardiovascular, skeletal, and renal defects in mice with a targeted disruption of the Pkd1 gene

Autosomal dominant polycystic kidney disease (ADPKD) is characterized by cyst formation in the kidney, liver, and pancreas and is associated often with cardiovascular abnormalities such as hypertension, mitral valve prolapse, and intracranial aneurysms. It is caused by mutations in PKD1 or PKD2, encoding polycystin-1 and -2, which together form a cell surface nonselective cation ion channel. Pkd2-/- mice have cysts in the kidney and pancreas and defects in cardiac septation, whereas Pkd1del34 -/- and Pkd1L -/- mice have cysts but no cardiac abnormalities, although vascular fragility was reported in the latter. Here we describe mice carrying a targeted mutation in Pkd1 (Pkd1del17–21ßgeo), which defines its expression pattern by using a lacZ reporter gene and may identify novel functions for polycystin-1. Although Pkd1del17–21ßgeo +/- adult mice develop renal and hepatic cysts, Pkd1del17–21ßgeo -/- embryos die at embryonic days 13.5–14.5 from a primary cardiovascular defect that includes double outflow right ventricle, disorganized myocardium, and abnormal atrio-ventricular septation. Skeletal development is also severely compromised. These abnormalities correlate with the major sites of Pkd1 expression. During nephrogenesis, Pkd1 is expressed in maturing tubular epithelial cells from embryonic day 15.5. This expression coincides with the onset of cyst formation in Pkd1del34 -/-, Pkd1L -/-, and Pkd2-/- mice, supporting the hypothesis that polycystin-1 and polycystin-2 interact in vivo and that their failure to do so leads to abnormalities in tubule morphology and function. Autosomal dominant polycystic kidney disease (ADPKD) is a common inherited disorder that affects 1 in 800 people and accounts for ˜8% of patients with end-stage renal failure. It is characterized by the formation of multiple cysts in the kidneys and liver and, less frequently, in the pancreas. Cardiovascular abnormalities including hypertension, mitral valve prolapse, and intracranial aneurysms are also frequently recognized. Extensive characterization of the cellular defects in cyst-lining epithelial cells derived from kidneys affected by ADPKD and from a variety of rodent models of renal cystic disease has demonstrated generalized abnormalities in cell proliferation, differentiation, and apoptosis (1–3). More specific defects in cell polarity and extracellular matrix production are also seen and have been implicated directly in the process of cyst formation (4, 5). However, the primary events that give rise to this cystic phenotype have not been elucidated. The cloning of PKD1 and PKD2, the genes mutated in almost all cases of ADPKD, has provided the opportunity to investigate the molecular basis of cyst formation and develop appropriate model systems to study the functions of their protein products, polycystin-1 and -2. Both are predicted to be polytopic membrane proteins, with polycystin-1 having a large predicted extracellular region comprising multiple discrete domains while polycystin-2 has homology to ion channels (6, 7). An interaction mediated by the C-terminal regions of both proteins results in the formation of calcium-permeable nonselective cation channels in vitro, suggesting that extracellular signals can be transduced by the polycystin complex to regulate diverse cellular processes (8). Indeed, the cytoplasmic tail of polycystin-1 has been shown to activate several signal transduction pathways (9, 10), whereas Madin-Darby canine kidney cells transfected with PKD1 are resistant to apoptosis and undergo spontaneous tubulogenesis (11). The nature of the extracellular signals or protein ligands that activate polycystin-1 signaling have not been determined. The formation of a polycystin complex suggests that Pkd1 and Pkd2 should have considerable overlap in their expression patterns, and detailed analysis of the cellular and subcellular distribution has been performed. Expression of polycystin-2 has been defined in renal tubular epithelial cells with widespread expression reported in other tissues including the heart and vasculature (12–14). Unfortunately, considerable differences have been reported in the expression pattern of polycystin-1 by using both antibodies directed against different epitopes and RNA in situ hybridization (15–23). This has made meaningful comparisons of Pkd1 and Pkd2 expression difficult. Mice carrying targeted mutations in Pkd1 and Pkd2 or a PKD1 transgene have been reported (24–28). They all have renal cysts, suggesting that alterations in the level of polycystin-1 lead to cyst formation. Both Pkd1del34 -/- and Pkd1L -/- mice develop renal, hepatic, and pancreatic cystic disease. However Pkd1L -/- embryos also develop gross edema and s.c. hemorrhage, which may be caused by a defect in vascular wall integrity (24). Unlike these models, Pkd2 mutant mice also have major defects in cardiac development manifested by septal abnormalities in addition to the cystic phenotype (27). Here we describe a mouse model of ADPKD that allows the accurate description of Pkd1 expression by using a lacZ reporter gene and identifies a major function for polycystin-1 in cardiovascular and skeletal development in addition to its role in embryonic and adult kidney.