Cellular and Subcellular Distribution of Polycystin-2, the Protein Product of the PKD2 Gene LUKAS FOGGENSTEINER, † A. PAUL BEVAN , * RUTH THOMAS,* NICHOLAS COLEMAN, ‡ CATHERINE BOULTER, § JOHN BRADLEY, † OXANA IBRAGHIMOV-BESKROVNAYA,  KATHERINE KLINGER,  and RICHARD SANDFORD* Departments of *Medical Genetics, † Medicine, and ‡ Pathology, University of Cambridge, Addenbrooke’s Hospital, Cambridge, United Kingdom; § Department of Genetics, University of Cambridge, Cambridge, United Kingdom; and  Genzyme Genetics, Framingham, Massachusetts. Abstract. Mutations in the PKD1 and PKD2 genes account for 85 and 15% of cases of autosomal dominant polycystic kidney disease, respectively. Polycystin-2, the product of the PKD2 gene, is predicted to be an integral membrane protein with homology to a family of voltage-activated Ca 2+ channels. In vitro studies suggest that it may interact with polycystin-1, the PKD1 gene product, via coiled-coil domains present in their C-terminal domains. In this study, the cellular and subcellular distribution of polycystin-2 is defined and compared with polycystin-1. A panel of rabbit polyclonal antisera was raised against polycystin-2 and shown to recognize a single band consistent with polycystin-2 in multiple tissues and cell lines by immunoprecipitation and Western blotting. Immunostaining of human and murine renal tissues demonstrated widespread and developmentally regulated expression of polycytin-2, with highest levels in the kidney in the thick ascending limbs of the loop of Henle and the distal convoluted tubule. In contrast, polycystin-1 expression, while localizing to the same tubular segments, was highest in the collecting ducts. Immunohisto- chemical staining and immunofluorescence microscopy local- ized polycystin-2 to the basolateral plasma membrane of kid- ney tubular epithelial cells compared with the junctional localization of polycystin-1. Differences in the developmental, cellular, and subcellular expression of polycystin-1 and poly- cystin-2 suggest that they may be able to function indepen- dently of each other in addition to a potential in vivo interaction via their C-termini. Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common inherited disorders of humans, with a prevalence of approximately 1 in 800 in all ethnic groups (1). It is characterized by progressive bilateral renal cyst formation, which results in a gradual decline in renal function such that half of affected individuals require renal replacement therapy by their sixth decade (2). Eight percent of all individuals in renal replacement programs have a diagnosis of ADPKD. Cysts are also seen in the liver and more rarely in the pancreas. Other more variable features of the disease include cardiovas- cular and connective tissue abnormalities such as hypertension, mitral valve prolapse, berry aneurysms of the cerebral circu- lation, and abdominal wall hernias (3). Eighty-five percent of cases of ADPKD are due to mutations in PKD1 with the remainder occurring in PKD2 (4). A few ADPKD families appear unlinked to either PKD1 or PKD2, suggesting the presence of a rare third locus, PKD3 (5). While mutations in PKD2 cause a milder phenotype than those in PKD1 with a slower decline in renal function, the clinical features are indistinguishable between the two groups (6). Targeted mutations in the murine pkd1 and pkd2 genes also produce a cystic phenotype, suggesting that the protein prod- ucts of these two genes, polycystin-1 and polycystin-2, may interact directly or form separate parts of a common cellular pathway (7,8). Polycystin-1 is composed of a novel array of structural and functional domains that suggest it is an integral plasma mem- brane glycoprotein involved in cell-cell or cell-matrix interac- tions (9,10). Its precise function is still unknown. A consensus derived from the many published reports of the cellular and subcellular localization of polycystin-1 defines it as a devel- opmentally regulated integral membrane protein expressed at highest levels in fetal tissues (11–15). In adult tissues it is predominantly localized to the distal part of the nephron and collecting ducts and other epithelial structures such as bile and pancreatic ducts (13,15). Polycystin-2 is predicted to be an integral membrane protein with intracellular N- and C-termini (16). It has significant homology to the pore-forming domains of a number of voltage- activated cation channels, suggesting that it may function as an ion channel subunit (16,17). A new member of the polycystin family, polycystin-L/polycystin-2L, has recently been de- scribed (17,18). It has extensive homology to polycystin-2 but Received June 4, 1999. Accepted September 21, 1999. Correspondence to Dr. Richard Sandford, Department of Medical Genetics, Cambridge Institute for Medical Research, Addenbrooke’s Hospital, Hills Road, Cambridge CB2 2XY, United Kingdom. Phone: +44 1223 762616; Fax: +44 1223 331206; E-mail: rns13@cam.ac.uk 1046-6673/1105-0814 Journal of the American Society of Nephrology Copyright © 2000 by the American Society of Nephrology J Am Soc Nephrol 11: 814 – 827, 2000