Identification and characterization of polycystin-2, the PKD2 gene product

PKD2, the second gene for the autosomal dominant polycystic kidney disease (ADPKD), encodes a protein, polycystin-2, with predicted structural similarity to cation channel subunits. However, the function of polycystin-2 remains unknown. We used polyclonal antisera specific for the intracellular NH₂ and COOH termini to identify polycystin-2 as an ~110-kDa integral membrane glycoprotein. Polycystin-2 from both native tissues and cells in culture is sensitive to Endo H suggesting the continued presence of high- mannose oligosaccharides typical of pre-middle Golgi proteins. Immunofluorescent cell staining of polycystin-2 shows a pattern consistent with localization in the endoplasmic reticulum. This finding is confirmed by co-localization with protein-disulfide isomerase as determined by double indirect immunofluorescence and co-distribution with calnexin in subcellular fractionation studies. Polycystin-2 translation products truncated at or after Gly⁸²¹ retain their exclusive endoplasmic reticulum localization while products truncated at or before Glu⁷⁸⁷ additionally traffic to the plasma membrane. Truncation mutants that traffic to the plasma membrane acquire Endo H resistance and can be biotinylated on the cell surface in intact cells. The 34-amino acid region Glu⁷⁸⁷-Ser⁸²⁰, containing two putative phosphorylation sites, is responsible for the exclusive endoplasmic reticulum localization of polycystin-2 and is the site of specific interaction with an as yet unidentified protein binding partner for polycystin-2. The localization of full-length polycystin-2 to intracellular membranes raises the possibility that the PKD2 gene product is a subunit of intracellular channel complexes.