Hoffmeister, Helen and Gallagher, Anna-Rachel and Rascle, Anne and Witzgall, Ralph (2011) The human polycystin-2 protein represents an integral membrane protein with six membrane-spanning domains and intracellular N- and C-termini. BIOCHEMICAL JOURNAL, 433. pp. 285-294. ISSN 0264-6021, 1470-8728
Full text not available from this repository. (Request a copy)Abstract
PKD2 is one of the two genes mutated in ADPKD (autosomal-dominant polycystic kidney disease). The protein product of PKD2, polycystin-2, functions as a non-selective cation channel in the endoplasmic reticulum and possibly at the plasma membrane. Hydrophobicity plots and its assignment to the TRP (transient receptor potential) family of cation channels suggest that polycystin-2 contains six transmembrane domains and that both the N- and C-termini extend into the cytoplasm. However, no experimental evidence for this model has so far been provided. To determine the orientation of the different loops of polycystin-2, we truncated polycystin-2 within the predicted loops 1-5 and tagged Me constructs at the C-terminus with an HA (haemagglutinin) epitope. After transient expression and selective membrane permeabilization, immunofluorescence staining for the HA epitope revealed that loops 1, 3 and 5 extend into the lumen of the endoplasmic reticulum or the extracellular space, whereas loops 2 and 4 extend into the cytoplasm. This approach also confirmed the cytoplasmic orientation of the N- and C-termini of polycystin-2. In accordance with the immunofluorescence data, protease protection assays from microsomal preparations yielded protected fragments when polycystin-2 was truncated in loops 1, 3 and 5, whereas no protected fragments could be detected when polycystin-2 was truncated in loops 2 and 4. The results of the present study therefore provide the first experimental evidence for the topological orientation of polycystin-2.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | KIDNEY-DISEASE; PKD1 GENE; CHANNEL REGULATION; CILIA; LOCALIZES; MECHANISM; ENCODES; TRPP2; membrane topology; polycystin-2; transient receptor potential family |
| Subjects: | 500 Science > 570 Life sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Anatomie > Lehrstuhl für Molekulare und zelluläre Anatomie > Prof. Dr. Ralph Witzgall |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 29 Jun 2020 08:13 |
| Last Modified: | 29 Jun 2020 08:13 |
| URI: | https://pred.uni-regensburg.de/id/eprint/21397 |
Actions (login required)
 |
View Item |
