Identification and functional characterization of an N-terminal oligomerization domain for polycystin-2.
Autosomal dominant polycystic kidney disease (ADPKD), the most common inherited cause of kidney failure, is caused by mutations in either PKD1 (85%) or PKD2 (15%). The PKD2 protein, polycystin-2 (PC2 or TRPP2), is a member of the transient receptor potential (TRP) superfamily and functions as a non-selective calcium channel. PC2 has been found to form oligomers in native tissues suggesting that it may form functional homo- or heterotetramers with other subunits, similar to other TRP channels. Our experiments unexpectedly revealed that PC2 mutant proteins lacking the known C-terminal dimerization domain were still able to form oligomers and co-immunoprecipitate full-length PC2, implying the possible existence of a proximal dimerization domain. Using yeast two-hybrid and biochemical assays, we have mapped an alternative dimerization domain to the N terminus of PC2 (NT2-1-223, L224X). Functional characterization of this domain demonstrated that it was sufficient to induce cyst formation in zebrafish embryos and inhibit PC2 surface currents in mIMCD3 cells probably by a dominant-negative mechanism. In summary, we propose a model for PC2 assembly as a functional tetramer which depends on both C- and N-terminal dimerization domains. These results have significant implications for our understanding of PC2 function and disease pathogenesis in ADPKD and provide a new strategy for studying PC2 function.
Feng
Shuang
S
Academic Unit of Nephrology, Sheffield Kidney Institute, University of Sheffield, Sheffield S10 2RX, United Kingdom.
Okenka
Genevieve M
GM
Bai
Chang-Xi
CX
Streets
Andrew J
AJ
Newby
Linda J
LJ
DeChant
Brett T
BT
Tsiokas
Leonidas
L
Obara
Tomoko
T
Ong
Albert C M
AC
D000818:Animals; D019281:Dimerization; D004594:Electrophysiology / Q000379:methods; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D008954:Models, Biological; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D007690:Polycystic Kidney Diseases / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D050396:TRPP Cation Channels / Q000737:chemistry* / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D015027:Zebrafish
D050396:TRPP Cation Channels