Protein kinase D-mediated phosphorylation of polycystin-2 (TRPP2) is essential for its effects on cell growth and calcium channel activity.
PKD2 is mutated in 15% of patients with autosomal dominant polycystic kidney disease. The PKD2 protein, polycystin-2 or TRPP2, is a nonselective Ca2+-permeable cation channel that has been shown to function at several locations, including primary cilia, basolateral membrane, and at the endoplasmic reticulum (ER). Nevertheless, the factors that regulate the channel activity of polycystin-2 are not well understood. Polycystin-2 has been shown previously to be regulated by phosphorylation at two serine residues (Ser812 and Ser76) with distinct functional consequences. Here, we report the identification of a previously unrecognized phosphorylation site within the polycystin-2 C terminus (Ser801), and we demonstrate that it is phosphorylated by protein kinase D. Phosphorylation at this site was significantly increased in response to serum and epidermal growth factor stimulation. In nonciliated Madin-Darby canine kidney I cells, inducible expression of polycystin-2 inhibited cell proliferation compared with wild-type cells. Mutagenesis at Ser801 abolished these effects and reduced ATP-stimulated Ca2+ release from ER stores. Finally, we show that a pathogenic mutation (S804N) within the consensus kinase recognition sequence abolished Ser801 phosphorylation. These results suggest that growth factor-stimulated, protein kinase D-mediated phosphorylation of polycystin-2 is essential for its ER channel function and links extracellular stimuli to its effects on cell growth and intracellular calcium regulation.
Streets
Andrew J
AJ
Kidney Genetics Group, Academic Nephrology Unit, Sheffield Kidney Institute, University of Sheffield, Sheffield S10 2RX, United Kingdom.
Needham
Andrew J
AJ
Gill
Sharonjit K
SK
Ong
Albert C M
AC
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D049109:Cell Proliferation; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D057809:HEK293 Cells; D006801:Humans; D009154:Mutation; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D034622:RNA Interference; D012694:Serine / Q000235:genetics / Q000378:metabolism; D050396:TRPP Cation Channels / Q000235:genetics / Q000378:metabolism*
D000255:Adenosine Triphosphate; D002118:Calcium; D015220:Calcium Channels; D004815:Epidermal Growth Factor; D011493:Protein Kinase C; D012694:Serine; D050396:TRPP Cation Channels; C089212:protein kinase D