Acid-induced off-response of PKD2L1 channel in Xenopus oocytes and its regulation by Ca(2.).
Polycystic kidney disease (PKD) protein 2 Like 1 (PKD2L1), also called transient receptor potential polycystin-3 (TRPP3), regulates Ca(2+)-dependent hedgehog signalling in primary cilia, intestinal development and sour tasting but with an unclear mechanism. PKD2L1 is a Ca(2+)-permeable cation channel that is activated by extracellular Ca(2+) (on-response) in Xenopus oocytes. PKD2L1 co-expressed with PKD protein 1 Like 3 (PKD1L3) exhibits extracellular acid-induced activation (off-response, i.e., activation following acid removal) but whether PKD1L3 participates in acid sensing remains unclear. Here we used the two-microelectrode voltage-clamp, site directed mutagenesis, Western blotting, reverse transcriptase-polymerase chain reaction (RT-PCR) and immunofluorescence, and showed that PKD2L1 expressed in oocytes exhibits sustained off-response currents in the absence of PKD1L3. PKD1L3 co-expression augmented the PKD2L1 plasma membrane localization but did not alter the observed properties of the off-response. PKD2L1 off-response was inhibited by an increase in intracellular Ca(2+). We also identified two intra-membrane residues aspartic acid 349 (D349) and glutamic acid 356 (E356) in the third transmembrane domain that are critical for PKD2L1 channel function. Our study suggests that PKD2L1 may itself sense acids and defines off-response properties in the absence of PKD1L3.
Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada.
Shaimaa
S
Hussein
Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada.
Wang
W
Zheng
Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada.
Chris
C
Dyte
Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada.
Qian
Q
Wang
Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada.
JungWoo
J
Yang
School of Life Sciences and Technology, Tongji University, Shanghai, China.
Fan
F
Zhang
Membrane Protein Disease and Cancer Research Center, Hubei University of Technology, Wuhan, China.
Jingfeng
J
Tang
School of Life Sciences and Technology, Tongji University, Shanghai, China.
Ying
Y
Cao
Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada.|Membrane Protein Disease and Cancer Research Center, Hubei University of Technology, Wuhan, China.
Xing-Zhen
XZ
Chen
D000143:Acids / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000737:chemistry / Q000378:metabolism*; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D006801:Humans; D007473:Ion Channels / Q000235:genetics / Q000378:metabolism; D007477:Ions / Q000737:chemistry / Q000378:metabolism; D008839:Microelectrodes; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D018408:Patch-Clamp Techniques; D012333:RNA, Messenger; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D014981:Xenopus / Q000254:growth & development / Q000378:metabolism*
D000143:Acids; D015220:Calcium Channels; D007473:Ion Channels; D007477:Ions; C550142:PKD1L3 protein, human; C117185:PKD2L1 protein, human; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D002118:Calcium