title: A novel PKD2L1 C-terminal domain critical for trimerization and channel function.
abstract: As a transient receptor potential (TRP) superfamily member, polycystic kidney disease 2-like-1 (PKD2L1) is also called TRPP3 and has similar membrane topology as voltage-gated cation channels. PKD2L1 is involved in hedgehog signaling, intestinal development, and sour tasting. PKD2L1 and PKD1L3 form heterotetramers with 3:1 stoichiometry. C-terminal coiled-coil-2 (CC2) domain (G699-W743) of PKD2L1 was reported to be important for its trimerization but independent studies showed that CC2 does not affect PKD2L1 channel function. It thus remains unclear how PKD2L1 proteins oligomerize into a functional channel. By SDS-PAGE, blue native PAGE and mutagenesis we here identified a novel C-terminal domain called C1 (K575-T622) involved in stronger homotrimerization than the non-overlapping CC2, and found that the PKD2L1 N-terminus is critical for dimerization. By electrophysiology and Xenopus oocyte expression, we found that C1, but not CC2, is critical for PKD2L1 channel function. Our co-immunoprecipitation and dynamic light scattering experiments further supported involvement of C1 in trimerization. Further, C1 acted as a blocking peptide that inhibits PKD2L1 trimerization as well as PKD2L1 and PKD2L1/PKD1L3 channel function. Thus, our study identified C1 as the first PKD2L1 domain essential for both PKD2L1 trimerization and channel function, and suggest that PKD2L1 and PKD2L1/PKD1L3 channels share the PKD2L1 trimerization process.
authors.affiliation: Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, T6G 2H7, Edmonton, AB, Canada.
authors.forename: Wang
authors.initials: W
authors.lastname: Zheng
authors.affiliation: Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, T6G 2H7, Edmonton, AB, Canada.
authors.forename: Shaimaa
authors.initials: S
authors.lastname: Hussein
authors.affiliation: Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, T6G 2H7, Edmonton, AB, Canada.
authors.forename: JungWoo
authors.initials: J
authors.lastname: Yang
authors.affiliation: Department of Chemical and Materials Engineering, Faculty of Engineering, University of Alberta, T6G 2V4, Edmonton, AB, Canada.
authors.forename: Jun
authors.initials: J
authors.lastname: Huang
authors.affiliation: School of Life Sciences and Technology, Tongji University, 200092, Shanghai, China.
authors.forename: Fan
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Department of Biochemistry, Faculty of Medicine and Dentistry, University of Alberta, T6G 2H7, Edmonton, AB, Canada.
authors.forename: Samuel
authors.initials: S
authors.lastname: Hernandez-Anzaldo
authors.affiliation: Department of Biochemistry, Faculty of Medicine and Dentistry, University of Alberta, T6G 2H7, Edmonton, AB, Canada.
authors.forename: Carlos
authors.initials: C
authors.lastname: Fernandez-Patron
authors.affiliation: School of Life Sciences and Technology, Tongji University, 200092, Shanghai, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Cao
authors.affiliation: Department of Chemical and Materials Engineering, Faculty of Engineering, University of Alberta, T6G 2V4, Edmonton, AB, Canada.
authors.forename: Hongbo
authors.initials: H
authors.lastname: Zeng
authors.affiliation: Membrane Protein Disease and Cancer Research Center, Hubei University of Technology, 430068, Wuhan, Hubei, China.
authors.forename: Jingfeng
authors.initials: J
authors.lastname: Tang
authors.affiliation: 1] Membrane Protein Disease Research Group, Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, T6G 2H7, Edmonton, AB, Canada [2] Membrane Protein Disease and Cancer Research Center, Hubei University of Technology, 430068, Wuhan, Hubei, China.
authors.forename: Xing-Zhen
authors.initials: XZ
authors.lastname: Chen
mesh_terms: D015220:Calcium Channels / Q000737:chemistry* / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007473:Ion Channels / Q000737:chemistry* / Q000378:metabolism; D016296:Mutagenesis; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000378:metabolism
chemical_list: D015220:Calcium Channels; D007473:Ion Channels; C550142:PKD1L3 protein, human; C117185:PKD2L1 protein, human; D011956:Receptors, Cell Surface
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