title: Different KChIPs compete for heteromultimeric assembly with pore-forming Kv4 subunits.
abstract: Auxiliary Kv channel-interacting proteins 1-4 (KChIPs1-4) coassemble with pore-forming Kv4 α-subunits to form channel complexes underlying somatodendritic subthreshold A-type current that regulates neuronal excitability. It has been hypothesized that different KChIPs can competitively bind to Kv4 α-subunit to form variable channel complexes that can exhibit distinct biophysical properties for modulation of neural function. In this study, we use single-molecule subunit counting by total internal reflection fluorescence microscopy in combinations with electrophysiology and biochemistry to investigate whether different isoforms of auxiliary KChIPs, KChIP4a, and KChIP4bl, can compete for binding of Kv4.3 to coassemble heteromultimeric channel complexes for modulation of channel function. To count the number of photobleaching steps solely from cell membrane, we take advantage of a membrane tethered k-ras-CAAX peptide that anchors cytosolic KChIP4 proteins to the surface for reduction of background noise. Single-molecule subunit counting reveals that the number of KChIP4 isoforms in Kv4.3-KChIP4 complexes can vary depending on the KChIP4 expression level. Increasing the amount of KChIP4bl gradually reduces bleaching steps of KChIP4a isoform proteins, and vice versa. Further analysis of channel gating kinetics from different Kv4-KChIP4 subunit compositions confirms that both KChIP4a and KChIP4bl can modulate the channel complex function upon coassembly. Taken together, our findings show that auxiliary KChIPs can heteroassemble with Kv4 in a competitive manner to form heteromultimeric Kv4-KChIP4 channel complexes that are biophysically distinct and regulated under physiological or pathological conditions.
authors.affiliation: Department of Neurobiology, Neuroscience Research Institute, Peking University Health Science Center, Beijing, China.
authors.forename: Jingheng
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Department of Molecular and Cellular Pharmacology, PKU-IDG/McGovern Institute for Brain Research, Peking University School of Pharmaceutical Sciences, Beijing, China; Qingdao University School of Pharmacy, Qingdao, China.
authors.forename: Yiquan
authors.initials: Y
authors.lastname: Tang
authors.affiliation: Department of Neurobiology, Neuroscience Research Institute, Peking University Health Science Center, Beijing, China.
authors.forename: Qin
authors.initials: Q
authors.lastname: Zheng
authors.affiliation: Department of Neurobiology, Neuroscience Research Institute, Peking University Health Science Center, Beijing, China.
authors.forename: Meng
authors.initials: M
authors.lastname: Li
authors.affiliation: Institute of Molecular Medicine, Peking University, Beijing, China.
authors.forename: Tianyi
authors.initials: T
authors.lastname: Yuan
authors.affiliation: Institute of Molecular Medicine, Peking University, Beijing, China.
authors.forename: Liangyi
authors.initials: L
authors.lastname: Chen
authors.affiliation: Department of Molecular and Cellular Pharmacology, PKU-IDG/McGovern Institute for Brain Research, Peking University School of Pharmaceutical Sciences, Beijing, China.
authors.forename: Zhuo
authors.initials: Z
authors.lastname: Huang
authors.affiliation: Department of Neurobiology, Neuroscience Research Institute, Peking University Health Science Center, Beijing, China; Department of Molecular and Cellular Pharmacology, PKU-IDG/McGovern Institute for Brain Research, Peking University School of Pharmaceutical Sciences, Beijing, China; Qingdao University School of Pharmacy, Qingdao, China. Electronic address: wangkw@bjmu.edu.cn.
authors.forename: KeWei
authors.initials: K
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D015640:Ion Channel Gating; D007700:Kinetics; D051605:Kv Channel-Interacting Proteins / Q000737:chemistry* / Q000378:metabolism*; D016062:Porosity; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry* / Q000378:metabolism; D051672:Shal Potassium Channels / Q000737:chemistry* / Q000378:metabolism; D014981:Xenopus
chemical_list: D051605:Kv Channel-Interacting Proteins; D021122:Protein Subunits; D051672:Shal Potassium Channels