title: Role of amino-terminal half of the S4-S5 linker in type 1 ryanodine receptor (RyR1) channel gating.
abstract: The type 1 ryanodine receptor (RyR1) is a Ca(2+) release channel found in the sarcoplasmic reticulum of skeletal muscle and plays a pivotal role in excitation-contraction coupling. The RyR1 channel is activated by a conformational change of the dihydropyridine receptor upon depolarization of the transverse tubule, or by Ca(2+) itself, i.e. Ca(2+)-induced Ca(2+) release (CICR). The molecular events transmitting such signals to the ion gate of the channel are unknown. The S4-S5 linker, a cytosolic loop connecting the S4 and S5 transmembrane segments in six-transmembrane type channels, forms an α-helical structure and mediates signal transmission in a wide variety of channels. To address the role of the S4-S5 linker in RyR1 channel gating, we performed alanine substitution scan of N-terminal half of the putative S4-S5 linker (Thr(4825)-Ser(4829)) that exhibits high helix probability. The mutant RyR1 was expressed in HEK cells, and CICR activity was investigated by caffeine-induced Ca(2+) release, single-channel current recordings, and [(3)H]ryanodine binding. Four mutants (T4825A, I4826A, S4828A, and S4829A) had reduced CICR activity without changing Ca(2+) sensitivity, whereas the L4827A mutant formed a constitutive active channel. T4825I, a disease-associated mutation for malignant hyperthermia, exhibited enhanced CICR activity. An α-helical wheel representation of the N-terminal S4-S5 linker provides a rational explanation to the observed activities of the mutants. These results suggest that N-terminal half of the S4-S5 linker may form an α-helical structure and play an important role in RyR1 channel gating.
authors.affiliation: Department of Cellular and Molecular Pharmacology, Juntendo University Graduate School of Medicine, Tokyo 113-8421, Japan. Electronic address: takashim@juntendo.ac.jp.
authors.forename: Takashi
authors.initials: T
authors.lastname: Murayama
authors.affiliation: Department of Cellular and Molecular Pharmacology, Juntendo University Graduate School of Medicine, Tokyo 113-8421, Japan.
authors.forename: Nagomi
authors.initials: N
authors.lastname: Kurebayashi
authors.affiliation: Department of Cell Physiology, Nagoya City University Graduate School of Medical Sciences, Nagoya 467-8601, Japan.
authors.forename: Toshiharu
authors.initials: T
authors.lastname: Oba
authors.affiliation: Department of Pharmacology, School of Medicine, Showa University, Tokyo 142-8555, Japan.
authors.forename: Hideto
authors.initials: H
authors.lastname: Oyamada
authors.affiliation: Department of Pharmacology, School of Medicine, Showa University, Tokyo 142-8555, Japan.
authors.forename: Katsuji
authors.initials: K
authors.lastname: Oguchi
authors.affiliation: Department of Cellular and Molecular Pharmacology, Juntendo University Graduate School of Medicine, Tokyo 113-8421, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Sakurai
authors.affiliation: Department of Cellular and Molecular Pharmacology, Juntendo University Graduate School of Medicine, Tokyo 113-8421, Japan.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Ogawa
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D015640:Ion Channel Gating / Q000502:physiology*; D020125:Mutation, Missense; D017433:Protein Structure, Secondary; D011817:Rabbits; D019837:Ryanodine Receptor Calcium Release Channel / Q000235:genetics / Q000378:metabolism*
chemical_list: D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium