Structural characterization of the mechanosensitive channel candidate MCA2 from Arabidopsis thaliana.
Mechanosensing in plants is thought to be governed by sensory complexes containing a Ca²⁺-permeable, mechanosensitive channel. The plasma membrane protein MCA1 and its paralog MCA2 from Arabidopsis thaliana are involved in mechanical stress-induced Ca²⁺ influx and are thus considered as candidates for such channels or their regulators. Both MCA1 and MCA2 were functionally expressed in Sf9 cells using a baculovirus system in order to elucidate their molecular natures. Because of the abundance of protein in these cells, MCA2 was chosen for purification. Purified MCA2 in a detergent-solubilized state formed a tetramer, which was confirmed by chemical cross-linking. Single-particle analysis of cryo-electron microscope images was performed to depict the overall shape of the purified protein. The three-dimensional structure of MCA2 was reconstructed at a resolution of 26 Å from 5,500 particles and appears to comprise a small transmembrane region and large cytoplasmic region.
Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, Okazaki, Aichi, Japan.
Hideki
H
Shigematsu
Biomembrane Laboratory, Tokyo Metropolitan Institute of Medical Science, Setagaya-ku, Tokyo, Japan.
Kazuko
K
Iida
Department of Biology, Tokyo Gakugei University, Koganei, Tokyo, Japan.
Masataka
M
Nakano
Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, Okazaki, Aichi, Japan.
Pratima
P
Chaudhuri
Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, Okazaki, Aichi, Japan ; Department of Biology, Tokyo Gakugei University, Koganei, Tokyo, Japan.
Hidetoshi
H
Iida
Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, Okazaki, Aichi, Japan ; National Institute for Physiological Sciences, National Institutes of Natural Sciences, Okazaki, Aichi, Japan ; Department of Physiological Sciences, Graduate University for Advanced Studies, Okazaki, Aichi, Japan.
Kuniaki
K
Nagayama
D000818:Animals; D017360:Arabidopsis / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D020285:Cryoelectron Microscopy; D040542:Mechanotransduction, Cellular / Q000235:genetics*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae; D061987:Sf9 Cells; D018411:Spodoptera
D029681:Arabidopsis Proteins; D015220:Calcium Channels; C520286:MCA1 protein, Arabidopsis; C000591896:MCA2 protein, Arabidopsis; D008565:Membrane Proteins