title: Quality control of photosystem II. Cleavage of reaction center D1 protein in spinach thylakoids by FtsH protease under moderate heat stress.
abstract: When spinach thylakoids were subjected to moderate heat stress (40 degrees C for 30 min), oxygen evolution was inhibited, and cleavage of the reaction center-binding protein D1 of photosystem II took place, producing 23-kDa N-terminal fragments. The D1 cleavage was greatly facilitated by the addition of 0.15 mM ZnCl2 and 1 mM ATP and was completely inhibited by 1 mM EDTA, indicating the participation of an ATP-dependent metalloprotease(s) in the D1 cleavage. Herbicides 3-(3,4-dichlorophenyl)-1,1-dimethyl urea, bromoxynil, and ioxynil, all of which bind to the Q(B) site, inhibited the D1 cleavage, suggesting that the DE-loop of the D1 protein is the heat-sensitive cleavage site. We solubilized the protease by treating the thylakoids with 2 M KSCN and detected a protease activity in the supernatant by gelatin activity gel electrophoresis in the 70-80-kDa region. The antibodies against tobacco FtsH and Arabidopsis FtsH2 reacted with a 70-80-kDa band of the KSCN-solubilized fraction, which suggests the presence of FtsH in the fraction. In accordance with this finding, we identified the homolog to Arabidopsis FtsH8 in the 70-80-kDa region by matrix-assisted laser desorption ionization time-of-flight mass analysis of the thylakoids. The KSCN-solubilized fraction was successively reconstituted with thylakoids to show heat-induced cleavage of the D1 protein and production of the D1 fragment. These results strongly suggest that an FtsH protease(s) is involved in the primary cleavage of the D1 protein under moderate heat stress.
authors.affiliation: Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530.
authors.forename: Miho
authors.initials: M
authors.lastname: Yoshioka
authors.affiliation: Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530.
authors.forename: Suguru
authors.initials: S
authors.lastname: Uchida
authors.affiliation: School of Pharmacy, Shujitsu University, Okayama 703-8516, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Mori
authors.affiliation: Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530.
authors.forename: Keisuke
authors.initials: K
authors.lastname: Komayama
authors.affiliation: Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Ohira
authors.affiliation: Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530.
authors.forename: Noriko
authors.initials: N
authors.lastname: Morita
authors.affiliation: School of Pharmacy, Shujitsu University, Okayama 703-8516, Japan.
authors.forename: Tohru
authors.initials: T
authors.lastname: Nakanishi
authors.affiliation: Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530. Electronic address: yasusiya@cc.okayama-u.ac.jp.
authors.forename: Yasusi
authors.initials: Y
authors.lastname: Yamamoto
mesh_terms: D049069:ATP-Dependent Proteases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000378:metabolism*; D006358:Hot Temperature; D008565:Membrane Proteins / Q000378:metabolism*; D010446:Peptide Fragments / Q000096:biosynthesis; D045332:Photosystem II Protein Complex / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D018724:Spinacia oleracea; D020524:Thylakoids / Q000737:chemistry* / Q000378:metabolism; D015032:Zinc / Q000494:pharmacology
chemical_list: D049069:ATP-Dependent Proteases; D000255:Adenosine Triphosphate; D029681:Arabidopsis Proteins; D008565:Membrane Proteins; D010446:Peptide Fragments; D045332:Photosystem II Protein Complex; D010940:Plant Proteins; D015032:Zinc; C513982:FtsH8 protein, Arabidopsis; C402651:AT2G30950 protein, Arabidopsis