A Critical Role for the Var2 FtsH Homologue of Arabidopsis thaliana in the Photosystem II Repair Cycle in Vivo

Abstract

Using a var2-2 mutant of Arabidopsis thaliana, which lacks a homologue of the zinc-metalloprotease, FtsH, we demonstrate that this protease is required for the efficient turnover of the D1 polypeptide of photosystem II and protection against photoinhibition in vivo. We show that var2-2 leaves are much more susceptible to lightinduced photosystem II photoinhibition than wild-type leaves. Furthermore, the rate of photosystem II photoinhibition in untreated var2-2 leaves is equivalent to that of var2-2 and wild-type leaves, which have been treated with lincomycin, an inhibitor of the photosystem II repair cycle at the level of D1 synthesis. This is in contrast to untreated wild-type leaves, which show a much slower rate of photosystem II photoinhibition due to an efficient photosystem II repair cycle. The recovery of var2-2 leaves from photosystem II photoinhibition is also impaired relative to wild-type. Using Western blot analysis in the presence of lincomycin we show that the D1 polypeptide remains stable in leaves of the var2-2 mutant under photoinhibitory conditions that lead to D1 degradation in wild-type leaves and that the abundance of DegP2 is not affected by the var2-2 mutation. We conclude, therefore, that the Var2 FtsH homologue is required for the cleavage of the D1 polypeptide in vivo. In addition, we identify a conserved lumenal domain in Var2 that is unique to FtsH homologues from oxygenic phototrophs. The Photosystem II (PSII)1 complex is a large protein-pigment assembly that catalyzes the light-dependent oxidation of water to molecular oxygen in chloroplasts and cyanobacteria. At the core of PSII lies the D1/D2 heterodimer, which binds the pigments and co-factors necessary for primary photochemistry (1). The D1 polypeptide is also important because of its high rate of turnover (2). This high turnover rate is related to the vulnerability of PSII to light, with D1 being the main target for photoinactivation and subsequent damage. An efficient repair cycle for D1 is therefore of paramount importance in oxygenic phototrophs. When the rate of photoinactivation and damage of D1 exceeds the capacity for repair, photoinhibition occurs, resulting in a decrease in the maximum efficiency of PSII photochemistry.A key feature of the D1 repair cycle is the degradation of the damaged polypeptide. It is generally accepted that damaged D1 is initially cleaved at a site on the stromal loop between transmembrane helices D and E yielding a 23-kDa N-terminal fragment (3) and a 10-kDa C-terminal fragment (4). This cleavage step is believed to be initiated by structural changes within the D1 polypeptide (5), although the precise nature of the cleavage event remains unclear. One proposal is that the action of active oxygen species acts to cleave the D1 polypeptide during strong illumination (6). However, the temperature dependence of the process (7) and its sensitivity to protease inhibitors (8) indicates the involvement of enzymatic proteolysis by an unidentified protease. Following cleavage, the breakdown fragments of ...

Coordinated Regulation and Complex Formation of YELLOW VARIEGATED1 and YELLOW VARIEGATED2, Chloroplastic FtsH Metalloproteases Involved in the Repair Cycle of Photosystem II in Arabidopsis Thylakoid Membranes

“…Similarly, leaf variegation appears as a typical phenotype of FtsH deficiency in higher plants (Sakamoto, 2003). The loss of VAR1 or VAR2 is accompanied by a decline in PSII activity when plants are exposed to high light (Bailey et al, 2002;Sakamoto et al, 2002). In particular, one of the main substrates in PSII appears to be a photodamaged D1 protein of the PSII reaction center, although it is unknown how the protease recognizes the damaged D1 protein.…”

“…Therefore, we concluded that although several highly similar FtsHs are present in chloroplasts, only the loss of VAR1 or VAR2 is associated with leaf variegation. In addition to their typical variegated phenotype, var1 and var2 mutants are characterized by their lesser ability to deal with photoinhibition (Bailey et al, 2002;Sakamoto et al, 2002). To test the behavior of the other mutants in this respect, we measured the maximal photochemical efficiency of PSII as a ratio of variable to maximum fluorescence using leaf discs from ftsh1 , ftsh6 , and ftsh8 knockout mutants.…”

“…The maximum yield of photochemistry in PSII measured by chlorophyll fluorescence indicated that var1 and var2 are highly sensitive to photoinhibitory light exposure, suggesting that VAR1 and VAR2 play a critical role in the photoprotection of chloroplasts. Further analysis in vivo suggested that one of the substrates for VAR2 is a photodamaged D1 protein of the PSII reaction center (Bailey et al, 2002). Degradation of the D1 protein is a key process in the repair cycle of photodamaged PSII (Aro et al, 1993;Melis, 1999), and FtsH appears to act in concert with another protease, DegP (Itzhaki et al, 1998;Haußühl et al, 2001).…”

Functional implications on the mechanism of the function of photosystem II including water oxidation based on the structure of photosystem II

“…The well-known phenomenon of photoinhibition may be the reason for the lack of the central antenna domain in PSII. The D1 protein is degraded during the process of photodamage and repair in the PSII complex (see Aro et al 1993;Spetea et al 1999;Bailey et al (2002) and references therein). During repair, the whole PSII complex has to be opened up.…”

Photosystem II core phosphorylation and photosynthetic acclimation require two different protein kinases

“…1). Consequently, lack of the D1 protease impairs the PSII repair cycle 13 . To test whether suppression of PSII core phosphorylation also impairs PSII repair by changing the rate of D1 turnover, we investigated the inactivation of PSII under high light, the recovery of PSII activity and the degradation of D1.…”