Energy-dissipative supercomplex of photosystem II associated with LHCSR3 in Chlamydomonas reinhardtii

Abstract

Plants and green algae have a low pH-inducible mechanism in photosystem II (PSII) that dissipates excess light energy, measured as the nonphotochemical quenching of chlorophyll fluorescence (qE). Recently, nonphotochemical quenching 4 (npq4), a mutant strain of the green alga Chlamydomonas reinhardtii that is qE-deficient and lacks the light-harvesting complex stress-related protein 3 (LHCSR3), was reported [Peers G, et al. (2009) Nature 462(7272):518-521]. Here, applying a newly established procedure, we isolated the PSII supercomplex and its associated light-harvesting proteins from both WT C. reinhardtii and the npq4 mutant grown in either low light (LL) or high light (HL). LHCSR3 was present in the PSII supercomplex from the HLgrown WT, but not in the supercomplex from the LL-grown WT or mutant. The purified PSII supercomplex containing LHCSR3 exhibited a normal fluorescence lifetime at a neutral pH (7.5) by single-photon counting analysis, but a significantly shorter lifetime at pH 5.5, which mimics the acidified lumen of the thylakoid membranes in HL-exposed chloroplasts. The switch from light-harvesting mode to energy-dissipating mode observed in the LHCSR3-containing PSII supercomplex was sensitive to dicyclohexylcarbodiimide, a protein-modifying agent specific to protonatable amino acid residues. We conclude that the PSII-LHCII-LHCSR3 supercomplex formed in the HL-grown C. reinhardtii cells is capable of energy dissipation on protonation of LHCSR3.high light stress | light acclimation | photosynthesis | time-resolved fluorescence P hotosynthetic reactions in plants and algae fix CO 2 by converting solar energy into electrochemical energy. In nature, unexpected changes in light intensity could lead to overexcitation of the photosystems, resulting in the accumulation of harmful reactive oxygen species (1). Plants and green algae have developed protective nonphotochemical quenching (NPQ) mechanisms that alleviate such photo-oxidative stress. Among these mechanisms, quenching of chlorophyll (Chl) fluorescence (qE)-a feedback process of photosystem II (PSII) regulation-dissipates excess light energy captured by PSII as heat on luminal acidification of the thylakoid membranes, which occurs along with elevated electron flow.Numerous previous studies have focused on elucidating the molecular mechanism of qE quenching (see refs. 1-3 for reviews). In higher plants, qE induction depends on activation of the xanthophyll cycle (4-6) and the sensing of luminal acidification by PsbS, a protein homologous to light harvesting complex (LHC) proteins (7,8). Protonation of PsbS induces a change in the macroorganization of the thylakoid membranes (3, 9, 10) that results in the aggregation of LHCII proteins (11) and/or induces conformational change(s) in LHCII (12), allowing for the formation of energyquenching sites (13). Although both PsbS protein and zeaxanthin are thought to have crucial roles in qE quenching in higher plants, such molecular effectors have not been studied in depth in other photosynthetic organisms. The ...

PHOTOSYSTEM II SUBUNIT R Is Required for Efficient Binding of LIGHT-HARVESTING COMPLEX STRESS-RELATED PROTEIN3 to Photosystem II-Light-Harvesting Supercomplexes in Chlamydomonas reinhardtii

“…As the diminishment of PSBR expression impacts the magnitude of NPQ but not the extent of LHCSR3 induction, we rationalized that PSBR reduction might affect the formation of an LHCSR3-PSII supercomplex that was described previously (Tokutsu and Minagawa, 2013). To this end, C. reinhardtii cells (the wild type and ami-RNA-psbr-2) were grown in TAP LL and shifted to HL in HSM for 2 and 24 h. C. reinhardtii ami-RNA-psbr-1 cells were additionally analyzed after 24 h of HL.…”

“…S2). The SDG separation displayed three green bands for the wild type, designated as LHCII, PSI-LHCI, and PSII-LHCII according to preceding work (Tokutsu and Minagawa, 2013). For ami-RNApsbr-1 and ami-RNA-psbr-2 strains, visual inspection of the Suc gradients indicated that the PSII-LHCII bands were less pronounced in comparison with the wild-type SDG separation.…”

“…A recent study in Chlamydomonas reinhardtii identified PSII-LHCII supercomplexes with three LHCII trimers attached to each side of the core (C 2 S 2 M 2 L 2 ; Tokutsu et al, 2012). Interestingly, such PSII-LHCII supercomplexes associate with LIGHT-HARVESTING COMPLEX STRESS-RELATED PROTEIN3 (LHCSR3; Tokutsu and Minagawa, 2013), an ancient light-harvesting protein required for efficient energy-dependent (qE) quenching in the alga (Peers et al, 2009). The qE component of nonphotochemical quenching (NPQ) is an energy-dependent constituent of NPQ and regulates the thermal dissipation of excess absorbed light energy (Li et al, 2000;Peers et al, 2009).…”

Chlamydomonas reinhardtii PsbS Protein Is Functional and Accumulates Rapidly and Transiently under High Light

“…LhcSR3 is homologous to the Lhc proteins: it has a similar structure with three transmembrane helices and it is also able to bind Chls and Cars (Bonente et al, 2011). It has been suggested that LhcSR3 itself is the energy quencher due to its pigment binding ability, the unusual fluorescence properties that show very short lifetime components, and its pH sensitivity (Bonente et al, 2011;Liguori et al, 2013;Tokutsu and Minagawa, 2013). A recent investigation suggests the LhcSR is localized both in grana membranes and stroma lamellae in the moss Physcomitrella patens (Pinnola et al, 2015), and investigations on Chlamydomonas suggest that LhcSR3 can bind both to PSI or PSII depending on illumination conditions (Allorent et al, 2013;Bergner et al, 2015).…”

Light-harvesting antenna complexes in the moss Physcomitrella patens: implications for the evolutionary transition from green algae to land plants

“…In C. reinhardtii, high light stress-related LHC (LHCSR [44]; originally known as LI818) protein is protonated on its lumenal side upon buildup of DpH [45], which enhances the induction of qE within the PSII-LHCII-LHCSR supercomplex [46]. PSII subunit S (PsbS) was recently shown to be expressed transiently during the activation of qE and stabilize the accumulation of LHCSR, implying a pivotal role for PsbS in qE induction in C. reinhardtii [47 ].…”