Evidences for interaction of PsbS with photosynthetic complexes in maize thylakoids.

The PsbS subunit of Photosystem II (PSII) has received much attention in the past few years, given its crucial role in photoprotection of higher plants. The exact location of this small subunit in thylakoids is also debated. In this work possible interaction partners of PsbS have been identified by immunoaffinity and immunoprecipitation, performed with mildly solubilized whole thylakoid membrane. The interacting proteins, as identified by mass spectrometry analysis of the immunoaffinity eluate, include CP29, some LHCII components, but also components of Photosystem I, of the cytochrome b(6)f complex as well as of ATP synthase. These proteins can be co-immunoprecipitated by using highly specific anti-PsbS antibodies and, vice-versa, PsbS is co-immunoprecipitated by antisera against components of the interacting complexes. We also find that PsbS co-migrates with bands containing PSII, ATP synthase and cytochrome b(6)f as well as with LHCII-containing bands on non-denaturing Deriphat PAGE. These results suggest multiple location of PsbS in the thylakoid membrane and point to an unexpected lateral mobility of this PSII subunit. As revealed by immunogold labelling with antibody against PsbS, the protein is associated either with granal membranes or prevalently with stroma lamellae in low or high-intensity light-treated intact leaves, respectively. This finding is consistent with the capability of PsbS to interact with complexes located in stroma lamellae, even though the exact physiological condition(s) under which these interactions may take place remain to be clarified.

Light- and pH-dependent structural changes in the PsbS subunit of photosystem II.

In higher plants, the PsbS subunit of photosystem II (PSII) plays a crucial role in pH- and xanthophyll-dependent nonphotochemical quenching of excess absorbed light energy, thus contributing to the defense mechanism against photoinhibition. We determined the amino acid sequence of Zea mays PsbS and produced an antibody that recognizes with high specificity a region of the protein located in the stroma-exposed loop between the second and third putative helices. By means of this antiserum, the thylakoid membranes of various higher plant species revealed the presence of a 42-kDa protein band, indicating the formation of a dimer of the 21-kDa PsbS protein. Crosslinking experiments and immunoblotting with other antisera seem to exclude the formation of a heterodimer with other PSII protein components. The PsbS monomer/dimer ratio in isolated thylakoid membranes was found to vary with luminal pH in a reversible manner, the monomer being the prevalent form at acidic and the dimer at alkaline pH. In intact chloroplasts and whole plants, dimer-to-monomer conversion is reversibly induced by light, known to cause luminal acidification. Sucrose-gradient centrifugation revealed a prevalent association of the PsbS monomer and dimer with light-harvesting complex and PSII core complexes, respectively. The finding of the existence of a light-induced change in the quaternary structure of the PsbS subunit may contribute to understanding the mechanism of PsbS action during nonphotochemical quenching.

Structural and functional role of the PsbH protein in resistance to light stress in Synechocystis PCC 6803.

Four mutants of the cyanobacterium Synechocystis sp. PCC 6803, carrying a modified PsbH subunit on a PSI-less background, were characterized by optically-detected magnetic resonance (ODMR), electron transport kinetics, and oxygen-evolving activity. Their relative tolerance to light stress was measured. Results indicate that: (i) the PsbH protein is deeply involved in determining structural and functional properties of the QB site on the D1 protein, whereas the environment of the primary donor P680 and its acceptors pheophytin and QA are not significantly affected by modifications of this subunit or its deletion; (ii) the charge recombination rate, in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), is reduced by a factor of 2, independently of the particular modification. The same result is found with the strain in which the subunit has been deleted. This result is taken as an indication that PsbH is important in regulating protein dynamics of the entire PSII core complex; (iii)all investigated mutants display reduced tolerance to light stress, the extent of which depends on the particular modification. In this respect, mutations introduced in the transmembrane portion of the polypeptide are more effective than those involving the extramembrane N-terminal extension.