Complexing Photosynthesis: Photosynthesis operates through a series of protein complexes that harvest sunlight and turn it into chemical energy. The separate complexes—including photosystems I and II, phycobilisome antennae, and reaction centers—are understood for a number of photosynthetic organisms; however, the large-scale organization and interactions between them are less clear. Using protein cross-linking, Liu et al. (p. 1104 ) demonstrate how the individual components are organized when present as a megacomplex in the cyanobacterium Synechocystis PCC 6803. Time-resolved fluorescence spectroscopy indicated that the phycobilisomes transfer energy to both photosystems, which is consistent with their molecular arrangement.

Precise genome editing of plants has the potential to reshape global agriculture through the targeted engineering of endogenous pathways or the introduction of new traits. To develop a CRISPR nuclease-based platform that would enable higher efficiencies of precise gene insertion or replacement, we screened the Cpf1 nucleases from Francisella novicida and Lachnospiraceae bacterium ND2006 for their capability to induce targeted gene insertion via homology directed repair. Both nucleases, in the presence of a guide RNA and repairing DNA template flanked by homology DNA fragments to the target site, were demonstrated to generate precise gene insertions as well as indel mutations at the target site in the rice genome. The frequency of targeted insertion for these Cpf1 nucleases, up to 8%, is higher than most other genome editing nucleases, indicative of its effective enzymatic chemistry. Further refinements and broad adoption of the Cpf1 genome editing technology have the potential to make a dramatic impact on plant biotechnology.

Interfering RNA was used to suppress the expression of the genes At1g06680 and At2g30790 in Arabidopsis thaliana, which encode the PsbP-1 and PsbP-2 proteins, respectively, of photosystem II (PS II). A phenotypic series of transgenic plants was recovered that expressed intermediate and low amounts of PsbP. Chlorophyll fluorescence induction and Q(A)(-) decay kinetics analyses were performed. Decreasing amounts of expressed PsbP protein led to the progressive loss of variable fluorescence and a marked decrease in the fluorescence quantum yield (F(V)/F(M)). This was primarily due to the loss of the J to I transition. Analysis of the fast fluorescence rise kinetics indicated no significant change in the number of PS II(beta) centers present in the mutants. Analysis of Q(A)(-) decay kinetics in the absence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea indicated a defect in electron transfer from Q(A)(-) to Q(B), whereas experiments performed in the presence of this herbicide indicated that charge recombination between Q(A)(-) and the oxygen-evolving complex was seriously retarded in the plants that expressed low amounts of the PsbP protein. These results demonstrate that the amount of functional PS II reaction centers is compromised in the plants that exhibited intermediate and low amounts of the PsbP protein. Plants that lacked detectable PsbP were unable to survive in the absence of sucrose, indicating that the PsbP protein is required for photoautotrophy. Immunological analysis of the PS II protein complement indicated that significant losses of the CP47 and D2 proteins, and intermediate losses of the CP43 and D1 proteins, occurred in the absence of the PsbP protein. This demonstrates that the extrinsic protein PsbP is required for PS II core assembly/stability.

Photosystem II (PSII), a large multisubunit pigment–protein complex localized in the thylakoid membrane of cyanobacteria and chloroplasts, mediates light-driven evolution of oxygen from water. Recently, a high-resolution X-ray structure of the mature PSII complex has become available. Two PSII polypeptides, D1 and CP43, provide many of the ligands to an inorganic Mn ₄ Ca center that is essential for water oxidation. Because of its unusual redox chemistry, PSII often undergoes degradation followed by stepwise assembly. Psb27, a small luminal polypeptide, functions as an important accessory factor in this elaborate assembly pathway. However, the structural location of Psb27 within PSII assembly intermediates has remained elusive. Here we report that Psb27 binds to CP43 in such assembly intermediates. We treated purified genetically tagged PSII assembly intermediate complexes from the cyanobacterium Synechocystis 6803 with chemical cross-linkers to examine intermolecular interactions between Psb27 and various PSII proteins. First, the water-soluble 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) was used to cross-link proteins with complementary charged groups in close association to one another. In the His27△ ctpA PSII preparation, a 58-kDa cross-linked species containing Psb27 and CP43 was identified. This species was not formed in the HT3△ ctpA △ psb27 PSII complex in which Psb27 was absent. Second, the homobifunctional thiol-cleavable cross-linker 3,3′-dithiobis(sulfosuccinimidylpropionate) (DTSSP) was used to reversibly cross-link Psb27 to CP43 in His27△ ctpA PSII preparations, which allowed the use of liquid chromatography/tandem MS to map the cross-linking sites as Psb27K ⁶³ ↔CP43D ³²¹ (trypsin) and CP43K ²¹⁵ ↔Psb27D ⁵⁸ AGGLK ⁶³ ↔CP43D ³²¹ (chymotrypsin), respectively. Our data suggest that Psb27 acts as an important regulatory protein during PSII assembly through specific interactions with the luminal domain of CP43.