Regulation of pepc gene expression in Anabaena sp. PCC 7120 and its effects on cyclic electron flow around photosystem I and tolerances to environmental stresses.

Since pepc gene encoding phosphoenolpyruvate carboxylase (PEPCase) has been cloned from Anabaena sp. PCC 7120 and other cyanobacteria, the effects of pepc gene expression on photosynthesis have not been reported yet. In this study, we constructed mutants containing either upregulated (forward) or downregulated (reverse) pepc gene in Anabaena sp. PCC 7120. Results from real-time quantitative polymerase chain reaction (RT-qPCR), Western blot and enzymatic analysis showed that PEPCase activity was significantly reduced in the reverse mutant compared with the wild type, and that of the forward mutant was obviously increased. Interestingly, the net photosynthesis in both the reverse mutant and the forward mutant were higher than that of the wild type, but dark respiration was decreased only in the reverse mutant. The absorbance changes of P700 upon saturation pulse showed the photosystem I (PSI) activity was inhibited, as reflected by Y(I), and Y(NA) was elevated, and dark reduction of P700(+) was stimulated, indicating enhanced cyclic electron flow (CEF) around PSI in the reverse mutant. Additionally, the reverse mutant photosynthesis was higher than that of the wild type in low temperature, low and high pH, and high salinity, and this implies increased tolerance in the reverse mutant through downregulated pepc gene.

Recombinant expression and functional analysis of a Chlamydomonas reinhardtii bacterial-type phosphoenolpyruvate carboxylase gene fragment.

To investigate the function of a bacterial-type phosphoenolpyruvate carboxylase (PEPC2) derived from photosynthetically-grown Chlamydomonas reinhardtii, a fragment of the pepc2 gene was cloned and expressed in Escherichia coli. After optimal induction for 6 h, PEPC activity in the reverse mutant was lower than wild type (0.9 vs. 1.7 U/mg protein), and soluble protein was also lower than wild type (119 vs. 186 mg/g dry wt). In contrast, the total lipid content was increased from 56 (in wild type) to 71 mg/g dry wt, despite the growth rate being slightly diminished. The changes in PEPC activity, soluble protein and total lipid in the forward mutant were the opposite (2.4 U/mg, 230 mg/g, and 44 mg/g dry wt, respectively). Together, these data indicate that PEPC may function as a metabolic pivot in the regulation of protein and lipid accumulation in this alga.

A mutation of OSOTP 51 leads to impairment of photosystem I complex assembly and serious photo-damage in rice.

Gene expression in chloroplasts is regulated by many nuclear-encoded proteins. In this study, we isolated a rice (Oryza sativa subsp. japonica) mutant osotp51 with significant reduction in photosystem I (PSI). The osotp51 is extremely sensitive to light and accumulates a higher level of reactive oxygen species. Its leaves are almost albino when grown at 40 µmol photons/m(2) per s. However, grown at 4 µmol photons/m(2) per s, osotp51 has a similar phenotype to the wild-type. 77K chlorophyll fluorescence analysis showed a blue shift in the highest peak emission from PSI in osotp51. In addition, the level of PSI and PSII dimer is dramatically reduced in osotp51. OSOTP 51 encodes a pentatricopeptide repeats protein, homologous to organelle transcript processing 51 in Arabidopsis. Loss-of-function OSOTP51 affects intron splicing of a number of plastid genes, particularly the ycf3 coding a protein involved in the assembly of PSI complex. OSOTP51 is functionally conserved in higher plants. The mutation of osotp51 indirectly leads to a widespread change in the structure and functions of PSI, results in severe photoinhibition, and finally dies, even when grown under very low light intensity.

Reversible association of ribulose-1, 5-bisphosphate carboxylase/oxygenase activase with the thylakoid membrane depends upon the ATP level and pH in rice without heat stress.

Ribulose-1, 5-bisphosphate carboxylase/oxygenase (Rubisco) activase (RCA) in the thylakoid membrane (TM) has been shown to play a role in protection and regulation of photosynthesis under moderate heat stress. However, the physiological significance of RCA bound to the TM (TM-RCA) without heat stress remains unknown. In this study, it is first shown, using experiments in vivo, that the TM-RCA varies in rice leaves at different development stages, under different environmental conditions, and in a rice mutant. Furthermore, it is shown that the amount of TM-RCA always increased when the Rubisco activation state and the pH gradient across the TM (DeltapH) decreased. It was then demonstrated in vitro that the RCA bound dynamically to TM and the amount of TM-RCA increased during Rubisco activation. A high level of ATP and a high pH value promoted the dissociation of RCA from the TM. Both the RCA association with and dissociation from the TM showed conformational changes related to the ATP level or pH as indicated by the changes in fluorescence intensity of 1-anilinonaphthalene-8-sulphonic acid (ANS) binding to RCA. These results suggest that the reversible association of RCA with the TM is ATP and pH (or DeltapH) dependent; it might be involved in the RCA activation of Rubisco, in addition to the previously discovered role in the protection and regulation of photosynthesis under heat stress.

Construction of a chloroplast protein interaction network and functional mining of photosynthetic proteins in Arabidopsis thaliana.

Chloroplast is a typical plant cell organelle where photosynthesis takes place. In this study, a total of 1,808 chloroplast core proteins in Arabidopsis thaliana were reliably identified by combining the results of previously published studies and our own predictions. We then constructed a chloroplast protein interaction network primarily based on these core protein interactions. The network had 22,925 protein interaction pairs which involved 2,214 proteins. A total of 160 previously uncharacterized proteins were annotated in this network. The subunits of the photosynthetic complexes were modularized, and the functional relationships among photosystem I (PSI), photosystem II (PSII), light harvesting complex of photosystem I (LHC I) and light harvesting complex of photosystem I (LHC II) could be deduced from the predicted protein interactions in this network. We further confirmed an interaction between an unknown protein AT1G52220 and a photosynthetic subunit PSI-D2 by yeast two-hybrid analysis. Our chloroplast protein interaction network should be useful for functional mining of photosynthetic proteins and investigation of chloroplast-related functions at the systems biology level in Arabidopsis.

[The progress in research on the reduced nicotinamide adenine di(tri)-nucleotide phosophate [NAD(P)H] dehydrogenase complex and chlororespiration].

Besides the non-cyclic electron transport driven by the two photosystems (PSII and PSI), the cyclic electron transport pathways around PSI are also essential for efficient photosynthesis. As one of these pathways, the NAD(P)H dehydrogenase complex (NDH complex) mediated cyclic electron transport has been well studied. Along with the identification of the plastid terminal oxydase (PTOX), the functions of NDH-mediated cyclic and chlororepiratory electron transport in energy supply for photosynthesis as well as in the resistance to photooxidative stress have increasingly been brought to the researchers' attention. In the present paper, the structural characteristics of NDH complex, the regulatory mechanism, and the physiological significance of NDH mediated cyclic electron transport and chlororespiration are reviewed.

[Differences in plant hormone level of cucumber seedlings grown under sulfur or xenon lamp].

The primary mechanism of growth difference of cucumber (Cucumis sativus L.) seedlings cultured under sulfur lamp and xenon lamp in a phytotron was investigated. Compared with cucumber seedlings grown under xenon lamp, those under sulfur lamp were shorter, and the cell number in the middle hypocotyls epidermis and cortex of them were more (Fig. 1, Table 1). Endogenous hormone analysis indicates that the content of IAA and GA(3) of seedlings under sulfur lamp were 17% and 24% lower, while ABA content was 31% higher than that under xenon lamp (Fig. 2). Based on these results, it is suggested that the growth difference between cucumber seedlings grown under sulfur lamp and under xenon lamp might be related to the control of endogenous hormones.

[Separation of hydrophobic NAD(P)H dehydrogenase subcomplexes from cyanobacterium Synechocystis PCC6803].

Many efforts have been paid to the separation of an integrated NA(D)PH dehydrogenase (NDH) complex. Several hydrophilic subcomplexes of NDH have been purified from the cyanobacterium Synechocystis PCC6803. However, no hydrophobic NDH subcomplex has ever been separated from cyanobacteria yet. In this paper, two NDH subcomplexes were separated from n-dodecyl beta-D-maltoside(DM)-treated whole cell extracts of Synechocystis PCC6803 by anion exchange chromatography and gel filtration. Both subcomplexes contained the hydrophobic subunit NdhA, suggesting that they were hydrophobic NDH subcomplexes. Of the two subcomplexes, only one subcomplex contained NdhH. These subcomplexes showed NADPH-nitroblue tetrazolium (NBT) oxidoreductase activity and could specifically oxidize NADPH when several quinone analogues were used as electron acceptors, such as ferricyanide, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), 2,6-dichlorophenol indophenol (DCPIP), duroquinone, ubiquinone-0 (UQ-0), etc.

The Involvement of Plastoquinone in the Pyocyanine-mediated Cyclic Electron Transport around Photosystem I.

2,5 dibromo-3-methyl-5-isopropyl-p-benzoquinone (DBMIB), an inhibitor of plastoquinone, inhibited photosystem I cyclic electron transport mediated by pyocyanine of low concentration, but had no effect on that mediated by phenazine methosulphate (PMS). In the presence of pyocyanine, the thylakoids displayed a transient post-illumination increase in chlorophyll fluorescence which resembled that displayed in leaves. The above results indicate the involvement of plastoquinone in the pyocyanine-mediated cyclic electron transport around photosystem I.