Wuttinun Raksajit

Small chloroplast-targeted DnaJ proteins are involved in optimization of photosynthetic reactions in Arabidopsis thaliana

BackgroundDnaJ proteins participate in many metabolic pathways through dynamic interactions with various components of these processes. The role of three small chloroplast-targeted DnaJ proteins, AtJ8 (At1 g80920), AtJ11 (At4 g36040) and AtJ20 (At4 g13830), was investigated here using knock-out mutants of Arabidopsis thaliana. Photochemical efficiency, capacity of CO2 assimilation, stabilization of Photosystem (PS) II dimers and supercomplexes under high light illumination, energy distribution between PSI and PSII and phosphorylation of PSII-LHCII proteins, global gene expression profiles and oxidative stress responses of these DnaJ mutants were analyzed.ResultsKnockout of one of these proteins caused a series of events including a decrease in photosynthetic efficiency, destabilization of PSII complexes and loss of control for balancing the redox reactions in chloroplasts. Data obtained with DNA microarray analysis demonstrated that the lack of one of these DnaJ proteins triggers a global stress response and therefore confers the plants greater tolerance to oxidative stress induced by high light or methyl viologen treatments. Expression of a set of genes encoding enzymes that detoxify reactive oxygen species (ROS) as well as a number of stress-related transcription factors behaved in the mutants at growth light similarly to that when wild-type (WT) plants were transferred to high light. Also a set of genes related to redox regulation were upregulated in the mutants. On the other hand, although the three DnaJ proteins reside in chloroplasts, the expression of most genes encoding thylakoid membrane proteins was not changed in the mutants.ConclusionIt is proposed that the tolerance of the DnaJ protein knockout plants to oxidative stress occurs at the expense of the flexibility of photosynthetic reactions. Despite the fact that the effects of the individual protein knockout on the response of plants to high light treatment are quite similar, it is conceivable that both specific- and cross-talk functions exist between the three small chloroplast-targeted DnaJ proteins, AtJ8, AtJ11 and AtJ20.

Characterization of the substrate-binding PotD subunit in Synechocystis sp. strain PCC 6803

The potD gene encodes the bacterial substrate-binding subunit of the polyamine transport system. The uptake system, which belongs to the ABC transporters, has been characterized in Escherichia coli, but it has not been previously studied in cyanobacteria. Although the overall sequence identity between Synechocystis sp. strain PCC 6803 (hereafter Synechocystis) PotD and Escherichia coli PotD is 24%, the ligand-binding site in the constructed homology model of Synechocystis PotD is well conserved. The conservation of the five polyamine-binding residues (Asp206, Glu209, Trp267, Trp293, and Asp295 in Synechocystis PotD) between these two species indicated polyamine-binding capacity for Synechocystis PotD. The SynechocystispotD gene is functional and its expression is under environmental regulation at transcriptional as well as post-transcriptional levels. Furthermore, an in vitro binding assay with the purified recombinant PotD protein demonstrated that the Synechocystis PotD protein is able to bind polyamines and favors spermidine over putrescine. Finally, we confirmed that Synechocystis PotD plays a physiological role in the uptake of polyamines in vivo using a constructed Synechocystis potD-disruption mutant.

Characterization of trophic changes and a functional oxidative pentose phosphate pathway in Synechocystis sp. PCC 6803

Cyanobacteria have a tremendous activity to adapt to environmental changes of their growth conditions. In this study, Synechocystis sp. PCC 6803 was used as a model organism to focus on the alternatives of cyanobacterial energy metabolism. Glucose oxidation in Synechocystis sp. PCC6803 was studied by inactivation of slr1843, encoding glucose-6-phosphate dehydrogenase (G6PDH), the first enzyme of the oxidative pentose phosphate pathway (OPPP). The resulting zwf strain was not capable of glucose supported heterotrophic growth. Growth under autotrophy and under mixotrophy was similar to that of the wild-type strain, even though oxygen evolution and uptake rates of the mutant were decreased in the presence of glucose. The organic acids citrate and succinate supported photoheterotrophic growth of both WT and zwf. Proteome analysis of soluble and membrane fractions allowed identification of four growth condition-dependent proteins, pentose-5-phosphate 3-epimerase (slr1622), inorganic pyrophosphatase (sll0807), hypothetical protein (slr2032) and ammonium/methylammonium permease (sll0108) revealing details of maintenance of the cellular carbon/nitrogen/phosphate balance under different modes of growth.

Recombinant polyamine-binding protein of Synechocystis sp. PCC 6803 specifically binds to and is induced by polyamines.

His-tagged Synechocystis sp. PCC 6803 PotD protein (rPotD) involved in polyamine transport was overexpressed in Escherichia coli. The purified rPotD showed saturable binding kinetics with radioactively labeled polyamines. The rPotD exhibited a similar binding characteristic for three polyamines, with putrescine having less preference. The Kd values for putrescine, spermine, and spermidine were 13.2, 8.3, and 7.8 μM, respectively. Binding of rPotD with polyamines was maximal at pH 8.0. Docking of these polyamines into the homology model of Synechocystis PotD showed that all three polyamines are able to interact with Synechocystis PotD. The binding modes of the docked putrescine and spermidine in Synechocystis are similar to those of PotF and PotD in E. coli, respectively. Competition experiments showed specific binding of rPotD with polyamines. The presence of putrescine and spermidine in the growth medium could induce an increase in PotD contents, suggesting the role of PotD in mediating the transport of polyamine in Synechocystis sp. PCC 6803.

Transcriptional regulation and structural modeling of the FutC subunit of an ABC-type iron transporter in Synechocystis sp. strain PCC 6803

The futC gene encodes a subunit of an ATP-binding cassette (ABC)-type iron transporter in Synechocystis sp. strain PCC 6803. In the present study, we have focused on the environmental regulation of futC transcription in the model organism Synechocystis sp. strain PCC 6803 and, moreover, studied the transcriptional regulation of the other transporter subunits, futA1, futA2 and futB. The steady-state amounts of the futA1, futA2, futB and futC transcripts were regulated under several conditions studied including darkness, temperature, alternative nitrogen source, salt and osmotic stresses and iron deficiency. Transcription of all subunits of the FutABC-iron transporter seems to be under similar regulation, which, according to our results, may also apply to genes encoding subunits of other transporters in Synechocystis. The sequence alignment, including sequences from six different organisms, revealed the conserved nature of FutC. Based on the sequence alignment and the structural model of FutC, the monomer consists of a nucleotide-binding domain (NBD) and a regulatory domain. The NBD is well conserved indicating completely functional ATP binding.

Factors affecting photobiological hydrogen production in five filamentous cyanobacteria from Thailand

We report here the screening of sixteen cyanobacterial and three green algal strains from Thailand for their potential biohydrogen production. Five filamentous cyanobacterial species, namely Calothrix elenkinii, Fischerella muscicola, Nostoc calcicola, Scytonema bohneri, and Tolypothrix distorta, all possessing nitrogenase activity, showed potentially high biohydrogen production. These five strains showed higher hydrogen production in the absence than in the presence of nitrogen. In particular, F. muscicola had a 17-fold increased hydrogen production under combined nitrogen and sulfur deprived conditions. Among various sugars as a carbon source, glucose at 0.1% (w/v) gave the maximal hydrogen production of 10.9 μmol(H2) mg–1(Chl) h–1 in T. distorta grown in BG11 medium without nitrate. Increasing light intensity up to 250 μmol(photon) m–2 s–1 increased hydrogen production in F. muscicola and T. distorta. Overall results indicate that both F. muscicola and T. distorta have a high potential for hydrogen production amenable for further improvement by using molecular genetics technique.