UV radiation and temperature increase alter the PSII function and defense mechanisms in a bloom-forming cyanobacterium Microcystis aeruginosa.

The aim was to determine the response of a bloom-forming Microcystis aeruginosa to climatic changes. Cultures of M. aeruginosa FACHB 905 were grown at two temperatures (25°C, 30°C) and exposed to high photosynthetically active radiation (PAR: 400-700 nm) alone or combined with UVR (PAR + UVR: 295-700 nm) for specified times. It was found that increased temperature enhanced M. aeruginosa sensitivity to both PAR and PAR + UVR as shown by reduced PSII quantum yields (Fv/Fm) in comparison with that at growth temperature (25°C), the presence of UVR significantly exacerbated the photoinhibition. M. aeruginosa cells grown at high temperature exhibited lower PSII repair rate (Krec) and sustained nonphotochemical quenching (NPQs) induction during the radiation exposure, particularly for PAR + UVR. Although high temperature alone or worked with UVR induced higher SOD and CAT activity and promoted the removal rate of PsbA, it seemed not enough to prevent the damage effect from them showing by the increased value of photoinactivation rate constant (Kpi). In addition, the energetic cost of microcystin synthesis at high temperature probably led to reduced materials and energy available for PsbA turnover, thus may partly account for the lower Krec and the declination of photosynthetic activity in cells following PAR and PAR + UVR exposure. Our findings suggest that increased temperature modulates the sensitivity of M. aeruginosa to UVR by affecting the PSII repair and defense capacity, thus influencing competitiveness and abundance in the future water environment.

Hypothetical chloroplast reading frame 51 encodes a photosystem I assembly factor in cyanobacteria.

Hypothetical chloroplast open reading frames (ycfs) are putative genes in the plastid genomes of photosynthetic eukaryotes. Many ycfs are also conserved in the genomes of cyanobacteria, the presumptive ancestors of present-day chloroplasts. The functions of many ycfs are still unknown. Here, we generated knock-out mutants for ycf51 (sll1702) in the cyanobacterium Synechocystis sp. PCC 6803. The mutants showed reduced photoautotrophic growth due to impaired electron transport between photosystem II (PSII) and PSI. This phenotype results from greatly reduced PSI content in the ycf51 mutant. The ycf51 disruption had little effect on the transcription of genes encoding photosynthetic complex components and the stabilization of the PSI complex. In vitro and in vivo analyses demonstrated that Ycf51 cooperates with PSI assembly factor Ycf3 to mediate PSI assembly. Furthermore, Ycf51 interacts with the PSI subunit PsaC. Together with its specific localization in the thylakoid membrane and the stromal exposure of its hydrophilic region, our data suggest that Ycf51 is involved in PSI complex assembly. Ycf51 is conserved in all sequenced cyanobacteria, including the earliest branching cyanobacteria of the Gloeobacter genus, and is also present in the plastid genomes of glaucophytes. However, Ycf51 has been lost from other photosynthetic eukaryotic lineages. Thus, Ycf51 is a PSI assembly factor that has been functionally replaced during the evolution of oxygenic photosynthetic eukaryotes.

Elevated CO2 modulates the physiological responses of Thalassiosira pseudonana to ultraviolet radiation.

Diatoms account for a large proportion of marine primary productivity, they tend to be the predominant species in the phytoplankton communities in the surface ocean with frequent and large light fluctuations. To understand the impacts of increased CO2 on diatoms' capacity in exploitation of variable solar radiation, we cultured a model diatom Thalassiosira pseudonana with 400 or 1000ppmv CO2 and exposed it to high photosynthetically active radiation (PAR) alone or PAR plus ultraviolet radiation (UVR) to examine its physiological performances. The results showed that the maximum photochemical efficiency (Fv/fm) was significantly reduced by high PAR and PAR + UVR in T. pseudonana, UVR-induced inhibition on PSII activity was exacerbated by high CO2. PSII activity drops coincide approximately with PsbA content in the cells exposed to high PAR or PAR + UVR, which was pronounced at high CO2. The removal of PsbD in T. pseudonana cells declined under high CO2 during UVR exposure, limiting the repair capacity of PSII. In addition, high CO2 reversed the induction of energy-dependent form of NPQ by UVR to the increase of Y(No), indicating the severe damage of the photoprotective reactions. Our findings suggest that the adverse impacts of UVR on PSII function of T. pseudonana were aggravated by the elevated CO2 through modulating its capacity in repair and protection, which thereby would influence its abundance and competitiveness in phytoplankton communities.

Physiological responses of the diatoms Thalassiosira weissflogii and Thalassiosira pseudonana to nitrogen starvation and high light.

As oceans warm, the depth of the upper mixed layer is predicted to decrease, resulting in insufficient nutrient supply and higher solar radiation for phytoplankton. In order to understand the photophysiological responses of the key eukaryotic phytoplankton diatoms to high light and nutrient limitation, we grew two diatoms, Thalassiosira weissflogii and Thalassiosira pseudonana under N starvation conditions and exposed them to high visible light. It showed that the large-sized diatom T. weissflogii can maintain photosynthetic activity for a longer period of time under nitrogen starvation as compared with the small-sized diatom T. pseudonana. The electron transfer reaction was inhibited in both diatoms and the fast closing of reaction centers promoted the development of QB non-reducing PSII centers, thus facilitated the rapid induction of NPQ, however, the induction of NPQ depended on the degree of N starvation. N starvation exacerbated the photoinhibition caused by high light. The smaller-sized T. pseudonana had a higher σi value and was more sensitive to high-light, but its PSII repair rate was also higher. In contrast, T. weissflogii was more tolerant to high light with a lower σi value, but the tolerance was severely reduced under N-starvation. This study provides helpful insight into how climate change variables impact diatom's photosynthetic physiology.

Sycrp2 Is Essential for Twitching Motility in the Cyanobacterium Synechocystis sp. Strain PCC 6803.

Two cAMP receptor proteins (CRPs), Sycrp1 (encoded by sll1371) and Sycrp2 (encoded by sll1924), exist in the cyanobacterium Synechocystis sp. strain PCC 6803. Previous studies have demonstrated that Sycrp1 has binding affinity for cAMP and is involved in motility by regulating the formation of pili. However, the function of Sycrp2 remains unknown. Here, we report that sycrp2 disruption results in the loss of motility of Synechocystis sp. PCC 6803, and that the phenotype can be recovered by reintroducing the sycrp2 gene into the genome of sycrp2-disrupted mutants. Electron microscopy showed that the sycrp2-disrupted mutant lost the pilus apparatus on the cell surface, resulting in a lack of cell motility. Furthermore, the transcript level of the pilA9-pilA11 operon (essential for cell motility and regulated by the cAMP receptor protein Sycrp1) was markedly decreased in sycrp2-disrupted mutants compared with the wild-type strain. Moreover, yeast two-hybrid analysis and a pulldown assay demonstrated that Sycrp2 interacted with Sycrp1 to form a heterodimer and that Sycrp1 and Sycrp2 interacted with themselves to form homodimers. Gel mobility shift assays revealed that Sycrp1 specifically binds to the upstream region of pilA9 Together, these findings indicate that in Synechocystis sp. PCC 6803, Sycrp2 regulates the formation of pili and cell motility by interacting with Sycrp1.IMPORTANCE cAMP receptor proteins (CRPs) are widely distributed in cyanobacteria and play important roles in regulating gene expression. Although many cyanobacterial species have two cAMP receptor-like proteins, the functional links between them are unknown. Here, we found that Sycrp2 in the cyanobacterium Synechocystis sp. strain PCC 6803 is essential for twitching motility and that it interacts with Sycrp1, a known cAMP receptor protein involved with twitching motility. Our findings indicate that the two cAMP receptor-like proteins in cyanobacteria do not have functional redundancy but rather work together.

Outer Membrane Iron Uptake Pathways in the Model Cyanobacterium Synechocystis sp. Strain PCC 6803.

Cyanobacteria are foundational drivers of global nutrient cycling, with high intracellular iron (Fe) requirements. Fe is found at extremely low concentrations in aquatic systems, however, and the ways in which cyanobacteria take up Fe are largely unknown, especially the initial step in Fe transport across the outer membrane. Here, we identified one TonB protein and four TonB-dependent transporters (TBDTs) of the energy-requiring Fe acquisition system and six porins of the passive diffusion Fe uptake system in the model cyanobacterium Synechocystis sp. strain PCC 6803. The results experimentally demonstrated that TBDTs not only participated in organic ferri-siderophore uptake but also in inorganic free Fe (Fe') acquisition. 55Fe uptake rate measurements showed that a TBDT quadruple mutant acquired Fe at a lower rate than the wild type and lost nearly all ability to take up ferri-siderophores, indicating that TBDTs are critical for siderophore uptake. However, the mutant retained the ability to take up Fe' at 42% of the wild-type Fe' uptake rate, suggesting additional pathways of Fe' acquisition besides TBDTs, likely by porins. Mutations in four of the six porin-encoding genes produced a low-Fe-sensitive phenotype, while a mutation in all six genes was lethal to cell survival. These diverse outer membrane Fe uptake pathways reflect cyanobacterial evolution and adaptation under a range of Fe regimes across aquatic systems.IMPORTANCE Cyanobacteria are globally important primary producers and contribute about 25% of global CO2 fixation. Low Fe bioavailability in surface waters is thought to limit the primary productivity in as much as 40% of the global ocean. The Fe acquisition strategies that cyanobacteria have evolved to overcome Fe deficiency remain poorly characterized. We experimentally characterized the key players and the cooperative work mode of two Fe uptake pathways, including an active uptake pathway and a passive diffusion pathway in the model cyanobacterium Synechocystis sp. PCC 6803. Our finding proved that cyanobacteria use ferri-siderophore transporters to take up Fe', and they shed light on the adaptive mechanisms of cyanobacteria to cope with widespread Fe deficiency across aquatic environments.

Characterization of the sulfur-formation (suf) genes in Synechocystis sp. PCC 6803 under photoautotrophic and heterotrophic growth conditions.

MAIN CONCLUSION: The sulfur-formation ( suf ) genes play important roles in both photosynthesis and respiration of cyanobacteria, but the organism prioritizes Fe-S clusters for respiration at the expense of photosynthesis. Iron-sulfur (Fe-S) clusters are important to all living organisms, but their assembly mechanism is poorly understood in photosynthetic organisms. Unlike non-photosynthetic bacteria that rely on the iron-sulfur cluster system, Synechocystis sp. PCC 6803 uses the Sulfur-Formation (SUF) system as its major Fe-S cluster assembly pathway. The co-expression of suf genes and the direct interactions among SUF subunits indicate that Fe-S assembly is a complex process in which no suf genes can be knocked out completely. In this study, we developed a condition-controlled SUF Knockdown mutant by inserting the petE promoter, which is regulated by Cu2+ concentration, in front of the suf operon. Limited amount of the SUF system resulted in decreased chlorophyll contents and photosystem activities, and a lower PSI/PSII ratio. Unexpectedly, increased cyclic electron transport and a decreased dark respiration rate were only observed under photoautotrophic growth conditions. No visible effects on the phenotype of SUF Knockdown mutant were observed under heterotrophic culture conditions. The phylogenetic distribution of the SUF system indicates that it has a co-evolutionary relationship with photosynthetic energy storing pathways.

Toll-like receptor-2 and -4 are associated with hyperlipidemia.

Recent studies have suggested that toll-like receptors (TLRs) contribute to insulin resistance, and that fatty acids have a role in TLR activation. Other studies have found that TLR2 and TLR4 upregulation is consistent with an increase in serum lipid. Therefore, it was hypothesized that TLRs are associated with hyperlipidemia. The aim of the present study was to investigate whether TLR2 or TLR4 was associated with hyperlipidemia and to provide novel targets for hyperlipidemia therapy. Volunteers were selected at the Medical Examination Center of Hebei General Hospital (Shijiazhuang, China), including 43 patients with high triglyceride (TG) levels, 84 with high total cholesterol (TC) levels and 55 with high TG and high TC levels. In addition, 68 healthy volunteers were selected as a control group. For the animal study, the TLR gene and protein levels were assessed in the skeletal muscle of rats fed a high­fat diet. As expected, TLR2 and TLR4 gene expression were upregulated when TC increased, TG increased, or TC and TG increased. In rats fed a high­fat diet, the levels of gene and protein expression in the skeletal muscle of the two TLRs were all increased compared with the control group, this was consistent with an increase in TC and TG. In addition, in drug treatment groups the mRNA and protein expression levels of TLR in the skeletal muscle of rats fed a high fat diet were decreased, as were the TC and TG levels. In conclusion, these findings suggest that TLR2 and TLR4 are associated with hyperlipidemia.

Chinese medicine Jinlida (JLD) ameliorates high-fat-diet induced insulin resistance in rats by reducing lipid accumulation in skeletal muscle.

The present paper reports the effects of Jinlida (JLD), a traditional Chinese medicine which has been given as a treatment for high-fat-diet (HFD)-induced insulin resistance. A randomized controlled experiment was conducted to provide evidence in support of the affects of JLD on insulin resistance induced by HFD. The affect of JLD on blood glucose, lipid, insulin, adiponectin, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and total bilirubin (TBIL) in serum and lipid content in skeletal muscle was measured. Genes and proteins of the AMPK signaling pathway were analyzed by real time RT-PCR and Western blot. Adiponectin receptor 1 and 2 (ADIPOR1, ADIPOR2) and other genes involved in mitochondrial function and fat oxidation were analyzed by real time RT-PCR. Histological staining was also performed. JLD or pioglitazone administration ameliorated fasting plasma levels of glucose, insulin, triglyceride (TG), total cholesterol (TC), ALT, AST and non-esterified fatty acid (NEFA) (P < 0.05). Treatment with JLD or pioglitazone significantly reverted muscle lipid content (P < 0.05). JLD (1.5 g/kg) significantly increased plasma adiponectin concentration by 60.17% and increased AMPK and acetyl-CoA carboxylase (ACC) phosphorylation in skeletal muscle (P < 0.05). JLD administration increased levels of ADIPOR1 and ADIPOR2 by 1.48 and 1.29 respectively. Levels of genes involved in mitochondrial function and fat oxidation were increased. This study provides the molecular mechanism by which JLD ameliorates HFD-induced insulin resistance in rats.

Jinlida reduces insulin resistance and ameliorates liver oxidative stress in high-fat fed rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Jinlida (JLD) is a compound preparation formulated on the basis of traditional Chinese medicine and is officially approved for the treatment of type 2 diabetes (T2DM) in China. We aimed to elucidate the mechanism of JLD treatment, in comparison to metformin treatment, on ameliorating insulin sensitivity in insulin resistant rats and to reveal its anti-oxidant properties. MATERIALS AND METHODS: Rats were fed with standard or high-fat diet for 6 weeks. After 6 weeks, the high-fat fed rats were subdivided into five groups and orally fed with JLD or metformin for 8 weeks. Fasting blood glucose (FBG), fasting blood insulin, blood lipid and antioxidant enzymes were measured. Intraperitoneal glucose tolerance test (IPGTT) and hyperinsulinemic euglycemic clamp technique were carried out to measure insulin sensitivity. Gene expression of the major signaling pathway molecules that regulate glucose uptake, including insulin receptor (INSR), insulin receptor substrate-1 (IRS-1), phosphoinositide-3-kinase (PI3K), protein kinase beta (AKT), and glucose transporter type 2 (GLUT2), were assessed by quantitative RT-PCR. The totle and phosphorylation expression of IRS-1, AKT, JNK and p38MAPK were determined by Western blot. RESULTS: Treatment with JLD effectively ameliorated the high-fat induced hyperglycemia, hyperinsulinemia and hyperlipidemia. Similar to metformin, the high insulin resistance in high-fat fed rats was significantly decreased by JLD treatment. JLD displayed anti-oxidant effects, coupled with up-regulation of the insulin signaling pathway. The attenuation of hepatic oxidative stress by JLD treatment was associated with reduced phosphorylation protein levels of JNK and p38MAPK. CONCLUSIONS: Treatment with JLD could moderate glucose and lipid metabolism as well as reduce hepatic oxidative stress, most likely through the JNK and p38MAPK pathways.