Thioredoxin Dependent Changes in the Redox States of FurA from Anabaena sp. PCC 7120.

FurA is a multifunctional regulator in cyanobacteria that contains five cysteines, four of them arranged into two CXXC motifs. Lack of a structural zinc ion enables FurA to develop disulfide reductase activity. In vivo, FurA displays several redox isoforms, and the oxidation state of its cysteines determines its activity as regulator and its ability to bind different metabolites. Because of the relationship between FurA and the control of genes involved in oxidative stress defense and photosynthetic metabolism, we sought to investigate the role of type m thioredoxin TrxA as a potential redox partner mediating dithiol-disulfide exchange reactions necessary to facilitate the interaction of FurA with its different ligands. Both in vitro cross-linking assays and in vivo two-hybrid studies confirmed the interaction between FurA and TrxA. Light to dark transitions resulted in reversible oxidation of a fraction of the regulator present in Anabaena sp. PCC7120. Reconstitution of an electron transport chain using E. coli NADPH-thioredoxin-reductase followed by alkylation of FurA reduced cysteines evidenced the ability of TrxA to reduce FurA. Furthermore, the use of site-directed mutants allowed us to propose a plausible mechanism for FurA reduction. These results point to TrxA as one of the redox partners that modulates FurA performance.

2-oxoglutarate modulates the affinity of FurA for the ntcA promoter in Anabaena sp. PCC 7120.

2-oxoglutarate (2-OG) is a central metabolite that acts as a signaling molecule informing about the status of the carbon/nitrogen balance of the cell. In recent years, some transcriptional regulators and even two-component systems have been described as 2-OG sensors. In the nitrogen-fixing cyanobacterium Anabaena sp. PCC 7120, two master regulators, NtcA and FurA, are deeply involved in the regulation of nitrogen metabolism. Both of them show a complex intertwined regulatory circuit to achieve a suitable regulation of nitrogen fixation. In this work, 2-OG is found to bind FurA, modulating the specific binding of FurA to the ntcA promoter. This study provides evidence of a new additional control point in the complex network controlled by the NtcA and FurA proteins.

Redox-Based Transcriptional Regulation in Prokaryotes: Revisiting Model Mechanisms.

SIGNIFICANCE: The successful adaptation of microorganisms to ever-changing environments depends, to a great extent, on their ability to maintain redox homeostasis. To effectively maintain the redox balance, cells have developed a variety of strategies mainly coordinated by a battery of transcriptional regulators through diverse mechanisms. Recent Advances: This comprehensive review focuses on the main mechanisms used by major redox-responsive regulators in prokaryotes and their relationship with the different redox signals received by the cell. An overview of the corresponding regulons is also provided. CRITICAL ISSUES: Some regulators are difficult to classify since they may contain several sensing domains and respond to more than one signal. We propose a classification of redox-sensing regulators into three major groups. The first group contains one-component or direct regulators, whose sensing and regulatory domains are in the same protein. The second group comprises the classical two-component systems involving a sensor kinase that transduces the redox signal to its DNA-binding partner. The third group encompasses a heterogeneous group of flavin-based photosensors whose mechanisms are not always fully understood and are often involved in more complex regulatory networks. FUTURE DIRECTIONS: Redox-responsive transcriptional regulation is an intricate process as identical signals may be sensed and transduced by different transcription factors, which often interplay with other DNA-binding proteins with or without regulatory activity. Although there is much information about some key regulators, many others remain to be fully characterized due to the instability of their clusters under oxygen. Understanding the mechanisms and the regulatory networks operated by these regulators is essential for the development of future applications in biotechnology and medicine.

Microcystin-LR Binds Iron, and Iron Promotes Self-Assembly.

The microcystin-producing Microcystis aeruginosa PCC 7806 and its close strain, the nonproducing Microcystis aeruginosa PCC 7005, grow similarly in the presence of 17 µM iron. Under severe iron deficient conditions (0.05 µM), the toxigenic strain grows slightly less than in iron-replete conditions, while the nonproducing microcystin strain is not able to grow. Isothermal titration calorimetry performed at cyanobacterial cytosol or meaningful environmental pHs values shows a microcystin-LR dissociaton constant for Fe2+ and Fe3+ of 2.4 µM. Using atomic force microscopy, 40% of microcystin-LR dimers were observed, and the presence of iron promoted its oligomerization up to six units. Microcystin-LR binds also Mo6+, Cu2+, and Mn2+. Polymeric microcystin binding iron may be related with a toxic cell colony advantage, providing enhanced iron bioavailability and perhaps affecting the structure of the gelatinous sheath. Inside cells, with microcystin implicated in the fitness of the photosynthetic machinery under stress conditions, the toxin would be involved in avoiding metal-dependent Fenton reactions when photooxidation causes disassembly of the iron-rich photosystems. Additionally, it could be hypothesized that polymerization-depolymerization dynamics may be an additional signal that could trigger changes (for example, in the binding of microcystin to proteins).

Cysteine Mutational Studies Provide Insight into a Thiol-Based Redox Switch Mechanism of Metal and DNA Binding in FurA from Anabaena sp. PCC 7120.

AIMS: The ferric uptake regulator (Fur) is the main transcriptional regulator of genes involved in iron homeostasis in most prokaryotes. FurA from Anabaena sp. PCC 7120 contains five cysteine residues, four of them arranged in two redox-active CXXC motifs. The protein needs not only metal but also reducing conditions to remain fully active in vitro. Through a mutational study of the cysteine residues present in FurA, we have investigated their involvement in metal and DNA binding. RESULTS: Residue C101 that belongs to a conserved CXXC motif plays an essential role in both metal and DNA binding activities in vitro. Substitution of C101 by serine impairs DNA and metal binding abilities of FurA. Isothermal titration calorimetry measurements show that the redox state of C101 is responsible for the protein ability to coordinate the metal corepressor. Moreover, the redox state of C101 varies with the presence or absence of C104 or C133, suggesting that the environments of these cysteines are mutually interdependent. INNOVATION: We propose that C101 is part of a thiol/disulfide redox switch that determines FurA ability to bind the metal corepressor. CONCLUSION: This mechanism supports a novel feature of a Fur protein that emerges as a regulator, which connects the response to changes in the intracellular redox state and iron management in cyanobacteria. Antioxid. Redox Signal. 00, 000-000.

γ-Lindane Increases Microcystin Synthesis in Microcystis aeruginosa PCC7806.

HCH factories, and the waste dumpsites associated to its production, have become a global environmental concern, and their runoff could pollute ground and surface waters with high levels of the pollutant. In this study, the influence of lindane (γ-HCH) on microcystin production has been investigated in Microcystis aeruginosa PCC7806. This toxic cyanobacterium is highly tolerant to γ-lindane (20 mg/L), and produces more toxin (microcystin) in the presence of the pollutant. Microcystis degrades γ-lindane and presence of γ-lindane induces genes involved in its own degradation (nirA). RT-PCRsq has been used to monitor changes in levels of transcripts encoded by the mcy operon (mcyD, mcyH and mcyJ), responsible for the microcystin synthesis machinery, as well as other genes involved in its transcriptional regulation, such as ntcA and fur family members. The presence of lindane in the culture media induces mcyD expression, as well as ntcA gene transcription, while other genes, such as mcyH, (putative ABC transporter), are downregulated. The amount of microcystin found in the cells and the culture media is higher when M. aeruginosa is treated with γ-lindane than in control cells. The results suggest that in a lindane polluted environment, Microcystis toxic strains may enhance their microcystin synthesis.

Usefulness of triglycerides-to-high-density lipoprotein cholesterol ratio for predicting the first coronary event in men.

Overweight and obesity potentiate the development of cardiovascular risk factors but many doubts have arisen recently regarding their role in coronary events. We evaluated the predictive value of a surrogate maker of insulin resistance, the ratio of triglyceride (TG) to high-density lipoprotein (HDL), for the incidence of a first coronary event in men workers according to body mass index (BMI). We designed a case-control study of active subjects collected from a single factory through their annual health examination and medical reports. Case subjects included those with myocardial infarction, unstable angina pectoris, or subclinical myocardial ischemia detected through electrocardiographic abnormalities. The sample was constituted by 208 case and 2,080 control subjects (mean age 49.9 years, 49.6 to 50.2). General characteristics of case and control subjects were well matched. The TG/HDL ratio was significantly higher in case subjects compared to controls. Stratification of the sample revealed an increasing prevalence of case subjects and mean TG/HDL in each category of BMI. Multivariable analysis, adjusted by smoking, demonstrated that TG/HDL increased 50% the risk of a first coronary event (odds ratio [OR] 1.47, 95% confidence interval [CI] 1.26 to 1.71), whereas low-density lipoprotein cholesterol values indicated a more moderate increased risk (OR 1.01, 95% CI 1.005 to 1.012); metabolic syndrome (OR 1.76, 95% CI 0.94 to 3.30) and hypertension (OR 1.50, 95% CI 0.81 to 2.79) did not reach statistical significance. The TG/HDL ratio was associated with a first coronary event in all categories of BMI. In conclusion, the TG/HDL ratio has a high predictive value of a first coronary event regardless of BMI.

Comparison of cardiovascular risk factors in young Spanish men between the 1980s and after the year 2000. Data from the AGEMZA study.

INTRODUCTION AND OBJECTIVES: To investigate the cardiovascular risk profile of a sample of young Spanish men taken in the 1980s, the initial AGEMZA study cohort, and to compare the findings with those in another sample with similar characteristics taken after 2000. METHODS: The two AGEMZA study cohorts comprised young men who were resident at the Zaragoza General Military Academy, where they were studying as aspiring cadets. A descriptive study of each cohort was carried out and the participants' anthropometric characteristics, sporting and dietary habits, exposure to toxins, and biochemical and lipid profiles were analyzed. Data on the prevalence of various risk factors were obtained for each cohort and the coronary disease risk was estimated using the Framingham equation. RESULTS: Comparison of data on 248 subjects from the current cohort with data on 260 from the initial cohort showed the following significant changes: weight (+6.03 kg), body mass index (BMI) (+1.57), cholesterol (+12.46 mg/dL), low-density lipoprotein cholesterol (+15.8 mg/dL), high-density lipoprotein cholesterol (-4.11 mg/dL), triglycerides (+3.64 mg/dL), apolipoprotein B (+24.8 mg/dL), estimated coronary disease risk in the next 10 years (+1/1000 individuals) and estimated coronary disease risk up to the age of 65 years (+23/1000 individuals). There were significant correlations between increases in weight and BMI and lipid profile alterations. CONCLUSIONS: The lipid profile and BMI were worse in the current sample. These findings make it essential that preventive measures for young people should be introduced and that an increased effort should be made to develop programs aimed at either stopping the progressive rise in obesity or even preventing it altogether.

Comparación de los factores de riesgo cardiovascular de jóvenes españoles de los años ochenta y tras el 2000. Datos del estudio AGEMZA / Comparison of cardiovascular risk factors in young spanish men between the 1980s and after the year 2000. Data from the AGEMZA study

Introducción y objetivos. Analizar el perfil de riesgo cardiovascular de una muestra de jóvenes varones españoles de los años ochenta, cohorte inicial del estudio AGEMZA, y compararla con otra muestra de similares características, tras el 2000. Métodos. Las dos muestras de AGEMZA están compuestas por varones jóvenes residentes en la Academia General Militar de Zaragoza, donde estudiaban en calidad de cadetes aspirantes. Realizamos un estudio descriptivo de cada cohorte, analizando datos antropométricos, hábitos deportivos, dietéticos y tóxicos y sus perfiles bioquímico y lipídico. Obtuvimos datos de la prevalencia de los diferentes factores de riesgo en cada una de ellas y estimamos el riesgo coronario para las dos cohortes mediante la ecuación de Framingham. Resultados. Comparamos datos de 260 sujetos de la muestra inicial y de 248 de la muestra actual, de lo que se obtuvo las siguientes variaciones significativas: peso (+6,03 kg), IMC (+1,57), colesterol total (+12,46 mg/dl), colesterol de las lipoproteínas de baja densidad (+15,8 mg/dl), colesterol de las lipoproteínas de alta densidad (-4,11 mg/dl), triglicéridos (+3,64 mg/dl) y apolipoproteína B (+24,8 mg/dl), el riesgo coronario estimado en los próximos 10 años (+1/1.000 sujetos) y el proyectado a los 65 años de edad (+23/1.000 sujetos). Los aumentos de peso y del IMC se correlacionan de forma significativa con los cambios encontrados en el perfil lipídico. Conclusiones. La muestra actual tiene peor perfil lipídico e IMC. Estos resultados nos obligan a tomar medidas preventivas en los jóvenes y extremar el desarrollo de campañas destinadas a frenar este aumento progresivo de la obesidad y para prevenir su aparición (AU)

Introduction and objectives. To investigate the cardiovascular risk profile of a sample of young Spanish men taken in the 1980s, the initial AGEMZA study cohort, and to compare the findings with those in another sample with similar characteristics taken after 2000. Methods. The two AGEMZA study cohorts comprised young men who were resident at the Zaragoza General Military Academy, where they were studying as aspiring cadets. A descriptive study of each cohort was carried out and the participants' anthropometric characteristics, sporting and dietary habits, exposure to toxins, and biochemical and lipid profiles were analyzed. Data on the prevalence of various risk factors were obtained for each cohort and the coronary disease risk was estimated using the Framingham equation. Results. Comparison of data on 248 subjects from the current cohort with data on 260 from the initial cohort showed the following significant changes: weight (+6.03 kg), body mass index (BMI) (+1.57), cholesterol (+12.46 mg/dL), low-density lipoprotein cholesterol (+15.8 mg/dL), high-density lipoprotein cholesterol (-4.11 mg/dL), triglycerides (+3.64 mg/dL), apolipoprotein B (+24.8 mg/dL), estimated coronary disease risk in the next 10 years (+1/1000 individuals) and estimated coronary disease risk up to the age of 65 years (+23/1000 individuals). There were significant correlations between increases in weight and BMI and lipid profile alterations. Conclusions. The lipid profile and BMI were worse in the current sample. These findings make it essential that preventive measures for young people should be introduced and that an increased effort should be made to develop programs aimed at either stopping the progressive rise in obesity or even preventing it altogether (AU)

Comparison of serum lipid values in subjects with and without the metabolic syndrome.

Insulin resistance is supposed to be the basis of metabolic syndrome (MS), although it is difficult to measure. The ratio of triglyceride (TG) to high-density lipoprotein (HDL) has been proposed as a surrogate marker of insulin resistance in overweight subjects. The aim of the present study was to assess the accuracy of the TG/HDL ratio for the diagnosis of MS. Data of 18,778 active workers (77.6% men) enrolled in 3 insurance companies in Spain were collected from their annual health examinations. Mean age was 42.2 +/- 10.7 years. MS was assessed according to modified Adult Treatment Panel III criteria. Prevalences of MS were 18.8% in men and 6.1% in women. Mean value of the TG/HDL ratio was 2.50 +/- 2.2 and increased in parallel to the number of MS components present. Subjects with MS had a ratio that was 2 times higher compared with those without (5.10 vs 2.03, p <0.001). Receiver operating characteristic curves were performed to assess the capability of the TG/HDL ratio to contribute to a diagnosis of MS and 80% sensitivity and 78% specificity were obtained for values >2.75 in men and >1.65 in women. In conclusion, the TG/HDL ratio is a feasible and accurate measurement for assessment of MS in healthy subjects. We propose cut-off values of 2.75 for men and 1.65 for women for a diagnosis of MS.