A low-cost and eco-friendly recombinant protein expression system using copper-containing industrial wastewater.

The development of innovative methods for highly efficient production of recombinant proteins remains a prominent focus of research in the biotechnology field, primarily due to the fact that current commercial protein expression systems rely on expensive chemical inducers, such as isopropyl ß-D-thiogalactoside (IPTG). In our study, we designed a novel approach for protein expression by creating a plasmid that responds to copper. This specialized plasmid was engineered through the fusion of a copper-sensing element with an optimized multiple cloning site (MCS) sequence. This MCS sequence can be easily customized by inserting the coding sequences of target recombinant proteins. Once the plasmid was generated, it was introduced into an engineered Escherichia coli strain lacking copA and cueO. With this modified E. coli strain, we demonstrated that the presence of copper ions can efficiently trigger the induction of recombinant protein expression, resulting in the production of active proteins. Most importantly, this expression system can directly utilize copper-containing industrial wastewater as an inducer for protein expression while simultaneously removing copper from the wastewater. Thus, this study provides a low-cost and eco-friendly strategy for the large-scale recombinant protein production. To the best of our knowledge, this is the first report on the induction of recombinant proteins using industrial wastewater.

FluPMT: Prediction of Predominant Strains of Influenza A Viruses Via Multi-task Learning.

Seasonal influenza vaccines play a crucial role in saving numerous lives annually. However, the constant evolution of the influenza A virus necessitates frequent vaccine updates to ensure its ongoing effectiveness. The decision to develop a new vaccine strain is generally based on the assessment of the current predominant strains. Nevertheless, the process of vaccine production and distribution is very time-consuming, leaving a window for the emergence of new variants that could decrease vaccine effectiveness, so predictions of influenza A virus evolution can inform vaccine evaluation and selection. Hence, we present FluPMT, a novel sequence prediction model that applies an encoder-decoder architecture to predict the hemagglutinin (HA) protein sequence of the upcoming season's predominant strain by capturing the patterns of evolution of influenza A viruses. Specifically, we employ time series to model the evolution of influenza A viruses, and utilize attention mechanisms to explore dependencies among residues of sequences. Additionally, antigenic distance prediction based on graph network representation learning is incorporated into the sequence prediction as an auxiliary task through a multi-task learning framework. Experimental results on two influenza datasets highlight the exceptional predictive performance of FluPMT, offering valuable insights into virus evolutionary dynamics, as well as vaccine evaluation and production.

Protein lipidation in health and disease: molecular basis, physiological function and pathological implication.

Posttranslational modifications increase the complexity and functional diversity of proteins in response to complex external stimuli and internal changes. Among these, protein lipidations which refer to lipid attachment to proteins are prominent, which primarily encompassing five types including S-palmitoylation, N-myristoylation, S-prenylation, glycosylphosphatidylinositol (GPI) anchor and cholesterylation. Lipid attachment to proteins plays an essential role in the regulation of protein trafficking, localisation, stability, conformation, interactions and signal transduction by enhancing hydrophobicity. Accumulating evidence from genetic, structural, and biomedical studies has consistently shown that protein lipidation is pivotal in the regulation of broad physiological functions and is inextricably linked to a variety of diseases. Decades of dedicated research have driven the development of a wide range of drugs targeting protein lipidation, and several agents have been developed and tested in preclinical and clinical studies, some of which, such as asciminib and lonafarnib are FDA-approved for therapeutic use, indicating that targeting protein lipidations represents a promising therapeutic strategy. Here, we comprehensively review the known regulatory enzymes and catalytic mechanisms of various protein lipidation types, outline the impact of protein lipidations on physiology and disease, and highlight potential therapeutic targets and clinical research progress, aiming to provide a comprehensive reference for future protein lipidation research.

The Correlation Between Serum Copper and Non-alcoholic Fatty Liver Disease in American Adults: an Analysis Based on NHANES 2011 to 2016.

Copper functions as an essential micronutrient influencing diverse metabolic processes in mammals, encompassing oxidative stress responses, lipid metabolism, and participation in enzymatic reactions. However, the impact of serum copper on non-alcoholic fatty liver disease (NAFLD) remains controversial. Our aim was to explore the precise correlation between serum copper and NAFLD in a large-scale population-based study. A total of 1377 participants from the National Health and Nutrition Examination Survey (NHANES) 2011-2016 were included in our study. The diagnosis of NAFLD and its progress to advanced liver fibrosis were based on serological indexes. One-way ANOVA, Kruskal-Wallis H test, and Chi-square test were used to access variations between quartiles groups of serum copper. We conducted multivariate-adjusted logistic regression models and subgroup analyses to investigate the association between serum copper and NAFLD, along with several metabolic diseases. Among the 1377 participants, 661 were diagnosed with NAFLD, and 141 of whom were classified into advanced liver fibrosis. Higher serum copper levels (≥ 21.00 µmol/L) were associated with an increased incidence of NAFLD (odds ratio (OR) = 2.07 (1.38-3.10), p < 0.001), as well as advanced liver fibrosis (OR = 2.40 (1.17-5.19), p = 0.025). Moreover, serum copper exhibited a positive correlation with hypertension, overweight, and abdominal obesity, all of which have been identified as risk factors of NAFLD. Additionally, female participants, under the age of 60, and with a higher body mass index (BMI) (> 24.9 kg/m2) emerged as the most vulnerable subgroup concerning the relationship between serum copper and NAFLD. In the U.S. population, a notable association has been identified, linking elevated serum copper to an increased susceptibility for both the onset and progression of NAFLD, along with several metabolic disorders associated with NAFLD. The adverse effects of excess copper warrant attention in the context of public health considerations.

Early-life stress contributes to depression-like behaviors in a two-hit mouse model.

BACKGROUND: Depression is a common psychological disorder with pathogenesis involving genetic and environmental interactions. Early life stress can adversely affect physical and emotional development and dramatically increase the risk for the development of depression and anxiety disorders. METHODS: To examine potential early life stress driving risk for anxiety and depression, we used a two-hit developmental stress model，injecting poly(I: C) into neonatal mice on P2-P6 followed by peripubertal unpredictable stress in adolescence. RESULTS: Our study shows that early-life and adolescent stress leads to anxiety and depression-related behavioral phenotypes in male mice. Early-life stress exacerbated depression-like behavior in mice following peripubertal unpredictable stress. We confirmed that early life stress might be involved in the decreased neuronal activity in the medial prefrontal cortex (mPFC) and might be involved in shaping behavioral phenotypes of animals. We found that increased microglia and neuroinflammation in the mPFC of two-hit mice and early life stress further boost microglia activation and inflammatory factors in the mPFC region of mice following adolescent stress. LIMITATIONS: The specific neural circuits and mechanisms by which microglia regulate depression-like behaviors require further investigation. CONCLUSIONS: Our findings provide a novel insight into developmental risk factors and biological mechanisms in depression and anxiety disorders.

Efficient synthesis of 6-membered cyclic monothiocarbonates from halohydrin and carbonyl sulfide.

We report herein an efficient method to synthesize 6-membered cyclic monothiocarbonates which are important intermediates for polymonothiocarbonate synthesis via the cycloaddition of carbonyl sulfide with 1,3-halohydrin using a low-cost base such as triethylamine and potassium carbonate. This protocol features excellent selectivity and efficiency, mild reaction conditions and easy-to-obtain starting materials.

Copper exerts cytotoxicity through inhibition of iron-sulfur cluster biogenesis on ISCA1/ISCA2/ISCU assembly proteins.

Copper is an essential mineral nutrient that provides the cofactors for some key enzymes. However, excess copper is paradoxically cytotoxic. Wilson's disease is an autosomal recessive hereditary disease characterized by pathological copper accumulation in many organs, with high mortality and disability. Nevertheless, many questions about the molecular mechanism in Wilson's disease remain unknown and there is an imperative need to address these questions to better exploit therapeutic strategy. In this study, we constructed the mouse model of Wilson's disease, ATP7A-/- immortalized lymphocyte cell line and ATP7B knockdown cells to explore whether copper could impair iron-sulfur cluster biogenesis in eukaryotic mitochondria. Through a series of cellular, molecular, and pharmacological analyses, we demonstrated that copper could suppress the assembly of Fe-S cluster, decrease the activity of the Fe-S enzyme and disorder the mitochondrial function both in vivo and in vitro. Mechanistically, we found that human ISCA1, ISCA2 and ISCU proteins have a strong copper-binding activity, which would hinder the process of iron-sulfur assembly. Of note, we proposed a novel mechanism of action to explain the toxicity of copper by providing evidence that iron-sulfur cluster biogenesis may be a primary target of copper toxicity both in cells and mouse models. In summary, the current work provides an in-depth study on the mechanism of copper intoxication and describes a framework for the further understanding of impaired Fe-S assembly in the pathological processes of Wilson's diseases, which helps to develop latent therapeutic strategies for the management of copper toxicity.

Use of lasers in gastrointestinal endoscopy: a review of the literature.

Lasers emit highly directional light with consistent wavelengths, and recent studies have demonstrated their successful applications in gastrointestinal endoscopic therapy. Although argon plasma coagulators (APC) became the preferred treatment option due to improved safety profile and lower costs, advancements in laser and optic fiber manufacturing have reignited interest in laser treatment. Different laser wavelengths have distinct features and applications based on their tissue absorption coefficient. Lasers with shorter wavelengths are effectively absorbed by hemoglobin, resulting in a good coagulation effect. Near-infrared lasers have ability to ablate solid tumors, while far-infrared lasers can make precise mucosal incisions without causing peripheral thermal damage. Lasers have proven to be highly applicable to endoscopy devices such as endoscopes, endoscopic ultrasound (EUS), double-balloon enteroscopes (DBE), and endoscopic retrograde cholangiopancreatography (ERCP), making them a potent tool to enhance the effectiveness of endoscopic treatments with minimal adverse events. This review aims to help readers understand the applications and effectiveness of lasers in gastrointestinal endoscopy, with the potential to promote the development and application of laser technology in the medical field.

Roles of conserved active site residues in the IscS cysteine desulfurase reaction.

Escherichia coli cysteine desulfurase (CD), IscS, modifies basal metabolism by transferring sulphur (S) from L-cysteine to numerous cellular pathways, whereas NFS1, a human CD, is active only in the formation of the [Acp]2:[ISD11]2:[NFS1]2 complex. Despite the accumulation of red-coloured IscS in E. coli cells as a result of the deficiency of accessible iron, as revealed in our previous studies, the mechanism of the potential enzymatic reaction remains unclear. In this study, the N-terminus of IscS was fused with the C-terminus of NFS1, which was reported to be almost fully active as IscS and exhibits a pyridoxal 5'-phosphate (PLP) absorption peak at 395 nm. Moreover, SUMO-EH-IscS exhibited significant growth recovery and NADH-dehydrogenase I activity in the iscS mutant cells. Furthermore, through in vitro and in vivo experiments combined with high-performance liquid chromatography and ultra-performance liquid chromatography-tandem mass spectrometry, it was shown that the new absorption peaks of the IscS H104Q, IscS Q183E, IscS K206A, and IscS K206A&C328S variants at 340 and 350 nm may correspond to the enzyme reaction intermediates, Cys-ketimine and Cys-aldimine, respectively. However, after mutation of the conserved active-site residues, additional absorption peaks at 420 and 430 nm were associated with PLP migration in the active-site pocket. Additionally, the corresponding absorption peaks of Cys-quinonoid, Ala-ketimine, and Ala-aldimine intermediates in IscS were 510, 325, and 345 nm, respectively, as determined by site-directed mutagenesis and substrate/product-binding analyses during the CD reaction process. Notably, red IscS formed in vitro by incubating IscS variants (Q183E and K206A) with excess L-alanine and sulphide under aerobic conditions produced an absorption peak similar to the wild-type IscS, at 510 nm. Interestingly, site-directed mutation of IscS with hydrogen bonds to PLP at Asp180 and Gln183 resulted in a loss of enzymatic activity followed by an absorption peak consistent with NFS1 (420 nm). Furthermore, mutations at Asp180 or Lys206 inhibited the reaction of IscS in vitro with L-cysteine (substrate) and L-alanine (product). These results suggest that the conserved active site residues (His104, Asp180, and Gln183) and their hydrogen bond with PLP in the N-terminus of IscS play a key role in determining whether the L-cysteine substrate can enter the active-site pocket and regulate the enzymatic reaction process. Therefore, our findings provide a framework for evaluating the roles of conserved active-site residues, motifs, and domains in CDs.

Novel antidepressant-like properties of the fullerenol in an LPS-induced depressive mouse model.

Depression is a common mental disease and is highly prevalent in populations. Dysregulated neuroinflammation and concomitant-activated microglia are involved in the pathogenesis of depression. Experimental evidence has indicated that fullerenol exerts anti-neuroinflammation and protective effects against neurological diseases. Here, we evaluated fullerenol's effects against lipopolysaccharide (LPS)-induced mouse depressive-like behaviors. Fullerenol treatment produced an antidepressant-like effect, as indicated by preventing the LPS-induced reduction in the sucrose preference and shortening the immobility durations in both the tail suspension test and the forced swim test. We found that fullerenol treatment mitigated LPS-induced hippocampal microglia activation and released proinflammatory cytokines. Meanwhile, fullerenol promoted hippocampus neurogenesis, evidenced by increased DCX-positive cells in LPS-treated mice. Hippocampal RNA-Seq analysis revealed proinflammatory cytokine and neurogenesis involved in fullerenol's antidepressant-like effects. Our data indicate that fullerenol exerts antidepressant effects, which might be due to beneficial functions in reducing neuroinflammatory processes and promoting neurogenesis in the hippocampus.

Transgenic RFP-RPS-30UbL strain of the nematode Caenorhabditis elegans as a biomonitor for environmental pollutants.

Environmental pollutants are recognized as one of the major concerns for public health. The free-living nematode Caenorhabditis elegans are widely used to evaluate the toxicity of environmental contaminants in biomonitoring researches. In the present study, a new transgenic strain, rps-30-/- ;RFP-RPS-30UbL was generated, with constitutively active rps-30 promoter used to control the expression of RFP-RPS-30UbL fusion protein. We found RFP-RPS-30UbL would accumulate to form 'rod-like' structures, when worms were exposed to environmental contaminants, including Cd, Hg, Pb, As, Paraquat and Dichlorvos. The number of the 'rod-like' structures was induced by environmental contaminants in a concentration- and time-dependent manner. The 'rod-like' structure formation could be detectable in response to the concentration of each contaminant as low as 24-h LC50 × 10-7 , and the detectable time could be within 2 h. Detecting the transcription and expression levels of RFP-RPS-30UbL in worms exposed to different kinds of environmental contaminants showed that the expression level of RFP-RPS-30UbL was not regulated by environmental contaminants, and the number differences of 'rod-like' structures were just due to the morphological change of RFP-RPS-30UbL from dispersion to accumulation induced by environmental contaminants. In addition, this transgenic strain was developed in rps-30-/- homozygous worm, which was a longevity strain. Detection of lifespan and brood size showed that rps-30-/- ;RFP-RPS-30UbL transgenic worm was more suitable to be cultured and used further than N2;GFP-RPS-30UbL , for expressing RPS-30UbL in wild type N2 worms shortened the lifespan and deceased the brood size. Therefore, rps-30-/- ;RFP-RPS-30UbL transgenic worm might play a potential role in versatile environmental biomonitoring, with the advantage of not only the convenient and quick fluorescence-based reporter assay, but also the quantificational evaluation of the toxicities of environmental contaminants using 'rod-like' structures with high sensitivity, off-limited the expression level of the reporter protein.

Methods of acoustic gas flux inversion-Investigation into the initial amplitude of bubble excitation.

Passive acoustic inversion techniques for measuring gas flux into the water column have the potential to be a powerful tool for the long-term monitoring and quantification of natural marine seeps and anthropogenic emissions. Prior inversion techniques have had limited precision due to lack of constraints on the initial amplitude of a bubble's excitation following its release into the water column ( R). R is determined by observing the acoustic signal of bubbles released from sediment in a controlled experiment and its use is demonstrated by quantifying the flux from a volcanic CO2 seep offshore Panarea (Italy), improving the precision by 78%.

ISCA2 deficiency leads to heme synthesis defects and impaired erythroid differentiation in K562 cells by indirect ROS-mediated IRP1 activation.

Iron­sulfur (Fe-S) clusters have been shown to play important roles in various cellular physiological process. Iron­sulfur cluster assembly 2 (ISCA2) is a vital component of the [4Fe-4S] cluster assembly machine. Several studies have shown that ISCA2 is highly expressed during erythroid differentiation. However, the role and specific regulatory mechanisms of ISCA2 in erythroid differentiation and erythroid cell growth remain unclear. RNA interference was used to deplete ISCA2 expression in human erythroid leukemia K562 cells. The proliferation, apoptosis, and erythroid differentiation ability of the cells were assessed. We show that knockdown of ISCA2 has profound effects on [4Fe-4S] cluster formation, diminishing mitochondrial respiratory chain complexes, leading to reactive oxygen species (ROS) accumulation and mitochondrial damage, inhibiting cell proliferation. Excessive ROS can inhibit the activity of cytoplasmic aconitase (ACO1) and promote ACO1, a bifunctional protein, to perform its iron-regulating protein 1(IRP1) function, thus inhibiting the expression of 5'-aminolevulinate synthase 2 (ALAS2), which is a key enzyme in heme synthesis. Deficiency of ISCA2 results in the accumulation of iron divalent. In addition, the combination of excessive ferrous iron and ROS may lead to damage of the ACO1 cluster and higher IRP1 function. In brief, ISCA2 deficiency inhibits heme synthesis and erythroid differentiation by double indirect downregulation of ALAS2 expression. We conclude that ISCA2 is essential for normal functioning of mitochondria, and is necessary for erythroid differentiation and cell proliferation.

Measurement of bistatic sea surface scattering with a parametric acoustic source.

This work presents the results from a series of bistatic sea surface scattering experiments conducted in shallow water using a parametric acoustic array as a source and a receiver comprising a horizontal linear array. The experiments measured scattering at three frequencies (4, 8, and 15 kHz) and at three incident grazing angles (13º, 20º, and 30º). The measurements were made over a 5 day period during which a variety of environmental conditions were encountered. This paper provides an outline of the experiments and presents some results for the forward scattering strength. The results show that the wave direction has a significant effect on the surface forward scattering. At each incident grazing angle, the fluctuations of scattering strength due to environmental conditions decreases as the frequency increases.

Heat shock transcription factor (Hsf) gene family in common bean (Phaseolus vulgaris): genome-wide identification, phylogeny, evolutionary expansion and expression analyses at the sprout stage under abiotic stress.

BACKGROUND: Common bean (Phaseolus vulgaris) is an essential crop with high economic value. The growth of this plant is sensitive to environmental stress. Heat shock factor (Hsf) is a family of antiretroviral transcription factors that regulate plant defense system against biotic and abiotic stress. To date, few studies have identified and bio-analyzed Hsfs in common bean. RESULTS: In this study, 30 Hsf transcription factors (PvHsf1-30) were identified from the PFAM database. The PvHsf1-30 belonged to 14 subfamilies with similar motifs, gene structure and cis-acting elements. The Hsf members in Arabidopsis, rice (Oryza sativa), maize (Zea mays) and common bean were classified into 14 subfamilies. Collinearity analysis showed that PvHsfs played a role in the regulation of responses to abiotic stress. The expression of PvHsfs varied across different tissues. Moreover, quantitative real-time PCR (qRT-PCR) revealed that most PvHsfs were differentially expressed under cold, heat, salt and heavy metal stress, indicating that PvHsfs might play different functions depending on the type of abiotic stress. CONCLUSIONS: In this study, we identified 30 Hsf transcription factors and determined their location, motifs, gene structure, cis-elements, collinearity and expression patterns. It was found that PvHsfs regulates responses to abiotic stress in common bean. Thus, this study provides a basis for further analysis of the function of PvHsfs in the regulation of abiotic stress in common bean.

Iron-sulphur cluster biogenesis factor LYRM4 is a novel prognostic biomarker associated with immune infiltrates in hepatocellular carcinoma.

BACKGROUND: LYRM4 is necessary to maintain the stability and activity of the human cysteine desulfurase complex NFS1-LYRM4-ACP. The existing experimental results indicate that cancer cells rely on the high expression of NFS1. However, the role of LYRM4 in liver hepatocellular carcinoma (LIHC) remains unclear. METHODS: In this study, we combined bioinformatics analysis and clinical specimens to evaluate the mRNA, protein expression, and gene regulatory network of LYRM4 in LIHC. Furthermore, we detected the activity of several classical iron-sulphur proteins in LIHC cell lines through UV-vis spectrophotometry. RESULTS: The mRNA and protein levels of LYRM4 were upregulated in LIHC. Subsequent analysis revealed that the LYRM4 mRNA expression was related to various clinical stratifications, prognosis, and survival of LIHC patients. In addition, the mRNA expression of LYRM4 was significantly associated with ALT, tumour thrombus, and encapsulation of HBV-related LIHC patients. IHC results confirmed that LYRM4 was highly expressed in LIHC tissues and showed that the expression of LYRM4 protein in LIHC was significantly correlated with age and serum low-density lipoprotein (LDL) and triglyceride (TG) content. In particular, the mRNA expression of key iron- sulphur proteins POLD1 and PRIM2 was significantly overexpressed and correlated with poor prognosis in LIHC patients. Compared with hepatocytes, the activities of mitochondrial complex I and aconitate hydratase (ACO2) in LIHC cell lines were significantly increased. These results indicated that the iron-sulphur cluster (ISC) biosynthesis was significantly elevated in LIHC, leading to ISC-dependent metabolic reprogramming. Changes in the activity of ISC-dependent proteins may also occur in paracancerous tissues. Further analysis of the biological interaction and gene regulation networks of LYRM4 suggested that these genes were mainly involved in the citric acid cycle and oxidative phosphorylation. Finally, LYRM4 expression in LIHC was significantly positively correlated with the infiltrating levels of six immune cell types, and both factors were strongly associated with prognosis. CONCLUSION: LYRM4 could be a novel prognostic biomarker and molecular target for LIHC therapy. In particular, the potential regulatory networks of LYRM4 overexpression in LIHC provide a scientific basis for future research on the role of the ISC assembly mechanism and LYRM4-mediated sulphur transfer routes in carcinogenesis.

Quercetin prevents primordial follicle loss via suppression of PI3K/Akt/Foxo3a pathway activation in cyclophosphamide-treated mice.

BACKGROUND: Chemotherapy improves the survival rates of patients with various cancers but often causes some adverse effects, including ovarian damage, characterised by a decrease in primordial follicle stockpiles. Recent studies have revealed that chemotherapy may stimulate the PI3K signalling pathway, thereby resulting in accelerated primordial follicle activation and a decreased ovarian reserve. Quercetin is an inhibitor of the PI3K pathway; however, its protective effects against chemotherapy-induced follicle loss in mice have not been established. In this study, the effects of quercetin in a mouse model of cyclophosphamide-induced ovarian dysfunction were investigated. METHODS: C57BL/6 female mice were used for the study. Paraffin sections of mouse ovaries (n = 30 mice) were stained with haematoxylin and eosin for differential follicle counts. Apoptosis (n = 5 mice per group) was evaluated by TUNEL assay. Immunohistochemical staining for ki67 and Foxo3a (n = 5 mice per group) was performed to evaluate the activation of primordial follicles. The role of the PI3K signalling pathway in the ovaries (n = 45 mice) was assessed by western blotting. RESULTS: Quercetin attenuated the cyclophosphamide-induced reduction in dormant primordial follicles. Analysis of the PI3K/Akt/Foxo3a pathway showed that quercetin decreased the phosphorylation of proteins that stimulate follicle activation in cyclophosphamide-induced ovaries. Furthermore, quercetin prevented cyclophosphamide-induced apoptosis in early growing follicles and early antral follicles, maintained anti-Müllerian hormone levels secreted by these follicles, and preserved the quiescence of the primordial follicle pool, as determined by intranuclear Foxo3a staining. CONCLUSIONS: Quercetin attenuates cyclophosphamide-induced follicle loss by preventing the phosphorylation of PI3K/Akt/Foxo3a pathway members and maintaining the anti-Müllerian hormone level through reduced apoptosis in growing follicles. Accordingly, quercetin is expected to improve fertility preservation and the prevention of endocrine-related side effects of chemotherapy.

The functional polymorphisms linked with interleukin-1ß gene expression are associated with bipolar disorder.

BACKGROUND: Bipolar disorder (BD) is a severe psychiatric illness attributable to multifactorial risk components (e.g. environmental stimuli, neuroinflammation, etc.), and genetic variations affecting these risk components are considered pivotal predisposing factors. The interleukin-1ß (IL-1ß) gene and its protein product have been repeatedly highlighted in the pathogenesis of BD. As functional polymorphisms and haplotypes linked with IL-1ß mRNA expression have been reported, whether they are correlated with the risk of developing BD remains to be tested. METHODS: To examine whether variations in the IL-1ß gene locus confer genetic risk of BD, we recruited 930 BD patients and 912 healthy controls for the current study. All subjects were Han Chinese, and were age- and gender-matched. We tested seven functional single nucleotide polymorphisms (SNPs) spanning the IL-1ß gene and one haplotype composed of three SNPs for their associations with risk of BD. RESULTS: We found that the functional SNPs in the promoter region of IL-1ß gene were significantly associated with risk of BD. The haplotype analyses further supported the involvement of IL-1ß promoter SNPs in BD. The BD risk SNPs in our study have been previously reported to predict higher IL-1ß levels in the brain and peripheral blood, which is consistent with the clinical observation of elevated IL-1ß levels in the lymphocytes or peripheral blood of patients with BD compared with healthy subjects. CONCLUSION: Our results support the contention that IL-1ß is likely a risk gene for BD, and further investigations on this gene may promote our understanding and clinical management of this illness.

Identification of an Intermediate Form of Ferredoxin That Binds Only Iron Suggests That Conversion to Holo-Ferredoxin Is Independent of the ISC System in Escherichia coli.

Escherichia coli [2Fe-2S]-ferredoxin and other ISC proteins encoded by the iscRSUA-hscBA-fdx-iscX (isc) operon are responsible for the assembly of iron-sulfur clusters. It is proposed that ferredoxin (Fdx) donates electrons from its reduced [2Fe-2S] center to iron-sulfur cluster biogenesis reactions. However, the underlying mechanisms of the [2Fe-2S] cluster assembly in Fdx remain elusive. Here, we report that Fdx preferentially binds iron, but not the [2Fe-2S] cluster, under cold stress conditions (≤16°C). The iron binding in Fdx is characterized by a unique absorption peak at 320 nm based on UV-visible spectroscopy. In addition, the iron-binding form of Fdx could be converted to the [2Fe-2S] cluster-bound form after transferring cold-stressed cells to normal cultivation temperatures above 25°C. In vitro experiments also revealed that Fdx could utilize bound iron to assemble the [2Fe-2S] cluster by itself. Furthermore, inactivation of the genes encoding IscS, IscU, and IscA did not limit [2Fe-2S] cluster assembly in Fdx, which was also observed by inactivating the isc or suf operon, indicating that iron-sulfur cluster biogenesis in Fdx arose from a unique pathway in E. coli Our results suggest that the intracellular assembly of [2Fe-2S] clusters in Fdx is susceptible to environmental temperatures. The iron binding form of Fdx (Fe-Fdx) is a precursor during its maturation to a cluster binding form ([2Fe-2S]-Fdx), and reassembly of the [2Fe-2S] clusters during temperature increases is not strictly reliant on other specific iron donors and scaffold proteins within the Isc or Suf system.IMPORTANCE Fdx is an electron carrier that is required for the maturation of many other iron-sulfur proteins. Its function strictly depends on its [2Fe-2S] center that bonds with the cysteinyl S atoms of four cysteine residues within Fdx. However, the assembly mechanism of the [2Fe-2S] clusters in Fdx remains controversial. This study reports that Fdx fails to form its [2Fe-2S] cluster under cold stress conditions but instead binds a single Fe atom at the cluster binding site. Moreover, when temperatures increase, Fdx can assemble clusters by itself from its iron-only binding form in E. coli cells. The possibility remains that Fdx can effectively accept clusters from multiple sources. Nevertheless, our results suggest that Fdx has a strong iron binding activity that contributes to the assembly of its own [2Fe-2S] cluster and that Fdx acts as a temperature sensor to regulate Isc system-mediated iron-sulfur cluster biogenesis.