Investigating the Role of Iron-Sulfur Signaling in Yeast Transcriptional Regulation

As an essential cofactor in a myriad of cellular processes, uptake and mobilization of iron must be tightly controlled. Iron homeostasis in fungi involves balancing iron uptake and storage with iron utilization to achieve adequate, non-toxic levels of this essential nutrient. Extensive work in the non-pathogenic yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe have uncovered unique iron regulation networks for each organism that control iron metabolism via distinct molecular mechanisms. The activities of all fungal iron-sensing transcription factors characterized to date are regulated via iron-sulfur cluster signaling. In Saccharomyces cerevisiae, expression of iron uptake, utilization and storage genes is primarily regulated by transcriptional activators Aft1 (and its paralogue Aft2) and Yap5. Upon iron repletion, a mitochondrially generated Fe-S cluster is delivered to Aft1/Aft2 by a cytosolic complex comprising Grx3/4 and Fra2, leading to the dimerization and eventual export of Aft1/2 to the cytosol. Genetic studies suggest that the protein Fra1 may also have a role in regulating Aft1/2 activity; however, its specific role is unknown. Deletion of either Fra1 or Fra2 in addition to deletion of the vacuolar iron importer Ccc1 restores iron-responsive control of the iron regulon despite the absence of Fra2 or Fra1. Before testing for possible alternate mechanisms for this restoration, we wanted to understand if the mitochondrial iron-sulfur (Fe-S) biogenesis machinery was contributing to inhibition of Aft1/2 in the strains in question. β-galactosidase reporter assay results show that in the absence of a functional mitochondrial iron-sulfur cluster biogenesis machinery (isu1Δ mutants), the iron regulon is constitutively active in all strains except for ccc1Δfra1Δisu1Δ, suggesting that the mitochondria generated inhibitory cluster is not required for inhibition of Aft1 and Aft2 in this mutant strain. We employed a genetic screen approach to corroborate the in vitro evidence of inhibitory cluster transfer from the cytosolic complex comprising Grx3-Fra2 to Aft1 and Aft2. With the help of error-prone PCR, we successfully created mutated pools of FRA1 and FRA2. Our goal was to isolate point mutations in each of these genes which would result in constitutive inhibition of the iron regulon via Aft1 and Aft2, and once identified, to test the effect of those mutations on inhibitory cluster transfer to Aft1 and Aft2 in vitro. The isolated FRA1 and FRA2 mutants were non-informative non-sense or missense mutations that led to probable defective protein yielding false positive phenotypes in the genetic screen. Iron homeostasis in Schizosaccharomyces pombe is maintained via transcriptional repression of iron uptake and iron utilization genes. The GATA-type transcriptional repressor Fep1 binds to the promoters of iron uptake and transport genes under iron replete conditions, turning off their expression to avoid iron overload. Fep1 function is controlled at the post-translational level by the cytosolic CGFS glutaredoxin Grx4 and its binding partner Fra2. Biochemical and spectroscopic characterization of Fep1 suggests that it bears a [Fe-S] cofactor and forms a [2Fe-2S]-bound heterocomplex with Grx4 as well as Grx4-Fra2. Monothiol glutaredoxins along with BolA like proteins (Fra2) have been repeatedly shown to be associated with the delivery and transfer of Fe-S clusters to proteins and thus effective communicators of the cellular iron status to iron-responsive transcription factors

IDENTIFICATION OF AMPC Î’-LACTAMASE-PRODUCING CLINICAL ISOLATES OF ESCHERICHIA COLI

  Objective: Indiscriminate use of β-lactam antibiotics has resulted in the emergence of β-lactamase enzymes. AmpC β-lactamases, in particular, confer resistance to penicillin, first-, second-, and third-generation cephalosporins as well as monobactams and are responsible for antibiotic resistance in nosocomial pathogens. Therefore, this study was undertaken to screen nosocomial Escherichia coli isolates for the presence and characterization of AmpC β-lactamases. The study also envisaged on the detection of inducible AmpC β-lactamases and extended-spectrum β-lactamases (ESBLs) in AmpC β-lactamase-producing E. coli.Methods: A total of 102 clinical isolates of E. coli, were subjected to cefoxitin screening, and screen-positive isolates were further subjected to inhibitor-based detection method, phenotypic confirmatory test, disc antagonism test, polymerase chain reaction (PCR), and isoelectric focusing (IEF).Results: In this study, 33% of E. coli were resistant to cefoxitin, of which 35% were found to be positive for AmpC β-lactamase by inhibitor-based phenotypic test. Of the AmpC-positive isolates, 83% were positive for ESBLs, whereas 25% were producing inducible AmpC β-lactamases. PCR and IEF showed CIT and EBC types of AmpC β-lactamases present in the tested isolates.Conclusion: Our study showed the presence of inducible AmpC enzymes and ESBLs in E. coli isolates and PCR identified more isolates to be AmpC producers

Predictors of candidemia in pediatric patients (0–12 years) admitted in a tertiary care hospital of Northern India

Background: Bloodstream infections due to Candida species are becoming a major cause of morbidity and mortality in hospitalized patients. The emergence of non-albicans Candida (NACs) species with lesser susceptibility to antifungals has added to the woes of clinicians. Objectives: The objectives of the study were to determine the clinical and laboratory predictors and microbiological profile of candidemia in pediatric patients. Materials and Methods: This is a hospital-based, prospective, and cross-sectional study conducted in the pediatric department of a tertiary care hospital. A total of 250 children aged 0–12 years with risk factors for fungal sepsis were enrolled. Demographic details, clinical, and laboratory parameters were noted and samples were sent for culture. Cultures yielding growth of Candida were included in the study, and antifungal susceptibility performed. Associations were assessed using Chi-square test first and then through logistic regression models. Results: Among the 250 patients with risk factors for fungal sepsis, 47 patients (18.8%) with culture proven candidemia were identified. Predictors of candidemia among neonates were prematurity (<30 weeks), prolonged ventilation (>72 h), and thrombocytopenia. Among pediatric patients, prolonged steroid intake, Candida isolation from sites other than blood and persistent neutropenia, were significantly associated with the candidemia. NAC species were the predominant isolates (78.7%). Conclusion: Candidemia should be suspected in premature neonates requiring prolonged ventilation with unexplained thrombocytopenia. Among pediatric patients, prolonged steroid intake, Candida isolation from sites other than blood and persistent neutropenia are predictors of candidemia

CATALYTIC CRACKING OF TOLUENE USING RED MUD: HYDROGEN EVOLUTION AND CARBON FORMATION.

Toluene has been used as a model tar compound in many cracking and reforming studies in the context of biomass gasification and pyrolysis. Low cost alternate catalysts such as biochar, limonite have been studied. Toluene has also been used in chemical vapour deposition of carbon nano tubes. In this study, red mud waste from the aluminium industry without any pre treatment was used as a catalyst for toluene cracking. The hydrogen evolution and nano carbon formation were studied. The reduction behaviour of the catalyst in hydrogen showed conversion of iron hydroxide and oxide to iron above 700 ?C. Hydrogen formation rate was maximum at 800 ?C after around 7 h and subsequently reduced with time. Depending on the carbon content, the post reaction catalyst was comprised of reduced iron oxides (magnetite, wustite) and mixture of iron, iron carbide and graphitic carbon. Carbon content up to 46% was obtained and the deposited carbon was in the form of nano fibers. While red mud has been used in catalytic steam gasification and pyrolysis of biomass, the hydrogen evolution using red mud and toluene has not been reported

A Computational Approach Identifies Immunogenic Features of Prognosis in Human Cancers

A large number of tumor intrinsic and extrinsic factors determine long-term survival in human cancers. In this study, we stratified 9120 tumors from 33 cancers with respect to their immune cell content and identified immunogenomic features associated with long-term survival. Our analysis demonstrates that tumors infiltrated by CD8+ T cells expressing higher levels of activation marker (PD1hi) along with TCR signaling genes and cytolytic T cell markers (IL2hi/TNF-αhi/IFN-γhi/GZMA-Bhi) extend survival, whereas survival benefit was absent for tumors infiltrated by anergic and hyperexhausted CD8+ T cells characterized by high expression of CTLA-4, TIM3, LAG3, and genes linked to PI3K signaling pathway. The computational approach of using robust and highly specific gene expression signatures to deconvolute the tumor microenvironment has important clinical applications, such as selecting patients who will benefit from checkpoint inhibitor treatment

STUDY OF DIFFERENT TREATMENT METHODS ON CHICKEN FEATHER BIOMASS

The chicken feathers (CFs)  consist of up to 10 % of total chicken dry mass and they have many potential industrial applications. CFs contains protein fibers named as keratin, which is an insoluble protein. Primary sanitization phases are complex because of the presence of lots of blood born microbes, pathogens and parasites in raw biomass. The extraction process of keratins from the unprocessed feathers is also a challenging task. Prior to the extraction cleaning/sanitization of feathers is a very necessary step. Thus, the present work was conducted to optimize  an efficient surfactant  for the cleaning process of the  CFs by using ionic and non-ionic surfactants. The experiment was conducted by the washing of feathers with double distilled water (ddH2O), detergents, ether and lastly with boiling water. The washed feathers treated with surfactants and the effect of each surfactant was analyzed by a microbiological test which tells about the extent of  the presence of different bacteria on the treated feathers. SEM, EDX, FTIR were used to study the morphology and composition of  untreated and treated CFs. SEM showed there was no detectable fiber damage after treatment. Cetrimonium bromide (CTAB) (t3) was one of the best surfactant for the treatment of CFs among all the surfactant used. The present study described the best treatment method  for the CFs.

Fracture Risk in Men With Congestive Heart Failure Risk Reduction With Spironolactone

ObjectivesThe purpose of this study was to determine whether spironolactone use is associated with fractures in men with congestive heart failure (CHF).BackgroundIn rats with aldosteronism, spironolactone preserves skeletal strength. However, in humans, the relationship of spironolactone to fractures is not known.MethodsThe medical records of all male patients with CHF from 1999 to 2005 treated at the Veterans Affairs Medical Center, Memphis, Tennessee, were reviewed (n = 4,735). Odds ratios with 95% confidence intervals of having a fracture associated with spironolactone use were estimated using conditional logistic regression.ResultsWe identified 167 cases with a single-incident fracture and matched these by age and race to 668 control subjects without fractures. After adjustment for covariates, spironolactone use was inversely associated with total fracture (odds ratio: 0.575; 95% confidence interval: 0.346 to 0.955, p = 0.0324).ConclusionsThe use of spironolactone is inversely associated with fractures in men with CHF