Transcriptional and Proteomic Analysis of the Aspergillus fumigatus ΔprtT Protease-Deficient Mutant

Aspergillus fumigatus is the most common opportunistic mold pathogen of humans, infecting immunocompromised patients. The fungus invades the lungs and other organs, causing severe damage. Penetration of the pulmonary epithelium is a key step in the infectious process. A. fumigatus produces extracellular proteases to degrade the host structural barriers. The A. fumigatus transcription factor PrtT controls the expression of multiple secreted proteases. PrtT shows similarity to the fungal Gal4-type Zn(2)-Cys(6) DNA-binding domain of several transcription factors. In this work, we further investigate the function of this transcription factor by performing a transcriptional and a proteomic analysis of the ΔprtT mutant. Unexpectedly, microarray analysis revealed that in addition to the expected decrease in protease expression, expression of genes involved in iron uptake and ergosterol synthesis was dramatically decreased in the ΔprtT mutant. A second finding of interest is that deletion of prtT resulted in the upregulation of four secondary metabolite clusters, including genes for the biosynthesis of toxic pseurotin A. Proteomic analysis identified reduced levels of three secreted proteases (ALP1 protease, TppA, AFUA_2G01250) and increased levels of three secreted polysaccharide-degrading enzymes in the ΔprtT mutant possibly in response to its inability to derive sufficient nourishment from protein breakdown. This report highlights the complexity of gene regulation by PrtT, and suggests a potential novel link between the regulation of protease secretion and the control of iron uptake, ergosterol biosynthesis and secondary metabolite production in A. fumigatus

Synthesis, characterization and application of composite derived from rice husk ash with aluminium oxide for sorption of uranium

A composite of rice husk (RH), caustic soda and aluminium oxide was synthesized at 500°C. The activated carbon and amorphous silica dispersed over the aluminium oxide selectively adsorbed uranium in the presence of other elements. At equilibrium time 1 h, phase ratio S/L (0.1 g/10 ml), pH = 5 and uranium initial concentration 120.6 mg/l uranium adsorption efficiency was 96.35%. The uranium stripping efficiency from the load RHA–alumina composite fulfilled 99.9% at 1 h equilibrium time, a phase ratio (S/A) of 0.05 g/10 ml and 0.5 mol/l HNO 3 . The scanning electron microscopy photos revealed that the rice husk ash (RHA)–alumina composite has vacant or regular cavities before the adsorption, and the cavities are fully occupied by uranium after adsorption. The Fourier transform infrared spectroscopy shows a more broadening of the band υ = 3526 and 3462 cm −1 which was ascribed to the uranium adsorption. The composite adsorbed 93.75% of uranium from a waste sample. The uranium adsorption exhibited a Langmuir isotherm

Platelet-Rich Plasma Ameliorates Monosodium Iodoacetate-Induced Ankle Osteoarthritis in the Rat Model via Suppression of Inflammation and Oxidative Stress

Until now, there is no treatment that cause complete cure of the chronic inflammatory and degenerative disease, osteoarthritis (OA). Moreover, the underlying mechanisms of OA development and progress are not fully elucidated, and the present pharmacological treatment alternatives are restricted and associated with adverse side effects. Thus, the present study was conducted to evaluate the role of platelet-rich plasma (PRP) in the remedy of OA in the rat model in terms of inflammation, ankle histopathological alterations, and oxidative stress. OA was induced in male Wistar rats by injection of MIA (2 mg)/50 µL isotonic saline in the right ankle joint for two successive days in each rat. After the 2nd MIA injection, the osteoarthritic rats were allocated into two groups such as the MIA group (group 2) and MIA + PRP group (group 3). The MIA + PRP group was treated with PRP (50 µL) by injection into the ankle joint of the right hind limb of each rat at days 14, 21, and 28 after the 2nd injection of MIA. The same equivalent volume of saline, as a substitute of PRP, was injected into the ankle joint of each rat of the normal control group (group 1) and MIA group (group 2) at the same tested periods. Swelling of joint, bodyweight, total leucocytes count (TLC), and morphological as well as histological changes of ankle joints were evaluated. Serum lipid peroxides (LPO), glutathione (GSH), and glutathione S-transferase (GST) levels were examined as biomarkers of oxidative stress. Serum tumor necrosis factor-α (TNF-α), interleukin-17 (IL-17), and interleukin-4 (IL-4) were investigated by ELISA as biomarkers of inflammation. In addition, magnetic resonance imaging (MRI) was carried out to investigate the soft tissues in joints. The obtained results revealed that PRP reduced LPO and increased GSH and GST levels in osteoarthritic rats. Also, PRP significantly diminished serum TNF-α and IL-17 levels, while it increased IL-4 serum levels in rats with MIA-induced OA. Morphological observations, histological analysis, and MRI revealed a gradual diminishing in joint inflammation and destruction of cartilage in PRP-injected osteoarthritic rats. Based on these results, it can be suggested that PRP has antiarthritic potential in MIA-induced OA, which may be mediated via suppression of inflammation and oxidative stress

Multiple introductions of reassorted highly pathogenic avian influenza viruses (H5N8) clade 2.3.4.4b causing outbreaks in wild birds and poultry in Egypt

Recently, an increased incidence of outbreaks of highly pathogenic avian influenza (HPAI) H5N8 in poultry linked to infected migratory birds has been reported from different European, Asian and African countries. In Egypt, incursion of HPAI H5N8 virus of clade 2.3.4.4b has been recently registered. Full genomic characterization of 3 virus isolates from wild birds and poultry (backyard and commercial farm sectors) showed high nucleotide similarity among the HA, NA, M, and NS gene segments of the three Egyptian HPAI H5N8 viruses, indicating that they are descendants of a common ancestral virus. However, the analyzed Egyptian H5N8 viruses revealed distinct genotypes involving different origins of the PB2, PB1, PA and/or NP segments. In genotype-1 represented by strain A/common-coot/Egypt/CA285/2016 the PB2 and NP segments showed closest relationship to H5N6 and H6N2 viruses, recently detected in Italy. The second is replacement of PB1 and NP genes A novel reassortant, represented by strain A/duck/Egypt/SS19/2017, showed an exchange of PB1 and NP genes which might have originated from H6N8 or H1N1 and H6N2 viruses. Finally, replacement of PA and NP genes characterized strain A/duck/Egypt/F446/2017. Bayesian phylogeographic analyses revealed that Egyptian H5N8 viruses are highly likely derived from Russian 2016 HPAI H5N8 virus (A/great_crested_grebe/Uvs-Nuur_Lake/341/2016 (H5N8)) and the reassortment likely occurred before incursion to Egypt

Top 50 downregulated genes plotted in ascending order by fold change.

<p>Black bars are the iron uptake genes, white bars are genes with various other annotations.</p

Inhibition of the Hypoxia-Inducible Factor 1α-Induced Cardiospecific HERNA1 Enhance-Templated RNA Protects From Heart Disease.

Enhancers are genomic regulatory elements conferring spatiotemporal and signal-dependent control of gene expression. Recent evidence suggests that enhancers can generate noncoding enhancer RNAs, but their (patho)biological functions remain largely elusive. We performed chromatin immunoprecipitation-coupled sequencing of histone marks combined with RNA sequencing of left ventricular biopsies from experimental and genetic mouse models of human cardiac hypertrophy to identify transcripts revealing enhancer localization, conservation with the human genome, and hypoxia-inducible factor 1α dependence. The most promising candidate, hypoxia-inducible enhancer RNA ( HERNA)1, was further examined by investigating its capacity to modulate neighboring coding gene expression by binding to their gene promoters by using chromatin isolation by RNA purification and λN-BoxB tethering-based reporter assays. The role of HERNA1 and its neighboring genes for pathological stress-induced growth and contractile dysfunction, and the therapeutic potential of HERNA1 inhibition was studied in gapmer-mediated loss-of-function studies in vitro using human induced pluripotent stem cell-derived cardiomyocytes and various in vivo models of human pathological cardiac hypertrophy. HERNA1 is robustly induced on pathological stress. Production of HERNA1 is initiated by direct hypoxia-inducible factor 1α binding to a hypoxia-response element in the histoneH3-lysine27acetylation marks-enriched promoter of the enhancer and confers hypoxia responsiveness to nearby genes including synaptotagmin XVII, a member of the family of membrane-trafficking and Ca &lt;sup&gt;2+&lt;/sup&gt; -sensing proteins and SMG1, encoding a phosphatidylinositol 3-kinase-related kinase. Consequently, a substrate of SMG1, ATP-dependent RNA helicase upframeshift 1, is hyperphoshorylated in a HERNA1- and SMG1-dependent manner. In vitro and in vivo inactivation of SMG1 and SYT17 revealed overlapping and distinct roles in modulating cardiac hypertrophy. Finally, in vivo administration of antisense oligonucleotides targeting HERNA1 protected mice from stress-induced pathological hypertrophy. The inhibition of HERNA1 postdisease development reversed left ventricular growth and dysfunction, resulting in increased overall survival. HERNA1 is a novel heart-specific noncoding RNA with key regulatory functions in modulating the growth, metabolic, and contractile gene program in disease, and reveals a molecular target amenable to therapeutic exploitation

qRT-PCR evaluation of transcripts levels from genes that showed a differential expression in the microarray (A) representative genes (<i>MirB, SidA, Erg3, Erg11</i> and <i>Alp1</i>) and (B) the genes <i>HapX</i>, <i>SreA, AcuM</i> and <i>SrbA</i> encoding transcription factors that regulate iron uptake.

<p>Expression rates were normalized relative to mRNA levels of the β-tubulin-encoding gene (AFUA_1G10910) and set arbitrarily to 1 for the WT strain grown in 1% SM medium. Values are given in relative quantity of template compared to the original sample (RQ). RQ values were calculated by use of the equation: RQ  =  2^–ΔΔCT, with ΔΔCT ± SD and ΔCT ± SDs. −Fe  =  growth medium lacking iron and containing 1 mM ferrozine. +Vori  =  growth medium supplemented with a sub-inhibitory concentration (0.125 µg/ml) of voriconazole. The experiment was repeated three times with similar results. Graphs show a representative experiment.</p

Selected enriched downregulated gene classes in the <i>ΔprtT</i> mutant vs. WT¹.

<p>1Top five genes with the highest fold change are shown in each category.</p>2<p>Ergosterol biosynthesis pathway.</p>3<p>Involved in amino acid catabolism.</p