Design and study of metal complexes with tetrabranched peptide systems as surrogates of superoxide dismutase

Enzymes offer numerous advantages as catalysts, such as selectivity and stereocontrol. However, their practical use is constrained by various limitations, including low thermal stability, low tolerance to diverse experimental conditions, and expensive processes of preparation and purification. To address these challenges, the development of enzyme mimics aims to mitigate these weaknesses. Enzyme mimics (EMs) are typically developed to replicate the binding and catalytic functions of natural enzymes, employing two primary methods: either mimicking enzyme activity through metal complexes with comparable properties, or reproducing the structure of the enzyme active site by means of suitable functional groups, such as oligopeptides [1,2]. Our approach in designing novel enzyme mimics integrates both strategies and relies on synthetic branched peptides to generate a previously unexplored category of EMs. A biocompatible central scaffold serves as the core of the EM structure, to which various oligopeptides can be attached (one for each maleimide chain, Figure 1) [3,4]. In this work we aimed at replicating the catalytic sites of metalloenzymes by introducing specific amino acid sequences capable of binding active metal ions. For instance, various surrogates of Cu/Zn-SOD, Fe/Mn-SOD, and Ni-SOD can be synthesized. Thermodynamic, spectroscopic and structural studies of single peptides and/or tetrabranched systems and their 100 metal complexes (e.g. Cu, Mn, Ni) will be carried out, together with the investigation of their redox behaviour and catalytic activity. Financial support of the National Recovery and Resilience Plan (NRRP), Mission 4 Component 2 Investment 1.1 – NextGenerationEU (PRIN PNRR 2022 - P2022EMY52) is gratefully acknowledged

Identification and characterisation of fosfomycin resistance in Escherichia coli urinary tract infection isolates from Australia

© 2020 Elsevier Ltd Of 1033 Escherichia coli urinary tract infection isolates collected from females >12 years of age in Australia in 2019, only 2 isolates were resistant to fosfomycin with a minimum inhibitory concentration (MIC) of >256 mg/L. Despite having different multilocus sequence types, the two isolates harboured an identical plasmid-encoded fosA4 gene. The fosA4 gene has previously been identified in a single clinical E. coli isolate cultured in Japan in 2014. Each fosfomycin-resistant isolate harboured two conjugative plasmids that possessed an array of genes conferring resistance to aminoglycosides, β-lactams, macrolides, quinolones, sulfonamides and/or trimethoprim

Influence of the allelic variants encoded at the Gli-B1 locus, responsible for a major allergen of wheat, on IgE reactivity for patients suffering from food allergy to wheat

Wheat presents an important genetic diversity that could be useful to look for cultivars with reduced allergencity. omega5-Gliadins have been described as major allergens for wheat allergic patients suffering from wheat-dependent exercise-induced anaphylaxis (WDEIA) and some cases of chronic urticaria (U). Our objective was to study the influence of genetic variability at the Gli-B1 locus encoding for omega5-gliadins on the reactivity of IgE antibodies from these patients. We selected cultivars expressing 13 alleles at Gli-B1 including a wheat/rye translocation and studied the reactivity to gliadins of a rabbit antiserum specific for omega5-gliadins and of IgE from 10 patients. The antiserum and IgE from nine patients with WDEIA and U strongly detected omega5-gliadins expressed by most of the Gli-B1 alleles but showed no or faint responses to the gliadins and secalins extracted from the translocated wheat. The selection of genotypes lacking the Gli-B1 locus may reduce wheat allergenicity

A direct link between MITF, innate immunity, and hair graying

<div><p>Melanocyte stem cells (McSCs) and mouse models of hair graying serve as useful systems to uncover mechanisms involved in stem cell self-renewal and the maintenance of regenerating tissues. Interested in assessing genetic variants that influence McSC maintenance, we found previously that heterozygosity for the melanogenesis associated transcription factor, <i>Mitf</i>, exacerbates McSC differentiation and hair graying in mice that are predisposed for this phenotype. Based on transcriptome and molecular analyses of <i>Mitf</i>^{<i>mi-vga9/+</i>} mice, we report a novel role for MITF in the regulation of systemic innate immune gene expression. We also demonstrate that the viral mimic poly(I:C) is sufficient to expose genetic susceptibility to hair graying. These observations point to a critical suppressor of innate immunity, the consequences of innate immune dysregulation on pigmentation, both of which may have implications in the autoimmune, depigmenting disease, vitiligo.</p></div

FACS isolation of adult McSCs.

<p><b>(A)</b> Immunolabeling for KIT protein in mouse skin at 8 weeks of age. At this time point, the majority of hairs exist in the telogen hair stage, and McSCs that are positive for both KIT (green) and DCT (red) are observed in the hair bulge (arrow) and secondary hair germ (arrowheads). The dotted white lines outline the hair follicles. <b>(B)</b> FACS of dermal cells from 8-week-old mice produces two KIT+ populations. McSCs are CD45− and mast cells are CD45+. The FACS gating strategy (center fluorescence plot) used to isolate McSCs successfully separates each of these cell types and is confirmed by visualizing their distinct morphologies; McSCs are small and often bipolar (left phase images), while mast cells are large and rough looking (right phase image). <b>(C)</b> KIT+/CD45− cells isolated by FACS were placed in culture and assessed for their expression of KIT and DCT by immunolabeling over a 5-day period. Total cells were identified by the nuclear marker DAPI. The table shows the percentage and total number (in parentheses) of cells exhibiting the indicated staining pattern. This FACS protocol produces a relatively pure McSC population, with >92% of cells being DCT+ 1 day after sorting. <b>(D)</b> FACS-isolated KIT+/CD45− McSCs remain positive for the melanocyte markers KIT (green) and DCT (red) and exhibit melanocyte-like traits while in tissue culture. These cells progress from being round at 1 day, to slightly spread at 3 days, to dendritic and pigmented over 5 days. <b>(E)</b> Evaluation of the indicated gates (boxes) on FACS fluorescence plots confirms that there is no significant difference by <i>t</i> test when comparing the percentage of each cell population between wild-type (left plot) and <i>Mitf</i>^{<i>mi-vga9/+</i>} (right plot) dermal cell suspensions (KIT+/CD45−, <i>p</i> = 0.85; KIT+/CD45+, <i>p</i> = 0.14; KIT−/CD45+, <i>p</i> = 0.28). Percentages are represented as the mean ± standard deviation, with <i>n</i> = 3 sorts per genotype. The raw data used to generate these graphs are available in <b><a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2003648#pbio.2003648.s001" target="_blank">S1 Data</a></b>. APC, allophycocyanin; CD45, cluster of differentiation 45; DCT, dopachrome tautomerase; FACS, fluorescence-activated cell sorting; FITC, fluorescein; McSC, melanocyte stem cell; <i>Mitf</i>, melanogenesis associated transcription factor; Neg, negative; PIG, pigment.</p

Relationship of MITF ChIP-seq peaks to the transcription start site (TSS) of innate immune genes.

<p>Relationship of MITF ChIP-seq peaks to the transcription start site (TSS) of innate immune genes.</p

Treatment with the viral mimic poly(I:C) worsens hair graying in Tg(Dct-Sox10)/0 susceptible mice.

<p><b>(A)</b> Wild-type (left) and Tg(Dct-Sox10)/0 (right) mice were plucked along their lower back (the plucked region is indicated by the dashed boxes) to synchronize and initiate the hair cycle in this area. Starting on the fourth day after plucking, mice received an intraperitoneal injection on three consecutive days of 100 μL of physiological water alone (vehicle) or physiological water containing 100 μg of poly(I:C). Hair regrowth was allowed to proceed normally and the mice were imaged within the same hair cycle, approximately 21 days after the initial plucking. Images shown here are representative of 3–5 animals for each genotype and treatment. <b>(B, C)</b> Histograms of the percent of hairs exhibiting DCT+ McSCs in the hair bulge (B) or DCT+ melanocytes in the hair bulb (C) of mice treated similarly to those described in (A). For this experiment, skins were harvested on the seventh day after plucking, sectioned, and immunolabeled for DCT. For the wild-type mice, each histogram represents 75 hairs evaluated across 3 animals for each anatomic location. For the Tg(Dct-Sox10)/0 mice, each histogram represents >190 hairs evaluated across 6 animals for each anatomic location. Histogram distributions were assessed for significance using the Kolmogorov-Smirnov test. <b>(D)</b> Quantification of pigmented and not pigmented DCT+ McSCs within the hair bulge of mice treated and prepared similarly to those described in (B). For the Tg(Dct-Sox10)/0 mice injected with the vehicle control, 256 McSCs were assessed across 3 animals. For the Tg(Dct-Sox10)/0 mice injected with poly(I:C), 260 McSCs were assessed across 5 animals. The difference in the proportion of pigmented and not pigmented DCT+ McSCs between treatments was tested for significance using the Fisher’s exact test (<i>p</i> = 0.001). The raw data used to generate the graphs in (B), (C), and (D) are available in <b><a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2003648#pbio.2003648.s001" target="_blank">S1 Data</a></b>. DCT, dopachrome tautomerase; McSC, melanocyte stem cell; poly(I:C); polyinosinic:polycytidylic acid.</p

Increased hair graying and McSC differentiation occurs in Tg(Dct-Sox10) animals that are haploinsufficient for <i>Mitf</i>^{<i>mi-vga9/+</i>}.

<p><b>(A)</b> Dorsal images of two female littermates taken at P110. At this age, the coat of Tg(Dct-Sox10)/0 mice appears solid black, while the coat of Tg(Dct-Sox10)/0; <i>Mitf</i>^{<i>mi-vga9/+</i>} mice exhibits a small number of gray hairs along its back. Bright-field images of histological sections of the skins from these mice reveal increased ectopic pigmentation (arrows) within the hair bulge (region between the dotted lines) of hairs from Tg(Dct-Sox10)/0; <i>Mitf</i>^{<i>mi-vga9/+</i>} animals in comparison to Tg(Dct-Sox10)/0 animals. <b>(B)</b> Dorsal images of two female littermates taken at P37 and P110. At P37, Tg(Dct-Sox10)/Tg(Dct-Sox10) mice exhibit a black coat with congenital white spotting on the belly and back. These mice experience progressive hair graying and by P110 exhibit a “salt and pepper” coat color in regions of the back fur that were originally black. In comparison to Tg(Dct-Sox10)/Tg(Dct-Sox10) mice, hair graying in Tg(Dct-Sox10)/Tg(Dct-Sox10); <i>Mitf</i>^{<i>mi-vga9/+</i>} mice at P110 is more severe, with a majority of the back fur exhibiting gray hairs. The images for this figure were generated in the original study presented in [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2003648#pbio.2003648.ref017" target="_blank">17</a>]. McSC, melanocyte stem cell; P, postnatal day; SG, sebaceous gland.</p