Short and dysfunctional telomeres protect from allergen‐induced airway inflammation

Asthma is a chronic inflammatory disease affecting 300 million people worldwide. As telomere shortening is a well‐established hallmark of aging and that asthma incidence decreases with age, here we aimed to study the role of short telomeres in asthma pathobiology. To this end, wild‐type and telomerase‐deficient mice with short telomeres (third‐generation (G3 Tert ^−/− mice)) were challenged with intranasal house dust mite (HDM) extract. We also challenged with HDM wild‐type mice in which we induced a telomere dysfunction by the administration of 6‐thio‐2´‐deoxyguanosine (6‐thio‐dG). Following HDM exposure, G3 Tert ^−/− and 6‐thio‐dG treated mice exhibited attenuated eosinophil counts and presence of hematopoietic stem cells in the bone marrow, as well as lower levels of IgE and circulating eosinophils. Accordingly, both G3 Tert ^−/− and 6‐thio‐dG treated wild‐type mice displayed reduced airway hyperresponsiveness (AHR), as indicated by decreased airway remodeling and allergic airway inflammation markers in the lung. Furthermore, G3 Tert ^−/− and 6‐thio‐dG treated mice showed lower differentiation of Club cells, attenuating goblet cell hyperplasia. Club cells of G3 Tert ^−/− and 6‐thio‐dG treated mice displayed increased DNA damage and senescence and reduced proliferation. Thus, short/dysfunctional telomeres play a protective role in murine asthma by impeding both AHR and mucus secretion after HDM exposure. Therefore, our findings imply that telomeres play a relevant role in allergen‐induced airway inflammation.     

G6PD overexpression protects from oxidative stress and age‐related hearing loss

Aging of the auditory system is associated with the incremental production of reactive oxygen species (ROS) and the accumulation of oxidative damage in macromolecules, which contributes to cellular malfunction, compromises cell viability, and, ultimately, leads to functional decline. Cellular detoxification relies in part on the production of NADPH, which is an important cofactor for major cellular antioxidant systems. NADPH is produced principally by the housekeeping enzyme glucose‐6‐phosphate dehydrogenase (G6PD), which catalyzes the rate‐limiting step in the pentose phosphate pathway. We show here that G6PD transgenic mice (G6PD‐Tg), which show enhanced constitutive G6PD activity and NADPH production along life, have lower auditory thresholds than wild‐type mice during aging, together with preserved inner hair cell (IHC) and outer hair cell (OHC), OHC innervation, and a conserved number of synapses per IHC. Gene expression of antioxidant enzymes was higher in 3‐month‐old G6PD‐Tg mice than in wild‐type counterparts, whereas the levels of pro‐apoptotic proteins were lower. Consequently, nitration of proteins, mitochondrial damage, and TUNEL⁺ apoptotic cells were all lower in 9‐month‐old G6PD‐Tg than in wild‐type counterparts. Unexpectedly, G6PD overexpression triggered low‐grade inflammation that was effectively resolved in young mice, as shown by the absence of cochlear cellular damage and macrophage infiltration. Our results lead us to propose that NADPH overproduction from an early stage is an efficient mechanism to maintain the balance between the production of ROS and cellular detoxification power along aging and thus prevents hearing loss progression.     

Phospholipase D1 promotes cervical cancer progression by activating the RAS pathway

This study aimed to further investigate the effect of PLD1 on the biological characteristics of human cervical cancer (CC) cell line, CASKI and the potential related molecular mechanism. CRISPR/Cas9 genome editing technology was used to knock out the PLD1 gene in CASKI cells. Cell function assays were performed to evaluate the effect of PLD1 on the biological function of CASKI cells in vivo and in vitro. A PLD1‐overexpression rescue experiment in these knockout cells was performed to further confirm its function. Two PLD1‐knockout CASKI cell lines (named PC‐11 and PC‐40, which carried the ins1/del4 mutation and del1/del2/ins1 mutation, respectively), were constructed by CRISPR/Cas9. PLD1 was overexpressed in these knockout cells (named PC11‐PLD1 and PC40‐PLD1 cells), which rescued the expression of PLD1 by approximately 71.33% and 74.54%, respectively. In vivo, the cell function assay results revealed that compared with wild‐type (WT)‐CASKI cells, the ability of PC‐11 and PC‐40 cells to proliferate, invade and migrate was significantly inhibited. The expression of H‐Ras and phosphorylation of Erk1/2 (p‐Erk1/2) was decreased in PC‐11 and PC‐40 cells compared with WT‐CASKI cells. PC‐11 and PC‐40 cells could sensitize CASKI cells to cisplatin. More importantly, the proliferation, migration and invasion of PC11‐PLD1 and PC40‐PLD1 cells with PLD1 overexpression were significantly improved compared with those of the two types of PLD1 knockout cells. The sensitivity to cisplatin was decreased in PC11‐PLD1 and PC40‐PLD1 cells compared with PC‐11 and PC‐40 cells. In vivo, in the PC‐11 and PC‐40 tumour groups, tumour growth was significantly inhibited and tumour weight (0.95 ± 0.27 g and 0.66 ± 0.43 g vs. 1.59 ± 0.67 g, p = 0.0313 and 0.0108) and volume (1069.41 ± 393.84 and 1077.72 mm³ ± 815.07 vs. 2142.94 ± 577.37 mm³, p = 0.0153 and 0.0128) were significantly reduced compared to those in the WT‐CASKI group. Tumour differentiation of the PC‐11 and PC40 cells was significantly better than that of the WT‐CASKI cells. The immunohistochemistry results confirmed that the expression of H‐Ras and p‐Erk1/2 was decreased in PC‐11 and PC‐40 tumour tissues compared with WT‐CASKI tumour tissues. PLD1 promotes CC progression by activating the RAS pathway. Inhibition of PLD1 may serve as an attractive therapeutic modality for CC.     

A high‐fat diet exacerbates the Alzheimer's disease pathology in the hippocampus of the App NL−F/NL−F knock‐in mouse model

Insulin resistance and diabetes mellitus are major risk factors for Alzheimer''s disease (AD), and studies with transgenic mouse models of AD have provided supportive evidence with some controversies. To overcome potential artifacts derived from transgenes, we used a knock‐in mouse model, App^{NL−F/NL−F} , which accumulates Aβ plaques from 6 months of age and shows mild cognitive impairment at 18 months of age, without the overproduction of APP. In the present study, 6‐month‐old male App^{NL−F/NL−F} and wild‐type mice were fed a regular or high‐fat diet (HFD) for 12 months. HFD treatment caused obesity and impaired glucose tolerance (i.e., T2DM conditions) in both wild‐type and App^{NL−F/NL−F} mice, but only the latter animals exhibited an impaired cognitive function accompanied by marked increases in both Aβ deposition and microgliosis as well as insulin resistance in the hippocampus. Furthermore, HFD‐fed App^{NL−F/NL−F} mice exhibited a significant decrease in volume of the granule cell layer in the dentate gyrus and an increased accumulation of 8‐oxoguanine, an oxidized guanine base, in the nuclei of granule cells. Gene expression profiling by microarrays revealed that the populations of the cell types in hippocampus were not significantly different between the two mouse lines, regardless of the diet. In addition, HFD treatment decreased the expression of the Aβ binding protein transthyretin (TTR) in App^{NL−F/NL−F} mice, suggesting that the depletion of TTR underlies the increased Aβ deposition in the hippocampus of HFD‐fed App^{NL−F/NL−F} mice.     

A Conserved Peptide Pattern from a Widespread Microbial Virulence Factor Triggers Pattern-Induced Immunity in Arabidopsis

Microbe- or host damage-derived patterns mediate activation of pattern-triggered immunity (PTI) in plants. Microbial virulence factor (effector)-triggered immunity (ETI) constitutes a second layer of plant protection against microbial attack. Various necrosis and ethylene-inducing peptide 1 (Nep1)-like proteins (NLPs) produced by bacterial, oomycete and fungal microbes are phytotoxic virulence factors that exert immunogenic activities through phytotoxin-induced host cell damage. We here show that multiple cytotoxic NLPs also carry a pattern of 20 amino acid residues (nlp20) that triggers immunity-associated plant defenses and immunity to microbial infection in Arabidopsis thaliana and related plant species with similar characteristics as the prototype pattern, bacterial flagellin. Characteristic differences in flagellin and nlp20 plant responses exist however, as nlp20s fail to trigger extracellular alkalinization in Arabidopsis cell suspensions and seedling growth inhibition. Immunogenic nlp20 peptide motifs are frequently found in bacterial, oomycete and fungal NLPs. Such an unusually broad taxonomic distribution within three phylogenetic kingdoms is unprecedented among microbe-derived triggers of immune responses in either metazoans or plants. Our findings suggest that cytotoxic NLPs carrying immunogenic nlp20 motifs trigger PTI in two ways as typical patterns and by inflicting host cell damage. We further propose that conserved structures within a microbial virulence factor might have driven the emergence of a plant pattern recognition system mediating PTI. As this is reminiscent of the evolution of immune receptors mediating ETI, our findings support the idea that there is a continuum between PTI and ETI.     

PPARα alleviates iron overload‐induced ferroptosis in mouse liver

Ferroptosis is an iron‐dependent form of non‐apoptotic cell death implicated in liver, brain, kidney, and heart pathology. How ferroptosis is regulated remains poorly understood. Here, we show that PPARα suppresses ferroptosis by promoting the expression of glutathione peroxidase 4 (Gpx4) and by inhibiting the expression of the plasma iron carrier TRF. PPARα directly induces Gpx4 expression by binding to a PPRE element within intron 3. PPARα knockout mice develop more severe iron accumulation and ferroptosis in the liver when fed a high‐iron diet than wild‐type mice. Ferrous iron (Fe²⁺) triggers ferroptosis via Fenton reactions and ROS accumulation. We further find that a rhodamine‐based "turn‐on" fluorescent probe(probe1) is suitable for the in vivo detection of Fe²⁺. Probe1 displays high selectivity towards Fe²⁺, and exhibits a stable response for Fe²⁺ with a concentration of 20 μM in tissue. Our data thus show that PPARα activation alleviates iron overload‐induced ferroptosis in mouse livers through Gpx4 and TRF, suggesting that PPARα may be a promising therapeutic target for drug discovery in ferroptosis‐related tissue injuries. Moreover, we identified a fluorescent probe that specifically labels ferrous ions and can be used to monitor Fe²⁺ in vivo.     

Structural analyses of the malaria parasite aminoacyl‐tRNA synthetases provide new avenues for antimalarial drug discovery

Malaria is a parasitic illness caused by the genus Plasmodium from the apicomplexan phylum. Five plasmodial species of P. falciparum (Pf), P. knowlesi, P. malariae, P. ovale, and P. vivax (Pv) are responsible for causing malaria in humans. According to the World Malaria Report 2020, there were 229 million cases and ~ 0.04 million deaths of which 67% were in children below 5 years of age. While more than 3 billion people are at risk of malaria infection globally, antimalarial drugs are their only option for treatment. Antimalarial drug resistance keeps arising periodically and thus threatens the main line of malaria treatment, emphasizing the need to find new alternatives. The availability of whole genomes of P. falciparum and P. vivax has allowed targeting their unexplored plasmodial enzymes for inhibitor development with a focus on multistage targets that are crucial for parasite viability in both the blood and liver stages. Over the past decades, aminoacyl‐tRNA synthetases (aaRSs) have been explored as anti‐bacterial and anti‐fungal drug targets, and more recently (since 2009) aaRSs are also the focus of antimalarial drug targeting. Here, we dissect the structure‐based knowledge of the most advanced three aaRSs—lysyl‐ (KRS), prolyl‐ (PRS), and phenylalanyl‐ (FRS) synthetases in terms of development of antimalarial drugs. These examples showcase the promising potential of this family of enzymes to provide druggable targets that stall protein synthesis upon inhibition and thereby kill malaria parasites selectively.     

Hepatoprotective effect of nanoniosome loaded Myristica fragrans phenolic compounds in mice‐induced hepatotoxicity

In this study, nanoniosome‐loaded Myristica fragrans'' (MF) phenolic compounds (NLMP) were synthesized and characterized for their physical properties, and hepatoprotective effects on mice with liver toxicity induced by L‐asparaginase (LA) injection. According to the results, NLMP has a spherical shape with a 263 nm diameter, a zeta potential of −26.55 mV and a polydispersity index (PDI) of 0.192. The weight and feed intake of mice induced with hepatotoxicity were significantly (p ≤ 0.05) increased after they were treated with NLMP (2.5 mg/kg body weight of mice). In addition, the blood levels of triglyceride (TG), cholesterol (Chol), liver enzymes (aspartate aminotransferase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP)) and total bilirubin were significantly (p ≤ 0.05) decreased. A significant increase (p ≤ 0.05) in the blood levels of the antioxidant defence system (glutathione peroxidase (GPX), superoxide dismutase (SOD) and catalase (CAT)) were also reported after NLMP treatment. NLMP was also led to a significant decrease (p ≤ 0.05) in inflammatory‐related gene expression of inducible nitric oxide synthase (iNOS) and Interferon‐gamma (IFN‐γ) in the liver, as well as a meaningful (p ≤ 0.05) increase in the expression of SOD as an antioxidant status biomarker. Consequently, the NLMP is recommended as a potential dietary supplement to alleviate the symptoms of LA‐induced hepatotoxicity.     

Parkin drives pS65‐Ub turnover independently of canonical autophagy in Drosophila

Parkinson''s disease‐related proteins, PINK1 and Parkin, act in a common pathway to maintain mitochondrial quality control. While the PINK1‐Parkin pathway can promote autophagic mitochondrial turnover (mitophagy) following mitochondrial toxification in cell culture, alternative quality control pathways are suggested. To analyse the mechanisms by which the PINK1–Parkin pathway operates in vivo, we developed methods to detect Ser65‐phosphorylated ubiquitin (pS65‐Ub) in Drosophila. Exposure to the oxidant paraquat led to robust, Pink1‐dependent pS65‐Ub production, while pS65‐Ub accumulates in unstimulated parkin‐null flies, consistent with blocked degradation. Additionally, we show that pS65‐Ub specifically accumulates on disrupted mitochondria in vivo. Depletion of the core autophagy proteins Atg1, Atg5 and Atg8a did not cause pS65‐Ub accumulation to the same extent as loss of parkin, and overexpression of parkin promoted turnover of both basal and paraquat‐induced pS65‐Ub in an Atg5‐null background. Thus, we have established that pS65‐Ub immunodetection can be used to analyse Pink1‐Parkin function in vivo as an alternative to reporter constructs. Moreover, our findings suggest that the Pink1‐Parkin pathway can promote mitochondrial turnover independently of canonical autophagy in vivo.     

C‐terminal deletion‐induced condensation sequesters AID from IgH targets in immunodeficiency

In activated B cells, activation‐induced cytidine deaminase (AID) generates programmed DNA lesions required for antibody class switch recombination (CSR), which may also threaten genome integrity. AID dynamically shuttles between cytoplasm and nucleus, and the majority stays in the cytoplasm due to active nuclear export mediated by its C‐terminal peptide. In immunodeficient‐patient cells expressing mutant AID lacking its C‐terminus, a catalytically active AID‐delC protein accumulates in the nucleus but nevertheless fails to support CSR. To resolve this apparent paradox, we dissected the function of AID‐delC proteins in the CSR process and found that they cannot efficiently target antibody genes. We demonstrate that AID‐delC proteins form condensates both in vivo and in vitro, dependent on its N‐terminus and on a surface arginine‐rich patch. Co‐expression of AID‐delC and wild‐type AID leads to an unbalanced nuclear AID‐delC/AID ratio, with AID‐delC proteins able to trap wild‐type AID in condensates, resulting in a dominant‐negative phenotype that could contribute to immunodeficiency. The co‐condensation model of mutant and wild‐type proteins could be an alternative explanation for the dominant‐negative effect in genetic disorders.     

Cryo‐EM structures of pentameric autoinducer‐2 exporter from Escherichia coli reveal its transport mechanism

Bacteria utilize small extracellular molecules to communicate in order to collectively coordinate their behaviors in response to the population density. Autoinducer‐2 (AI‐2), a universal molecule for both intra‐ and inter‐species communication, is involved in the regulation of biofilm formation, virulence, motility, chemotaxis, and antibiotic resistance. While many studies have been devoted to understanding the biosynthesis and sensing of AI‐2, very little information is available on its export. The protein TqsA from Escherichia coli, which belongs to the AI‐2 exporter superfamily, has been shown to export AI‐2. Here, we report the cryogenic electron microscopic structures of two AI‐2 exporters (TqsA and YdiK) from E. coli at 3.35 Å and 2.80 Å resolutions, respectively. Our structures suggest that the AI‐2 exporter exists as a homo‐pentameric complex. In silico molecular docking and native mass spectrometry experiments were employed to demonstrate the interaction between AI‐2 and TqsA, and the results highlight the functional importance of two helical hairpins in substrate binding. We propose that each monomer works as an independent functional unit utilizing an elevator‐type transport mechanism.     

Demographic effects of a megafire on a declining prairie grouse in the mixed‐grass prairie

Recent studies have documented benefits of small, prescribed fire and wildfire for grassland‐dependent wildlife, such as lesser prairie‐chickens (Tympanuchus pallidicintus), but wildlife demographic response to the scale and intensity of megafire (wildfire >40,000 ha) in modern, fragmented grasslands remains unknown. Limited available grassland habitat makes it imperative to understand if increasing frequency of megafires could further reduce already declining lesser prairie‐chicken populations, or if historical evolutionary interactions with fire make lesser prairie‐chickens resilient. To evaluate lesser prairie‐chicken demographic response to megafires, we compared lek counts, nest density, and survival rates of adults, nests, and chicks before (2014–2016) and after (2018–2020) a 2017 megafire in the mixed‐grass prairie of Kansas, USA (Starbuck fire ~254,000 ha). There was a 67% decline in attending males on leks post‐fire and a 57% decline in occupied leks post‐fire. Despite population declines as indicated by lek counts, adult female breeding season survival (S^) was similar pre‐ (S^ = 0.65 ± 0.08 [SE]) and post‐fire (0.61 ± 0.08), as was chick survival (pre‐fire: 0.23 ± 0.07; post‐fire: 0.27 ± 0.11). Nest survival appeared lower post‐fire (pre‐fire: 0.38 ± 0.06; post‐fire: 0.20 ± 0.06), but did not differ at the 95% confidence interval. Nest density of marked females declined 73% in areas burned by megafire. Although lesser prairie‐chickens persisted in the study area and we documented minimal effects on most demographic rates, reduced lesser prairie‐chicken abundance and reproductive output suggests full recovery may take >3 years. Increased propensity for megafire resulting from suppression of smaller fires, compounded by climate change and woody encroachment, may impose a short‐term (3–5 year) threat to already declining lesser prairie‐chicken populations.     

Caveolin‐1 negatively regulates inflammation and fibrosis in silicosis

Inhalation of crystalline silica causes silicosis, the most common and serious occupational disease, which is characterized by progressive lung inflammation and fibrosis. Recent studies revealed the anti‐inflammatory and anti‐fibrosis role of Caveolin‐1 (Cav‐1) in lung, but this role in silicosis has not been investigated. Thus, this study evaluated Cav‐1 regulatory effects in silicosis. It was found that Cav‐1 levels were significantly reduced in the lung from silicosis patients and silicotic mice. The silicosis models were established in C57BL/6 (wild‐type) and Cav‐1 deficiency (Cav‐1 ^−/−) mice, and Cav‐1 ^−/− mice displayed wider alveolar septa, increased collagen deposition and more silicotic nodules. The mice peritoneal‐derived macrophages were used to explore the role of Cav‐1 in silica‐induced inflammation, which plays a central role in mechanism of silicosis. Cav‐1 inhibited silica‐induced infiltration of inflammatory cells and secretion of inflammatory factors in vitro and in vivo, partly by downregulating NF‐κB pathway. Additionally, silica uptake and expression of 4‐hydroxynonenal in silicotic mice were observed, and it was found that Cav‐1 absence triggered excessive silica deposition, causing a stronger oxidative stress response. These findings demonstrate the protective effects of Cav‐1 in silica‐induced lung injury, suggesting its potential therapeutic value in silicosis.     

Prohibitin depletion extends lifespan of a TORC2/SGK‐1 mutant through autophagy and the mitochondrial UPR

Mitochondrial prohibitins (PHB) are highly conserved proteins with a peculiar effect on lifespan. While PHB depletion shortens lifespan of wild‐type animals, it enhances longevity of a plethora of metabolically compromised mutants, including target of rapamycin complex 2 (TORC2) mutants sgk‐1 and rict‐1. Here, we show that sgk‐1 mutants have impaired mitochondrial homeostasis, lipogenesis and yolk formation, plausibly due to alterations in membrane lipid and sterol homeostasis. Remarkably, all these features are suppressed by PHB depletion. Our analysis shows the requirement of SRBP1/SBP‐1 for the lifespan extension of sgk‐1 mutants and the further extension conferred by PHB depletion. Moreover, although the mitochondrial unfolded protein response (UPR^mt) and autophagy are induced in sgk‐1 mutants and upon PHB depletion, they are dispensable for lifespan. However, the enhanced longevity caused by PHB depletion in sgk‐1 mutants requires both, the UPR^mt and autophagy, but not mitophagy. We hypothesize that UPR^mt induction upon PHB depletion extends lifespan of sgk‐1 mutants through autophagy and probably modulation of lipid metabolism.     

Attenuation of SARS‐CoV‐2 replication and associated inflammation by concomitant targeting of viral and host cap 2'‐O‐ribose methyltransferases

The SARS‐CoV‐2 infection cycle is a multistage process that relies on functional interactions between the host and the pathogen. Here, we repurposed antiviral drugs against both viral and host enzymes to pharmaceutically block methylation of the viral RNA 2''‐O‐ribose cap needed for viral immune escape. We find that the host cap 2''‐O‐ribose methyltransferase MTr1 can compensate for loss of viral NSP16 methyltransferase in facilitating virus replication. Concomitant inhibition of MTr1 and NSP16 efficiently suppresses SARS‐CoV‐2 replication. Using in silico target‐based drug screening, we identify a bispecific MTr1/NSP16 inhibitor with anti‐SARS‐CoV‐2 activity in vitro and in vivo but with unfavorable side effects. We further show antiviral activity of inhibitors that target independent stages of the host SAM cycle providing the methyltransferase co‐substrate. In particular, the adenosylhomocysteinase (AHCY) inhibitor DZNep is antiviral in in vitro, in ex vivo, and in a mouse infection model and synergizes with existing COVID‐19 treatments. Moreover, DZNep exhibits a strong immunomodulatory effect curbing infection‐induced hyperinflammation and reduces lung fibrosis markers ex vivo. Thus, multispecific and metabolic MTase inhibitors constitute yet unexplored treatment options against COVID‐19.     

Gastric stem cells promote inflammation and gland remodeling in response to Helicobacter pylori via Rspo3‐Lgr4 axis

Helicobacter pylori is a pathogen that colonizes the stomach and causes chronic gastritis. Helicobacter pylori can colonize deep inside gastric glands, triggering increased R‐spondin 3 (Rspo3) signaling. This causes an expansion of the “gland base module,” which consists of self‐renewing stem cells and antimicrobial secretory cells and results in gland hyperplasia. The contribution of Rspo3 receptors Lgr4 and Lgr5 is not well explored. Here, we identified that Lgr4 regulates Lgr5 expression and is required for H. pylori‐induced hyperplasia and inflammation, while Lgr5 alone is not. Using conditional knockout mice, we reveal that R‐spondin signaling via Lgr4 drives proliferation of stem cells and also induces NF‐κB activity in the proliferative stem cells. Upon exposure to H. pylori, the Lgr4‐driven NF‐κB activation is responsible for the expansion of the gland base module and simultaneously enables chemokine expression in stem cells, resulting in gland hyperplasia and neutrophil recruitment. This demonstrates a connection between R‐spondin‐Lgr and NF‐κB signaling that links epithelial stem cell behavior and inflammatory responses to gland‐invading H. pylori.