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Summary: Stemness and pluripotency are mediated by transcriptional master regulators that promote self-renewal and repress cell differentiation, among which is the high-mobility group (HMG) box transcription factor SOX2. Dysregulated SOX2 expression, by contrast, leads to transcriptional aberrations relevant to oncogenic transformation, cancer progression, metastasis, therapy resistance, and relapse. Here, we report a post-transcriptional mechanism by which the cytosolic pool of SOX2 contributes to these events in an unsuspected manner. Specifically, a low-complexity region within SOX2’s C-terminal segment connects to the ribosome to modulate the expression of cognate downstream factors. Independent of nuclear structures or DNA, this C-terminal functionality alone changes metabolic properties and induces non-adhesive growth when expressed in the cytosol of SOX2 knockout cells. We thus propose a revised model of SOX2 action where nuclear and cytosolic fractions cooperate to impose cell fate decisions via both transcriptional and translational mechanisms.

Abstract The blue wildebeest (Connochaetes taurinus) is a keystone species in savanna ecosystems from southern to eastern Africa, and is well known for its spectacular migrations and locally extreme abundance. In contrast, the black wildebeest (C. gnou) is endemic to southern Africa, barely escaped extinction in the 1900s and is feared to be in danger of genetic swamping from the blue wildebeest. Despite the ecological importance of the wildebeest, there is a lack of understanding of how its unique migratory ecology has affected its gene flow, genetic structure and phylogeography. Here, we analyze whole genomes from 121 blue and 22 black wildebeest across the genus’ range. We find discrete genetic structure consistent with the morphologically defined subspecies. Unexpectedly, our analyses reveal no signs of recent interspecific admixture, but rather a late Pleistocene introgression of black wildebeest into the southern blue wildebeest populations. Finally, we find that migratory blue wildebeest populations exhibit a combination of long-range panmixia, higher genetic diversity and lower inbreeding levels compared to neighboring populations whose migration has recently been disrupted. These findings provide crucial insights into the evolutionary history of the wildebeest, and tangible genetic evidence for the negative effects of anthropogenic activities on highly migratory ungulates.

Abstract Background Natural cytokines are poorly suited as therapeutics for systemic administration due to suboptimal pharmacological and pharmacokinetic (PK) properties. Recombinant human interleukin-2 (rhIL-2) has shown promise for treatment of autoimmune (AI) disorders yet exhibits short systemic half-life and opposing immune responses that negate an appropriate therapeutic index. Methods A semi-synthetic microbial technology platform was used to engineer a site-specifically pegylated form of rhIL-2 with enhanced PK, specificity for induction of immune-suppressive regulatory CD4 + T cells (Tregs), and reduced stimulation of off-target effector T and NK cells. A library of rhIL-2 molecules was constructed with single site-specific, biorthogonal chemistry-compatible non-canonical amino acids installed near the interface where IL-2 engages its cognate receptor βγ (IL-2Rβγ) signaling complex. Biorthogonal site-specific pegylation and functional screening identified variants that retained engagement of the IL-2Rα chain with attenuated potency at the IL-2Rβγ complex. Results Phenotypic screening in mouse identifies SAR444336 (SAR’336; formerly known as THOR-809), rhIL-2 pegylated at H16, as a potential development candidate that specifically expands peripheral CD4+ Tregs with upregulation of markers that correlate with their suppressive function including FoxP3, ICOS and Helios, yet minimally expands CD8 + T or NK cells. In non-human primate, administration of SAR’336 also induces dose-dependent expansion of Tregs and upregulated suppressive markers without significant expansion of CD8 + T or NK cells. SAR’336 administration reduces inflammation in a delayed-type hypersensitivity mouse model, potently suppressing CD4+ and CD8 + T cell proliferation. Conclusion SAR’336 is a specific Treg activator, supporting its further development for the treatment of AI diseases.

Summary: The inherent ability of melanoma cells to alter the differentiation-associated transcriptional repertoire to evade treatment and facilitate metastatic spread is well accepted and has been termed phenotypic switching. However, how these facets of cellular behavior are controlled remains largely elusive. Here, we show that cysteine availability, whether from lysosomes (CTNS-dependent) or exogenously derived (SLC7A11-dependent or as N-acetylcysteine), controls melanoma differentiation-associated pathways by acting on the melanocyte master regulator MITF. Functional data indicate that low cysteine availability reduces MITF levels and impairs lysosome functions, which affects tumor ferroptosis sensitivity but improves metastatic spread in vivo. Mechanistically, cysteine-restrictive conditions reduce acetyl-CoA levels to decrease p300-mediated H3K27 acetylation at the melanocyte-restricted MITF promoter, thus forming a cysteine feedforward regulation that controls MITF levels and downstream lysosome functions. These findings collectively suggest that cysteine homeostasis governs melanoma differentiation by maintaining MITF levels and lysosome functions, which protect against ferroptosis and limit metastatic spread.

Abstract Several African mammals exhibit a phylogeographic pattern where closely related taxa are split between West/Central and East/Southern Africa, but their evolutionary relationships and histories remain controversial. Bushpigs (Potamochoerus larvatus) and red river hogs (P. porcus) are recognised as separate species due to morphological distinctions, a perceived lack of interbreeding at contact, and putatively old divergence times, but historically, they were considered conspecific. Moreover, the presence of Malagasy bushpigs as the sole large terrestrial mammal shared with the African mainland raises intriguing questions about its origin and arrival in Madagascar. Analyses of 67 whole genomes revealed a genetic continuum between the two species, with putative signatures of historical gene flow, variable F ST values, and a recent divergence time (

Abstract The mechanisms underlying Parkinson’s disease (PD) etiology are only partially understood despite intensive research conducted in the field. Recent evidence suggests that early neurodevelopmental defects might play a role in cellular susceptibility to neurodegeneration. To study the early developmental contribution of GBA mutations in PD we used patient-derived iPSCs carrying a heterozygous N370S mutation in the GBA gene. Patient-specific midbrain organoids displayed GBA-PD relevant phenotypes such as reduction of GCase activity, autophagy impairment, and mitochondrial dysfunction. Genome-scale metabolic (GEM) modeling predicted changes in lipid metabolism which were validated with lipidomics analysis, showing significant differences in the lipidome of GBA-PD. In addition, patient-specific midbrain organoids exhibited a decrease in the number and complexity of dopaminergic neurons. This was accompanied by an increase in the neural progenitor population showing signs of oxidative stress-induced damage and premature cellular senescence. These results provide insights into how GBA mutations may lead to neurodevelopmental defects thereby predisposing to PD pathology.

ObjectiveBi-allelic pathogenic variants in the MVK gene, which encodes mevalonate kinase (MK), an essential enzyme in isoprenoid biosynthesis, cause the autoinflammatory metabolic disorder mevalonate kinase deficiency (MKD). We generated and characterized MK-deficient monocytic THP-1 cells to identify molecular and cellular mechanisms that contribute to the pro-inflammatory phenotype of MKD.MethodsUsing CRISPR/Cas9 genome editing, we generated THP-1 cells with different MK deficiencies mimicking the severe (MKD-MA) and mild end (MKD-HIDS) of the MKD disease spectrum. Following confirmation of previously established disease-specific biochemical hallmarks, we studied the consequences of the different MK deficiencies on LPS-stimulated cytokine release, glycolysis versus oxidative phosphorylation rates, cellular chemotaxis and protein kinase activity.ResultsSimilar to MKD patients’ cells, MK deficiency in the THP-1 cells caused a pro-inflammatory phenotype with a severity correlating with the residual MK protein levels. In the MKD-MA THP-1 cells, MK protein levels were barely detectable, which affected protein prenylation and was accompanied by a profound pro-inflammatory phenotype. This included a markedly increased LPS-stimulated release of pro-inflammatory cytokines and a metabolic switch from oxidative phosphorylation towards glycolysis. We also observed increased activity of protein kinases that are involved in cell migration and proliferation, and in innate and adaptive immune responses. The MKD-HIDS THP-1 cells had approximately 20% residual MK activity and showed a milder phenotype, which manifested mainly upon LPS stimulation or exposure to elevated temperatures.ConclusionMK-deficient THP-1 cells show the biochemical and pro-inflammatory phenotype of MKD and are a good model to study underlying disease mechanisms and therapeutic options of this autoinflammatory disorder.

Abstract While targeted treatment against BRAF(V600E) improve survival for melanoma patients, many will see their cancer recur. Here we provide data indicating that epigenetic suppression of PGC1α defines an aggressive subset of chronic BRAF-inhibitor treated melanomas. A metabolism-centered pharmacological screen further identifies statins (HMGCR inhibitors) as a collateral vulnerability within PGC1α-suppressed BRAF-inhibitor resistant melanomas. Lower PGC1α levels mechanistically causes reduced RAB6B and RAB27A expression, whereby their combined re-expression reverses statin vulnerability. BRAF-inhibitor resistant cells with reduced PGC1α have increased integrin-FAK signaling and improved extracellular matrix detached survival cues that helps explain their increased metastatic ability. Statin treatment blocks cell growth by lowering RAB6B and RAB27A prenylation that reduces their membrane association and affects integrin localization and downstream signaling required for growth. These results suggest that chronic adaptation to BRAF-targeted treatments drive novel collateral metabolic vulnerabilities, and that HMGCR inhibitors may offer a strategy to treat melanomas recurring with suppressed PGC1α expression.

The objective of this research was applying air Micronanobubbles to reduce Arsenic present in an effluent generated at the laboratory level. In this research, an effluent with arsenic metal was used from a mixture of minerals with the presence of arsenopyrite with 19% As content. This mixture of minerals was stirred in a hot alkaline leaching medium at a temperature of 80 °C, with Na2S (1.5 M) and NaOH (1.0 M), then it was diluted with distilled water to obtain a volume of 100 L with a concentration of 150 ppm As. This effluent was treated with the air micronanobubble (MNB) generator equipment patented by Dr. Jhonny Valverde Flores. The variables of the pressure and the insufflation time of air as MNB in the effluent with content of the arsenic element were analyzed. A 40% decrease of As was obtained in the effluent. The results indicate that the use of MNB is promising in terms of the reduction of arsenic in solutions with concentrations of the same.

Abstract Aim To assess the anti-plaque effect of a high concentration sodium bicarbonate dentifrice on plaque formation, and gingivitis, as compared to a control toothpaste, irrespective of individual brushing technique and plaque quality. Methods The experimental gingivitis model, with a split-mouth design was used to assess the anti-plaque effect of a high concentration sodium bicarbonate dentifrice on plaque formation. By producing individual fitted trays, the toothpaste was applied in the test quadrant and a control dentifrice in the contralateral. The participants used the individual fitted trays for 1 min every morning and evening, for 21 days. In this period, the participants was only allowed to brush the teeth in the opposite jaw, as usual. Twenty healthy individuals successfully completed the study. Results At 21 days, there was no statistically significant difference between test quadrant and control quadrant with regard to plaque indices, gingival index and number of bleeding sites. Conclusion This study demonstrated that the high concentration sodium bicarbonate dentifrice used did not produce statistically significant anti-plaque effect compared to the control dentifrice, in terms of Plaque- and Gingival Indices, number of bleeding sites or by Quigely and Hein, the Turesky modification Plaque Index, irrespective of brushing technique and individual plaque quality. Trial registration Regional Committee for Medical Research and Ethics, South-East Norway in 2021 (REK.2021/370116). Clinical Trial Registration: NCT05441371 (First registered 09/06/2022, First posted 01/07/2022) ( http://www.clinicaltrials.gov ). (Retrospectively registered).

In vitro culture systems that structurally model human myogenesis and promote PAX7+ myogenic progenitor maturation have not been established. Here we report that human skeletal muscle organoids can be differentiated from induced pluripotent stem cell lines to contain paraxial mesoderm and neuromesodermal progenitors and develop into organized structures reassembling neural plate border and dermomyotome. Culture conditions instigate neural lineage arrest and promote fetal hypaxial myogenesis toward limb axial anatomical identity, with generation of sustainable uncommitted PAX7 myogenic progenitors and fibroadipogenic (PDGFRa+) progenitor populations equivalent to those from the second trimester of human gestation. Single-cell comparison to human fetal and adult myogenic progenitor /satellite cells reveals distinct molecular signatures for non-dividing myogenic progenitors in activated (CD44High/CD98+/MYOD1+) and dormant (PAX7High/FBN1High/SPRY1High) states. Our approach provides a robust 3D in vitro developmental system for investigating muscle tissue morphogenesis and homeostasis.

A diagnosis is presented for the Centrioncinae, the Afromontane Forest Flies or stalkless Diopsidae, while its taxonomic position within the Diopsidae is discussed. Arguments are presented for an eventual raising of the Centrioncinae to family level. The differential characters for its two genera, Centrioncus Speiser and Teloglabrus Feijen, are tabulated. The diagnosis for Centrioncus is updated and a key to the ten species now recognised (including three new species) is provided. Centrioncus crassifemur sp. nov. is described from a single female from Angola. This greatly extends the distribution range for the genus. Centrioncus bururiensis sp. nov. is described from Burundi, while Centrioncus copelandi sp. nov. originates from the Kasigau Massif of Kenya. Diagnoses, descriptive updates, illustrations and notes are presented for all other Centrioncus. Centrioncus aberrans Feijen, described from Uganda, is now also recorded for western Kenya, Rwanda, and possibly eastern DR Congo. This wide range of C. aberrans is unusual for the Centrioncinae species which have allopatric and usually very restricted distribution ranges. Defining characters of C. aberrans from the various regions were examined in detail, but only minor differences were found. Centrioncus decoronotus Feijen, described from Kenya, is now recorded for several other places in Kenya. A distribution map is given for the Eastern African Centrioncus species. The eastern branch of the Great Rift Valley appears to form a barrier between C. aberrans and C. decoronotus. The type species of the genus, C. prodiopsis Speiser from the Kilimanjaro in Tanzania, was only known from the 1905–1906 type series. After more than 100 years it has now been found again on the Kenya side of Kilimanjaro. Various differential characters of Centrioncus and Diopsidae are discussed, while brief discussions on sex ratio and fungal parasites are given. Centrioncus are known to occur on low shrubs and herbaceous plants in rain forests. Now, the possibility is indicated that they also might occur higher up in the tree canopies.

Molecular biomarkers correspond to all endogenous or exogenous biomolecules, whose presence or alteration in metabolic pathways indicate the response of a biological system to a disturbance caused by internal or external factors. The search for biomarkers has allowed deepening the knowledge of complex pathophysiological mechanisms of diseases, interactions with potential drugs, and the improvement of diagnostic and therapeutic tests, establishing itself as a central point in personalized medicine. In addition, the development of the omics sciences has fostered an increasing exploration in this field of research. This review presents essential and relevant concepts in biomarker research, explores the various definitions and classifications of biomarkers according to their purpose, the critical aspects to be considered in experimental design, the primary employed study models (in vivo, in vitro, and in silico) and the factors that have hindered their ultimate implementation. Finally, important, and current clinical and pharmacological research areas in which biomarker research play a central role are presented.

Peroxisomes are single-membrane bounded organelles that in humans play a dual role in lipid metabolism, including the degradation of very long-chain fatty acids and the synthesis of ether lipids/plasmalogens. The first step in de novo ether lipid synthesis is mediated by the peroxisomal enzyme glyceronephosphate O-acyltransferase, which has a strict substrate specificity reacting only with the long-chain acyl-CoAs. The aim of this study was to determine the origin of these long-chain acyl-CoAs. To this end, we developed a sensitive method for the measurement of de novo ether phospholipid synthesis in cells and, by CRISPR-Cas9 genome editing, generated a series of HeLa cell lines with deficiencies of proteins involved in peroxisomal biogenesis, beta-oxidation, ether lipid synthesis, or metabolite transport. Our results show that the long-chain acyl-CoAs required for the first step of ether lipid synthesis can be imported from the cytosol by the peroxisomal ABCD proteins, in particular ABCD3. Furthermore, we show that these acyl-CoAs can be produced intraperoxisomally by chain shortening of CoA esters of very long-chain fatty acids via beta-oxidation. Our results demonstrate that peroxisomal beta-oxidation and ether lipid synthesis are intimately connected and that the peroxisomal ABC transporters play a crucial role in de novo ether lipid synthesis.

Abstract Western chimpanzees (Pan troglodytes verus) are Critically Endangered and Guinea is a key stronghold for this subspecies. However, Guinea is also rich in minerals with some of the highest‐grade iron‐ore deposits in the world. Specifically, the Nimba Mountains, home to western chimpanzees, is one of the sites under consideration for mining activities. To assess the impact of mining activities in the area, we used non‐invasive genetic sampling to estimate chimpanzee population size, sex ratio, community composition, and range boundaries on the western flank of the massif. The level of genetic diversity and affinity between communities was estimated and recommendations for future genetic censusing provided. Between 2003 and 2018, we collected 999 fecal samples of which 663 were analyzed using a panel of 26 microsatellites. We identified a minimum of 136 chimpanzees in four communities, with evidence of migratory events, a high level of shared ancestry and genetic diversity. We assessed sampling intensities and capture rates for each community. Saturation was reached in two communities with sampling between 3.2 and 4.3 times the estimated number of chimpanzees. Our findings highlight the utility of genetic censusing for temporal monitoring of ape abundance, as well as capturing migratory events and gauging genetic diversity and population viability over time. We recommend genetic sampling, combined with camera trapping, for use in future Environmental and Social Impact Assessments, as these methods can yield robust baselines for implementing the mitigation hierarchy, future biomonitoring and conservation management.

Abstract In recent years, 3D cell culture has been gaining a more widespread following across many fields of biology. Tissue clearing enables optical analysis of intact 3D samples and investigation of molecular and structural mechanisms by homogenizing the refractive indices of tissues to make them nearly transparent. Here, we describe and quantify that common clearing solutions including benzyl alcohol/benzyl benzoate (BABB), PEG-associated solvent system (PEGASOS), immunolabeling-enabled imaging of solvent-cleared organs (iDISCO), clear, unobstructed brain/body imaging cocktails and computational analysis (CUBIC), and ScaleS4 alter the emission spectra of Alexa Fluor fluorophores and fluorescent dyes. Clearing modifies not only the emitted light intensity but also alters the absorption and emission peaks, at times to several tens of nanometers. The resulting shifts depend on the interplay of solvent, fluorophore, and the presence of cells. For biological applications, this increases the risk for unexpected channel crosstalk, as filter sets are usually not optimized for altered fluorophore emission spectra in clearing solutions. This becomes especially problematic in high throughput/high content campaigns, which often rely on multiband excitation to increase acquisition speed. Consequently, researchers relying on clearing in quantitative multiband excitation experiments should crosscheck their fluorescent signal after clearing in order to inform the proper selection of filter sets and fluorophores for analysis.

Tauopathies are a major type of proteinopathies underlying neurodegenerative diseases. Mutations in the tau-encoding MAPT-gene lead to hereditary cases of frontotemporal lobar degeneration (FTLD)-tau, which span a wide phenotypic and pathological spectrum. Some of these mutations, such as the N279K mutation, result in a shift of the physiological 3R/4R ratio towards the more aggregation prone 4R isoform. Other mutations such as V337M cause a decrease in the in vitro affinity of tau to microtubules and a reduced ability to promote microtubule assembly. Whether both mutations address similar downstream signalling cascades remains unclear but is important for potential rescue strategies. Here, we developed a novel and optimised forward programming protocol for the rapid and highly efficient production of pure cultures of glutamatergic cortical neurons from hiPSCs. We apply this protocol to delineate mechanisms of neurodegeneration in an FTLD-tau hiPSC-model consisting of MAPTN279K- or MAPTV337M-mutants and wild-type or isogenic controls. The resulting cortical neurons express MAPT-genotype-dependent dominant proteome clusters regulating apoptosis, ROS homeostasis and mitochondrial function. Related pathways are significantly upregulated in MAPTN279K neurons but not in MAPTV337M neurons or controls. Live cell imaging demonstrates that both MAPT mutations affect excitability of membranes as reflected in spontaneous and stimulus evoked calcium signals when compared to controls, albeit more pronounced in MAPTN279K neurons. These spontaneous calcium oscillations in MAPTN279K neurons triggered mitochondrial hyperpolarisation and fission leading to mitochondrial ROS production, but also ROS production through NOX2 acting together to induce cell death. Importantly, we found that these mechanisms are MAPT mutation-specific and were observed in MAPTN279K neurons, but not in MAPTV337M neurons, supporting a pathological role of the 4R tau isoform in redox disbalance and highlighting MAPT-mutation specific clinicopathological-genetic correlations, which may inform rescue strategies in different MAPT mutations.

Non-invasive biological samples benefit studies that investigate rare, elusive, endangered, or dangerous species. Integrating genomic techniques that use non-invasive biological sampling with advances in computational approaches can benefit and inform wildlife conservation and management. Here, we used non-invasive fecal DNA samples to generate low- to medium-coverage genomes (e.g., >90% of the complete nuclear genome at six X-fold coverage) and metagenomic sequences, combining widely available and accessible DNA collection cards with commonly used DNA extraction and library building approaches. DNA preservation cards are easy to transport and can be stored non-refrigerated, avoiding cumbersome or costly sample methods. The genomic library construction and shotgun sequencing approach did not require enrichment or targeted DNA amplification. The utility and potential of the data generated was demonstrated through genome scale and metagenomic analyses of zoo and free-ranging African savanna elephants (Loxodonta africana). Fecal samples collected from free-ranging individuals contained an average of 12.41% (5.54–21.65%) endogenous elephant DNA. Clustering of these elephants with others from the same geographic region was demonstrated by a principal component analysis of genetic variation using nuclear genome-wide SNPs. Metagenomic analyses identified taxa that included Loxodonta, green plants, fungi, arthropods, bacteria, viruses and archaea, showcasing the utility of this approach for addressing complementary questions based on host-associated DNA, e.g., pathogen and parasite identification. The molecular and bioinformatic analyses presented here contributes towards the expansion and application of genomic techniques to conservation science and practice.

Abstract Background and aims The image-forming methods copper chloride crystallization, capillary dynamolysis and circular chromatography are presented as an instrument for assessing wine quality. Wine quality of samples from a long-term field trial comparing integrated, organic and biodynamic management were investigated by using image-forming methods and sensory analysis. Methods and results Concerning the image-forming methods copper chloride crystallization, capillary dynamolysis and circular chromatography, the images of encoded samples were (i) grouped into pairs with similar image features; (ii) characterized based on reference images (e. g. high–low resistance to degradation); (iii) ranked (according to the characterization), and (iv) assigned to the different production methods (classified). Wine samples from organic and biodynamic management needed less wine per sample for a similar expression of structural characteristics than wine samples from integrated cultivation. Organic and biodynamic samples also show structures that indicate less degeneration than integrated samples. Due to these properties, nine coded wine samples from 2010 could be (i) grouped, (ii) characterized, (iii) ranked and (iv) classified without errors, i.e., assigned to the cultivation methods of integrated, organic and biodynamic agriculture. In sensory analysis, the wine derived from biodynamic management had the highest aroma intensity. In the other parameters the differences were not significant. Conclusion Analysis with the image-forming methods copper chloride crystallization, capillary dynamolysis and circular chromatography complements sensory analysis for a more complete description of the characteristic properties of wines originating from different management systems. Significance of the study If further studies confirm these results, the image-forming methods copper chloride crystallization, capillary dynamolysis and circular chromatography may be developed as a complementary tool to sensory and chemical analysis in assessing wine quality. Graphic abstract

Abstract Background Numerous Ebola virus outbreaks have occurred in Equatorial Africa over the past decades. Besides human fatalities, gorillas and chimpanzees have also succumbed to the fatal virus. The 2004 outbreak at the Odzala-Kokoua National Park (Republic of Congo) alone caused a severe decline in the resident western lowland gorilla (Gorilla gorilla gorilla) population, with a 95% mortality rate. Here, we explore the immediate genetic impact of the Ebola outbreak in the western lowland gorilla population. Results Associations with survivorship were evaluated by utilizing DNA obtained from fecal samples from 16 gorilla individuals declared missing after the outbreak (non-survivors) and 15 individuals observed before and after the epidemic (survivors). We used a target enrichment approach to capture the sequences of 123 genes previously associated with immunology and Ebola virus resistance and additionally analyzed the gut microbiome which could influence the survival after an infection. Our results indicate no changes in the population genetic diversity before and after the Ebola outbreak, and no significant differences in microbial community composition between survivors and non-survivors. However, and despite the low power for an association analysis, we do detect six nominally significant missense mutations in four genes that might be candidate variants associated with an increased chance of survival. Conclusion This study offers the first insight to the genetics of a wild great ape population before and after an Ebola outbreak using target capture experiments from fecal samples, and presents a list of candidate loci that may have facilitated their survival.

Abstract MI-773 is a recently developed small-molecule inhibitor of the mouse double minute 2 (MDM2) proto-oncogene. Preclinical data on the anti-tumour activity of MI-773 are limited and indicate that tumour cell lines (CLs) with mutated TP53 are more resistant to MI-773 than wild type TP53. Here, we explored the compound’s therapeutic potential in vitro using a panel of 274 annotated CLs derived from a diversity of tumours. MI-773 exhibited a pronounced selectivity and moderate potency, with anti-tumour activity in the sub-micromolar range in about 15% of the CLs. The most sensitive tumour types were melanoma, sarcoma, renal and gastric cancers, leukaemia, and lymphoma. A COMPARE analysis showed that the profile of MI-773 was similar to that of Nutlin-3a, the first potent inhibitor of p53–MDM2 interactions, and, in addition, had a superior potency. In contrast, it poorly correlates with profiles of compounds targeting the p53 pathway with another mechanism of action. OMICS analyses confirmed that MI-773 was primarily active in CLs with wild type TP53. In silico biomarker investigations revealed that the TP53 mutation status plus the aggregated expression levels of 11 genes involved in the p53 signalling pathway predicted sensitivity or resistance of CLs to inhibitors of p53–MDM2 interactions reliably. The results obtained for MI-773 could help to refine the selection of cancer patients for therapy.

Abstract Early detection of congenital disorders by newborn screening (NBS) programs is essential to prevent or limit disease manifestation in affected neonates. These programs balance between the detection of the highest number of true cases and the lowest number of false‐positives. In this case report, we describe four unrelated cases with a false‐positive NBS result for very‐long‐chain acyl‐CoA dehydrogenase deficiency (VLCADD). Three neonates presented with decreased but not deficient VLCAD enzyme activity and two of them carried a single heterozygous ACADVL c.1844G>A mutation. Initial biochemical investigations after positive NBS referral in these infants revealed acylcarnitine and organic acid profiles resembling those seen in multiple acyl‐CoA dehydrogenase deficiency (MADD). Genetic analysis did not reveal any pathogenic mutations in the genes encoding the electron transfer flavoprotein (ETF alpha and beta subunits) nor in ETF dehydrogenase. Subsequent further diagnostics revealed decreased levels of riboflavin in the newborns and oral riboflavin administration normalized the MADD‐like biochemical profiles. During pregnancy, the mothers followed a vegan, vegetarian or lactose‐free diet which probably caused alimentary riboflavin deficiency in the neonates. This report demonstrates that a secondary (alimentary) maternal riboflavin deficiency in combination with reduced VLCAD activity in the newborns can result in an abnormal VLCADD/MADD acylcarnitine profile and can cause false‐positive NBS. We hypothesize that maternal riboflavin deficiency contributed to the false‐positive VLCADD neonatal screening results.

Abstract Seventy years following the discovery of peroxisomes, their complete proteome, the peroxi‐ome, remains undefined. Uncovering the peroxi‐ome is crucial for understanding peroxisomal activities and cellular metabolism. We used high‐content microscopy to uncover peroxisomal proteins in the model eukaryote – Saccharomyces cerevisiae. This strategy enabled us to expand the known peroxi‐ome by ~40% and paved the way for performing systematic, whole‐organellar proteome assays. By characterizing the sub‐organellar localization and protein targeting dependencies into the organelle, we unveiled non‐canonical targeting routes. Metabolomic analysis of the peroxi‐ome revealed the role of several newly identified resident enzymes. Importantly, we found a regulatory role of peroxisomes during gluconeogenesis, which is fundamental for understanding cellular metabolism. With the current recognition that peroxisomes play a crucial part in organismal physiology, our approach lays the foundation for deep characterization of peroxisome function in health and disease.

Debaryomyces hansenii is considered an unconventional yeast with a strong biotechnological potential, which can produce and store high amounts of lipids. However, relatively little is known about its lipid metabolism, and genetic tools for this yeast have been limited. The aim of this study was to explore the fatty acid β-oxidation pathway in D. hansenii. To this end, we employed recently developed methods to generate multiple gene deletions and tag open reading frames with GFP in their chromosomal context in this yeast. We found that, similar as in other yeasts, the β-oxidation of fatty acids in D. hansenii was restricted to peroxisomes. We report a series of experiments in D. hansenii and the well-studied yeast Saccharomyces cerevisiae that show that the homeostasis of NAD+ in D. hansenii peroxisomes is dependent upon the peroxisomal membrane protein Pmp47 and two peroxisomal dehydrogenases, Mdh3 and Gpd1, which both export reducing equivalents produced during β-oxidation to the cytosol. Pmp47 is the first identified NAD+ carrier in yeast peroxisomes.

Peroxisomes serve as important centers for cellular redox metabolism and communication. However, fundamental gaps remain in our understanding of how the peroxisomal redox equilibrium is maintained. In particular, very little is known about the function of the nonenzymatic antioxidant glutathione in the peroxisome interior and how the glutathione antioxidant system balances with peroxisomal protein thiols. So far, only one human peroxisomal glutathione-consuming enzyme has been identified: glutathione S-transferase 1 kappa (GSTK1). To study the role of this enzyme in peroxisomal glutathione regulation and function, a GSTK1-deficient HEK-293 cell line was generated and fluorescent redox sensors were used to monitor the intraperoxisomal GSSG/GSH and NAD+/NADH redox couples and NADPH levels. We provide evidence that ablation of GSTK1 does not change the basal intraperoxisomal redox state but significantly extends the recovery period of the peroxisomal glutathione redox sensor po-roGFP2 upon treatment of the cells with thiol-specific oxidants. Given that this delay (i) can be rescued by reintroduction of GSTK1, but not its S16A active site mutant, and (ii) is not observed with a glutaredoxin-tagged version of po-roGFP2, our findings demonstrate that GSTK1 contains GSH-dependent disulfide bond oxidoreductase activity.

The use of synthetic organisms could provide opportunities for discovery and advanced manufacturing of medical drugs. Here the authors use a semi-synthetic organism with an expanded genetic code to generate site-specific chemical modifications in human IL-2.

For the recently established genus Madagopsina (Diopsidae, stalk-eyed flies), Madagopsina makayensis Feijen, Feijen & Feijen, sp. nov. is described from Madagascar. A concise catalogue is given for the genus and an identification key is presented for its six species. The differential character states are listed for the two species groups of the genus: the Madagopsina apollo species group and the Madagopsina apographica species group. The intrageneric relations are discussed based on morphology, geometric morphometrics analysis of wing shape, and allometric data for eye span against body length. Each of these three procedures places the new species in the M. apollo species group with Madagopsina parvapollina as its closest relative. New records are presented for M. apographica and M. parvapollina.

An induced pluripotent stem cell (hiPSC) line (MPIi008-A) was generated from fibroblasts of a 1-year-old male patient with Denys-Drash syndrome using lentiviral delivery of reprogramming factors OCT4, SOX2, KLF4 and c-MYC. The MPIi008-A iPSC line exhibited typical iPSC morphology and normal karyotype, expressed pluripotent stem cell markers, and showed developmental potential to differentiate into derivatives of all three germ layers in vivo. The hiPSC line harbours a heterozygous missense mutation (R394L) in exon 9 of the WT1 gene.

Abstract Background Congenital cytomegalovirus disease (cCMV) is common and can be fatal or cause severe sequelae. Circulating strains of cytomegalovirus carry a high number of variable or disrupted genes. One of these is UL146, a highly diverse gene with 14 distinct genotypes encoding a CXC-chemokine involved in viral dissemination. UL146 genotypes 5 and 6 lack the conserved ELR motif, potentially affecting strain virulence. Here, we investigate whether UL146 genotypes 5 and 6 were associated with congenital CMV infection. Methods Viral DNA was extracted and UL146 sequenced from 116 neonatal dried blood spots (DBS) stored in the Danish National Biobank since 1982 and linked to registered cCMV cases through a personal identifier. These sequences were compared to UL146 control sequences obtained from CMV DNA extracted from 83 urine samples from children with suspected bacterial urinary tract infections. Results Three non-ELR UL146 genotypes (5 and 6) were observed among the cases (2.6%) and two were observed among the controls (2.4%; P > 0.99). Additionally, no significant association with cCMV was found for the other 12 genotypes in a post-hoc analysis, although genotype 8 showed a tendency to be more frequent among cases with 12 observations against three (P = 0.10). All fourteen genotypes were found to have little intra-genotype variation. Viral load, gender, and sample age were not found to be associated with any particular UL146 genotype. Conclusions No particular UL146 genotype was associated with cCMV in this nationwide retrospective case-control study. Associations between CMV disease and disrupted or polymorph CMV genes among immunosuppressed people living with HIV/AIDS and transplant recipients should be investigated in future studies.

Most cardiomyocytes (CMs) in the adult mammalian heart are either binucleated or contain a single polyploid nucleus. Recent studies have shown that polyploidy in CMs plays an important role as an adaptive response to physiological demands and environmental stress and correlates with poor cardiac regenerative ability after injury. However, knowledge about the functional properties of polyploid CMs is limited. In this study, we generated tetraploid pluripotent stem cells (PSCs) by fusion of murine embryonic stem cells (ESCs) and somatic cells isolated from bone marrow or spleen and performed a comparative analysis of the electrophysiological properties of tetraploid fusion-derived PSCs and diploid ESC-derived CMs. Fusion-derived PSCs exhibited characteristics of genuine ESCs and contained a near-tetraploid genome. Ploidy features and marker expression were also retained during the differentiation of fusion-derived cells. Fusion-derived PSCs gave rise to CMs, which were similar to their diploid ESC counterparts in terms of their expression of typical cardiospecific markers, sarcomeric organization, action potential parameters, response to pharmacologic stimulation with various drugs, and expression of functional ion channels. These results suggest that the state of ploidy does not significantly affect the structural and electrophysiological properties of murine PSC-derived CMs. These results extend our knowledge of the functional properties of polyploid CMs and contribute to a better understanding of their biological role in the adult heart.

Abstract The Novel Coronavirus (SARS-CoV-2) was introduced into the United States via travel from Asia and Europe, although the extent of the spread of the disease was limited in the early days of the pandemic. Consequently, international travel may have played a role in the transmission of the disease into Iowa. This study seeks to determine how preferences for international travel changed as novel Coronavirus Disease (COVID-19) spread throughout the world and if any of these returning travelers developed COVID-19 as a result of their trips. This is a retrospective chart review of patients presenting to a travel clinic in Bettendorf, Iowa for pre-travel advice and vaccinations. From October 2019 to March 2020, four hundred twelve (n = 412) patients presented to the clinic. Intended travel to the Western Pacific region (China, Japan, Korea, etc.) decreased dramatically during the study period. All 412 patients were followed in the electronic medical record for the period after their planned travel and only three (3) presented for COVID-19 testing. Two (2) tested positive, and both of these infections were linked to workplace exposures and not due to travel. News of the growing pandemic and travel warnings likely altered patients’ travel plans and decreased travel to the most affected regions of the world in the early months of the COVID-19 pandemic. Based on our study, travel was not a significant source of COVID-19 exposure for patients seen at this clinic.

Diaschisis is a phenomenon observed in stroke that is defined as neuronal dysfunction in regions spared by the infarction but connected to the lesion site.We combined lesion network mapping and task-based functional MRI in 71 patients with post-stroke aphasia to investigate, whether diaschisis and its resolution contribute to early loss and recovery of language functions. Language activation acquired in the acute, subacute and chronic phase was analyzed in compartments with high and low normative resting-state functional connectivity to the lesion site on an individual basis.Regions with high compared to regions with low lesion connectivity showed a steeper increase in language reactivation from the acute to the subacute phase. This finding is compatible with the assumption of resolution of diaschisis. Additionally, language performance in the subacute phase and improvement from the subacute to the chronic phase significantly correlated with the diaschisis effect and its resolution, respectively, suggesting a behavioral relevance of this effect.We therefore assume that undamaged but functionally connected regions become dysfunctional due to missing input from the lesion contributing to the aphasic deficit. Since these regions are structurally intact, dysfunction resolves over time contributing to the rapid early behavioral improvement observed in aphasic stroke patients. Our results demonstrate that diaschisis and its resolution might be a relevant mechanism of early loss and recovery of language function in acute stroke patients.

The transcription factor Oct4 is essential for the maintenance and induction of stem cell pluripotency, but its functional roles are not fully understood. Here, we investigate the functions of Oct4 by depleting and subsequently recovering it in mouse embryonic stem cells (ESCs) and conducting a time-resolved multiomics analysis. Oct4 depletion leads to an immediate loss of its binding to enhancers, accompanied by a decrease in mRNA synthesis from its target genes that are part of the transcriptional network that maintains pluripotency. Gradual decrease of Oct4 binding to enhancers does not immediately change the chromatin accessibility but reduces transcription of enhancers. Conversely, partial recovery of Oct4 expression results in a rapid increase in chromatin accessibility, whereas enhancer transcription does not fully recover. These results indicate different concentration-dependent activities of Oct4. Whereas normal ESC levels of Oct4 are required for transcription of pluripotency enhancers, low levels of Oct4 are sufficient to retain chromatin accessibility, likely together with other factors such as Sox2.

Human cytomegalovirus (HCMV) is a major pathogen in immunocompromised patients. The UL146 gene exists as 14 diverse genotypes among clinical isolates, which encode 14 different CXC chemokines. One genotype (vCXCL1GT1) is a known agonist for CXCR1 and CXCR2, while two others (vCXCL1GT5 and vCXCL1GT6) lack the ELR motif considered crucial for CXCR1 and CXCR2 binding, thus suggesting another receptor targeting profile. To determine the receptor target for vCXCL1GT5, the chemokine was probed in a G protein signaling assay on all 18 classical human chemokine receptors, where CXCR2 was the only receptor being activated. In addition, vCXCL1GT5 recruited β-arrestin in a BRET-based assay and induced migration in a chemotaxis assay through CXCR2, but not CXCR1. In contrast, vCXCL1GT1 stimulated G protein signaling, recruited β-arrestin and induced migration through both CXCR1 and CXCR2. Both vCXCL1GT1 and vCXCL1GT5 induced equally potent and efficacious migration of neutrophils, and ELR vCXCL1GT4 and non-ELR vCXCL1GT6 activated only CXCR2. In contrast to most human chemokines, the 14 UL146 genotypes have remarkably long C-termini. Comparative modeling using Rosetta showed that each genotype could adopt the classic chemokine core structure, and predicted that the extended C-terminal tail of several genotypes (including vCXCL1GT1, vCXCL1GT4, vCXCL1GT5, and vCXCL1GT6) forms a novel β-hairpin not found in human chemokines. Secondary NMR shift and TALOS+ analysis of vCXCL1GT1 supported the existence of two stable β-strands. C-terminal deletion of vCXCL1GT1 resulted in a non-functional protein and in a shift to solvent exposure for tryptophan residues likely due to destabilization of the chemokine fold. The results demonstrate that non-ELR chemokines can activate CXCR2 and suggest that the UL146 chemokines have unique C-terminal structures that stabilize the chemokine fold. Increased knowledge of the structure and interaction partners of the chemokine variants encoded by UL146 is key to understanding why circulating HCMV strains sustain 14 stable genotypes.

ABSTRACT A large number of studies on salinity in hydroponic systems have been carried out, but few of them have evaluated the interaction between salinity and nutrient solution temperatures. Two experiments were carried out in a randomized block design from January to February (Experiment I) and June to July 2018 (Experiment II). Experiment I consisted of treatments with five combinations, as follows: unheated (control) and heated nutrient solution (NS) at a temperature of 32°C using fresh water with an electrical conductivity (ECw) of 0.3 dS m−1, and unheated and heated NS at temperatures of 30 and 32°C using brackish water with an ECw of 6.5 dS m−1 in the main plots, with two coriander cultivars (Tabocas and Verdão) in the subplots, which were grown in the same hydroponic channel. The cultivar Verdão was grown in Experiment II in a 2 × 2 factorial arrangement, consisting of two NS temperatures (unheated and heated at 30°C) and two ECw levels (0.3 and 6.5 dS m−1). Coriander cultivation is feasible with heated NS despite decreases in shoot fresh matter production compared to the control of approximately 37% for Experiment I (cultivar Verdão at a temperature of 32°C and using fresh water) and 17% for Experiment II (temperature of 30°C and using fresh and brackish waters) at 25 days after transplanting.

Embryonic diapause is a state of dormancy with poorly understood mechanisms of embryo intrinsic regulation. Here, the authors show that murine diapause is a dynamic process, where tissue-scale reorganization of the pluripotent lineage is controlled in an autocrine manner by the Wnt/b-catenin/Esrrb signalling cascade.

The literature on Sri Lankan Diopsidae is reviewed. Eight Diopsidae are now known to occur in Sri Lanka, five species in the genus Teleopsis and one species each in the genera Sphyracephala, Diopsis, and Cyrtodiopsis. The presence of Cyrtodiopsis requires confirmation to exclude the possibility of mislabelling. All five Teleopsis species are endemic, as are the Diopsis species and probably the Cyrtodiopsis species. Only Sphyracephala bipunctipennis Senior-White has a larger distribution as it also occurs in India. A key is presented for the Diopsidae of Sri Lanka. Three Teleopsis species were already known to occur in Sri Lanka: T. ferruginea Röder, T. krombeini Feijen and T. maculata Feijen. These species form the T. ferruginea species group. Two new species are now described for this group: Teleopsis neglecta sp. nov. and Teleopsis sorora sp. nov. Teleopsis ferruginea is redescribed, as an earlier redescription turned out to be based on a series of specimens of its sister species T. sorora sp. nov. The other three Diopsidae of Sri Lanka are listed and illustrated. Allometric aspects of the five Teleopsis species are discussed. Three Teleopsis species are sexually dimorphic with regard to eye span, while two species are monomorphic. It is assumed that sexual dimorphism developed independently in the T. ferruginea species group. This brings the number of known cases of independent development of sexual dimorphism in the Diopsidae to ten.

Hydrogen peroxide has been used in agriculture as a way to minimize the negative effects caused by biotic and abiotic stresses on plants. Thus, the objective of this study was to evaluate the mitigating effect of hydrogen peroxide on chlorophyll a fluorescence and growth of soursop subjected to salt stress in the rootstock production phase. The study was conducted in plastic bags under greenhouse conditions. The treatments were distributed in randomized blocks, in a 5 × 2 factorial arrangement, corresponding to five levels of irrigation water electrical conductivity—ECw (0.6; 1.2; 1.8; 2.4 and 3.0 dS m-1) and two concentrations of hydrogen peroxide—H2O2 (0 and 20 μM), with four replicates and two plants per plot. Irrigation water salinity hamper the quantum efficiency of photosystem II in soursop plants, at 120 days after sowing and it inhibits the growth of rootstocks in the period from 80 to 140 days after sowing. Hydrogen peroxide applications at concentration of 20 μM minimized the negative effects of salinity on the soursop initial fluorescence and favored the variable fluorescence and quantum efficiency of PSII.

Abstract Genetic white matter disorders have heterogeneous etiologies and overlapping clinical presentations. We performed a study of the diagnostic efficacy of genome sequencing in 41 unsolved cases with prior exome sequencing, resolving an additional 14 from an historical cohort (n = 191). Reanalysis in the context of novel disease‐associated genes and improved variant curation and annotation resolved 64% of cases. The remaining diagnoses were directly attributable to genome sequencing, including cases with small and large copy number variants (CNVs) and variants in deep intronic and technically difficult regions. Genome sequencing, in combination with other methodologies, achieved a diagnostic yield of 85% in this retrospective cohort.

Policymakers around the world are enforcing mobility restriction policies such as lockdowns, facemask requirements and social distancing to curb the spread of Covid-19. While these policies are effective in preventing the spread of virus, the economic implications are not well understood. We contribute to the literature by examining the impact of these policies on the offline retail sector. Specifically, we measure the effects of these policies on the daily number of shoppers passing by, which we refer to as 'footfall', along major shopping streets in Netherlands. We rely on unique proprietary Wifi data to accurately measure footfall. Our findings imply that all these policies attribute to a non-trivial reduction in footfall levels along shopping streets. While lockdowns led to a 50% reduction in footfall along major shopping streets, shopping streets faced with facemask regulations also experience a 25% drop in human traffic. A reduction in footfall translates into a substantial reduction in retail income of between 12% and 25%.

Peroxisomes are essential organelles involved in various metabolic processes, including fatty acid β-oxidation. Their metabolic functions require a controlled exchange of metabolites and co-factors, including ATP, across the peroxisomal membrane. We investigated which proteins are involved in the peroxisomal uptake of ATP in the yeast Saccharomyces cerevisiae. Using wild-type and targeted deletion strains, we measured ATP-dependent peroxisomal octanoate β-oxidation, intra-peroxisomal ATP levels employing peroxisome-targeted ATP-sensing reporter proteins, and ATP uptake in proteoliposomes prepared from purified peroxisomes. We show that intra-peroxisomal ATP levels are maintained by different peroxisomal membrane proteins each with different modes of action: 1) the previously reported Ant1p protein, which catalyzes the exchange of ATP for AMP or ADP, 2) the ABC transporter protein complex Pxa1p/Pxa2p, which mediates both uni-directional acyl-CoA and ATP uptake, and 3) the mitochondrial Aac2p protein, which catalyzes ATP/ADP exchange and has a dual localization in both mitochondria and peroxisomes. Our results provide compelling evidence for a complementary system for the uptake of ATP in peroxisomes.

Several mechanisms have been attributed to post-stroke loss and recovery of language functions. However, the significance and timing of domain-general and homotopic right-hemispheric activation is controversial. We aimed to examine the effect of left-hemispheric lesion location and time post-stroke on right-hemispheric activation. Voxel-based lesion analyses were informed by auditory language-related fMRI activation of 71 patients with left middle cerebral artery stroke examined longitudinally in the acute, subacute and early chronic phase. Language activation was determined in several right-hemispheric regions of interest and served as regressor of interest for voxel-based lesion analyses. We found that an acute to chronic increase of language activation in the right supplementary motor area was associated with lesions to the left extreme capsule as part of the ventral language pathway. Importantly, this activation increase correlated significantly with improvement of out-of-scanner comprehension abilities. We interpret our findings in terms of successful domain-general compensation in patients with critical left frontotemporal disconnection due to damage to the ventral language pathway but relatively spared cortical language areas.

Abstract The juvenile form of neuronal ceroid Lipofuscinosis (JNCL) is the most common form within this group of rare lysosomal storage disorders, causing pediatric neurodegeneration. The genetic disorder, which is caused by recessive mutations affecting the CLN3 gene, features progressive vision loss, cognitive and motor decline and other psychiatric conditions, seizure episodes, leading to premature death. Animal models have traditionally aid the understanding of the disease mechanisms and pathology and are very relevant for biomarker research and therapeutic testing. Nevertheless, there is a need for establishing reliable and predictive human cellular models to study the disease. Since patient material, particularly from children, is scarce and difficult to obtain, we generated an engineered a CLN3-mutant isogenic human induced pluripotent stem cell (hiPSC) line carrying the c.1054C → T pathologic variant, using state of the art CRISPR/Cas9 technology. To prove the suitability of the isogenic pair to model JNCL, we screened for disease-specific phenotypes in non-neuronal two-dimensional cell culture models as well as in cerebral brain organoids. Our data demonstrates that the sole introduction of the pathogenic variant gives rise to classical hallmarks of JNCL in vitro. Additionally, we discovered an alteration of the splicing caused by this particular mutation. Next, we derived cerebral organoids and used them as a neurodevelopmental model to study the particular effects of the CLN3Q352X mutation during brain formation in the disease context. About half of the mutation -carrying cerebral organoids completely failed to develop normally. The other half, which escaped this severe defect were used for the analysis of more subtle alterations. In these escapers, whole-transcriptome analysis demonstrated early disease signatures, affecting pathways related to development, corticogenesis and synapses. Complementary metabolomics analysis confirmed decreased levels of cerebral tissue metabolites, some particularly relevant for synapse formation and neurotransmission, such as gamma-amino butyric acid (GABA). Our data suggests that a mutation in CLN3 severely affects brain development. Furthermore, before disease onset, disease -associated neurodevelopmental changes, particular concerning synapse formation and function, occur.

Peroxisomes share metabolic pathways with other organelles and peroxisomes are embedded into key cellular processes. However, the specific function of many peroxisomal proteins remains unclear and restricted knowledge of the peroxisomal protein interaction network limits a precise mapping of this network into the cellular metabolism. Inborn peroxisomal disorders are autosomal or X-linked recessive diseases that affect peroxisomal biogenesis (PBD) and/or peroxisomal metabolism. Pathogenic variants in the PEX26 gene lead to peroxisomal disorders of the full Zellweger spectrum continuum. To investigate the phenotypic complexity of PEX26 deficiency, we performed a combined organelle protein interaction screen and network medicine approach and 1) analyzed whether PEX26 establishes interactions with other peroxisomal proteins, 2) deciphered the PEX26 interaction network, 3) determined how PEX26 is involved in further processes of peroxisomal biogenesis and metabolism, and 4) showed how variant-specific disruption of protein-protein interactions (edgetic perturbations) may contribute to phenotypic variability in PEX26 deficient patients. The discovery of 14 novel protein-protein interactions for PEX26 revealed a hub position of PEX26 inside the peroxisomal interactome. Analysis of edgetic perturbations of PEX26 variants revealed a strong correlation between the number of affected protein-protein interactions and the molecular phenotype of matrix protein import. The role of PEX26 in peroxisomal biogenesis was expanded encompassing matrix protein import, division and proliferation, and membrane assembly. Moreover, the PEX26 interaction network intersects with cellular lipid metabolism at different steps. The results of this study expand the knowledge about the function of PEX26 and refine genotype-phenotype correlations, which may contribute to our understanding of the underlying disease mechanism of PEX26 deficiency.

Mevalonate kinase deficiency (MKD) is an autoinflammatory metabolic disorder characterized by life-long recurring episodes of fever and inflammation, often without clear cause. MKD is caused by bi-allelic pathogenic variants in the MVK gene, resulting in a decreased activity of the encoded enzyme mevalonate kinase (MK). MK is an essential enzyme in the isoprenoid biosynthesis pathway, which generates both non-sterol and sterol isoprenoids. The inflammatory symptoms of patients with MKD point to a major role for isoprenoids in the regulation of the innate immune system. In particular a temporary shortage of the non-sterol isoprenoid geranylgeranyl pyrophosphate (GGPP) is increasingly linked with inflammation in MKD. The shortage of GGPP compromises protein prenylation, which is thought to be one of the main causes leading to the inflammatory episodes in MKD. In this review, we discuss current views and the state of knowledge of the pathogenetic mechanisms in MKD, with particular focus on the role of compromised protein prenylation.

The citrus yield is limited by soil and/or water salinity, but appropriate rootstocks can ensure the sustainability of the production system. Therefore, the objective of the present research was to evaluate the salt content in the soil and the production and physiological aspects of the ‘Tahiti’ acid lime combined with thirteen rootstocks, irrigated with saline water in the first two production years to identify indicators of salt tolerance. The rootstocks evaluated were: ‘Santa Cruz Rangpur’ lime, ‘Indio’, ‘Riverside’ and ‘San Diego’ citrandarins, ‘Sunki Tropical’ mandarin, and eight hybrids, obtained from the Citrus Breeding Program of Embrapa Cassava and Fruits. The waters used had three saline levels: 0.14, 2.40, and 4.80 dS m−1, in a randomized block adopting a split-plot design, with rootstocks in the plots and saline waters in the subplots, with four replicates. From August 2019 to February 2021, fruit harvests and agronomic traits were measured. At the end of each production year, the soil characteristics, leaf gas exchange, and chlorophyll a fluorescence analysis were performed. It was concluded that: (1) the effects of water salinity on citrus are of osmotic nature, reducing gas exchange, (2) the salinity did not significantly damage the photosynthetic apparatus until the second year of production, and (3) using more stable, salt-tolerant rootstocks makes it possible to cultivate ‘Tahiti’ acid lime under irrigation with waters of 2.4 dS m−1 electrical conductivity.

Toxicity testing is a crucial step in the development and approval of chemical compounds for human contact and consumption. However, existing model systems often fall short in their prediction of human toxicity in vivo because they may not sufficiently recapitulate human physiology. The complexity of three-dimensional (3D) human organ-like cell culture systems (“organoids”) can generate potentially more relevant models of human physiology and disease, including toxicity predictions. However, so far, the inherent biological heterogeneity and cumbersome generation and analysis of organoids has rendered efficient, unbiased, high throughput evaluation of toxic effects in these systems challenging. Recent advances in both standardization and quantitative fluorescent imaging enabled us to dissect the toxicities of compound exposure to separate cellular subpopulations within human organoids at the single-cell level in a framework that is compatible with high throughput approaches. Screening a library of 84 compounds in standardized human automated midbrain organoids (AMOs) generated from two independent cell lines correctly recognized known nigrostriatal toxicants. This approach further identified the flame retardant 3,3′,5,5′-tetrabromobisphenol A (TBBPA) as a selective toxicant for dopaminergic neurons in the context of human midbrain-like tissues for the first time. Results were verified with high reproducibility in more detailed dose-response experiments. Further, we demonstrate higher sensitivity in 3D AMOs than in 2D cultures to the known neurotoxic effects of the pesticide lindane. Overall, the automated nature of our workflow is freely scalable and demonstrates the feasibility of quantitatively assessing cell-type-specific toxicity in human organoids in vitro.

The present study aimed to evaluate the production and postharvest quality of yellow passion fruit, cultivated under different levels of irrigation water salinity and exogenous application of hydrogen peroxide. At low concentrations, hydrogen peroxide may favor plant acclimatization when subjected to saline stress conditions due to the activation of defense mechanisms. The experiment was conducted in greenhouse in the municipality of Campina Grande-PB, Brazil, using drainage lysimeters filled with sandy loam Entisol. The experimental design was randomized blocks, with 4×4 factorial arrangement with three replicates, consisting of four levels of irrigation water electrical conductivity—ECw (0.7, 1.4, 2.1 and 2.8 dS m-1) and four concentrations of hydrogen peroxide—H2O2 (0, 20, 40 and 60 μM). The different concentrations of H2O2 were applied by soaking the seeds for a 24-h period before sowing and spraying the leaves on the adaxial and abaxial sides. At the end of the cycle (205 days after transplanting-DAT), passion fruit production was evaluated by determining the number of fruits per plant, mean fruit weight and total fruit weight. Postharvest quality was determined by physical characterization of fruit (equatorial and polar diameters and rind and pulp thicknesses), hydrogen potential in the pulp, titratable acidity and ascorbic acid. Increasing levels of irrigation water salinity negatively affected the production and physical and chemical quality of passion fruits. Number and total weight of fruits per plant were the most compromised variables. There was significant interaction between irrigation water salinity levels and H2O2 concentrations on fruits’ polar diameter, at 205 days after transplanting. Foliar application at estimated concentration of 27.5 and 41.5 μM H2O2 led to the highest values of total titratable acidity and vitamin C, respectively, at 205 DAT. Water salinity above 0.7 dS m-1 resulted in changes in the physico-chemical characteristics of passion fruit. Peroxide concentrations up to 60 μM did not mitigate the deleterious effects of salt stress on passion fruit yield and quality in the first crop cycle.