Mastigoneme structure reveals insights into the O-linked glycosylation code of native hydroxyproline-rich helices.

Hydroxyproline-rich glycoproteins (HRGPs) are a ubiquitous class of protein in the extracellular matrices and cell walls of plants and algae, yet little is known of their native structures or interactions. Here, we used electron cryomicroscopy (cryo-EM) to determine the structure of the hydroxyproline-rich mastigoneme, an extracellular filament isolated from the cilia of the alga Chlamydomonas reinhardtii. The structure demonstrates that mastigonemes are formed from two HRGPs (a filament of MST1 wrapped around a single copy of MST3) that both have hyperglycosylated poly(hydroxyproline) helices. Within the helices, O-linked glycosylation of the hydroxyproline residues and O-galactosylation of interspersed serine residues create a carbohydrate casing. Analysis of the associated glycans reveals how the pattern of hydroxyproline repetition determines the type and extent of glycosylation. MST3 possesses a PKD2-like transmembrane domain that forms a heteromeric polycystin-like cation channel with PKD2 and SIP, explaining how mastigonemes are tethered to ciliary membranes.

Mechanism of IFT-A polymerization into trains for ciliary transport.

Intraflagellar transport (IFT) is the highly conserved process by which proteins are transported along ciliary microtubules by a train-like polymeric assembly of IFT-A and IFT-B complexes. IFT-A is sandwiched between IFT-B and the ciliary membrane, consistent with its putative role in transporting transmembrane and membrane-associated cargoes. Here, we have used single-particle analysis electron cryomicroscopy (cryo-EM) to determine structures of native IFT-A complexes. We show that subcomplex rearrangements enable IFT-A to polymerize laterally on anterograde IFT trains, revealing a cooperative assembly mechanism. Surprisingly, we discover that binding of IFT-A to IFT-B shields the preferred lipid-binding interface from the ciliary membrane but orients an interconnected network of ß-propeller domains with the capacity to accommodate diverse cargoes toward the ciliary membrane. This work provides a mechanistic basis for understanding IFT-train assembly and cargo interactions.

Human parainfluenza virus 3 vaccine candidates attenuated by codon-pair deoptimization are immunogenic and protective in hamsters.

Human parainfluenza virus type 3 (HPIV3) is a major pediatric respiratory pathogen lacking available vaccines or antiviral drugs. We generated live-attenuated HPIV3 vaccine candidates by codon-pair deoptimization (CPD). HPIV3 open reading frames (ORFs) encoding the nucleoprotein (N), phosphoprotein (P), matrix (M), fusion (F), hemagglutinin-neuraminidase (HN), and polymerase (L) were modified singly or in combination to generate 12 viruses designated Min-N, Min-P, Min-M, Min-FHN, Min-L, Min-NP, Min-NPM, Min-NPL, Min-PM, Min-PFHN, Min-MFHN, and Min-PMFHN. CPD of N or L severely reduced growth in vitro and was not further evaluated. CPD of P or M was associated with increased and decreased interferon (IFN) response in vitro, respectively, but had little effect on virus replication. In Vero cells, CPD of F and HN delayed virus replication, but final titers were comparable to wild-type (wt) HPIV3. In human lung epithelial A549 cells, CPD F and HN induced a stronger IFN response, viral titers were reduced 100-fold, and the expression of F and HN proteins was significantly reduced without affecting N or P or the relative packaging of proteins into virions. Following intranasal infection in hamsters, replication in the nasal turbinates and lungs tended to be the most reduced for viruses bearing CPD F and HN, with maximum reductions of approximately 10-fold. Despite decreased in vivo replication (and lower expression of CPD F and HN in vitro), all viruses induced titers of serum HPIV3-neutralizing antibodies similar to wt and provided complete protection against HPIV3 challenge. In summary, CPD of HPIV3 yielded promising vaccine candidates suitable for further development.

NAD activates olfactory receptor 1386 to regulate type I interferon responses in Plasmodium yoelii YM infection.

Olfactory receptors (Olfr) are G protein-coupled receptors that are normally expressed on olfactory sensory neurons to detect volatile chemicals or odorants. Interestingly, many Olfrs are also expressed in diverse tissues and function in cell-cell recognition, migration, and proliferation as well as immune responses and disease processes. Here, we showed that many Olfr genes were expressed in the mouse spleen, linked to Plasmodium yoelii genetic loci significantly, and/or had genome-wide patterns of LOD scores (GPLSs) similar to those of host Toll-like receptor genes. Expression of specific Olfr genes such as Olfr1386 in HEK293T cells significantly increased luciferase signals driven by IFN-ß and NF-κB promoters, with elevated levels of phosphorylated TBK1, IRF3, P38, and JNK. Mice without Olfr1386 were generated using the CRISPR/Cas9 method, and the Olfr1386-/- mice showed significantly lower IFN-α/ß levels and longer survival than wild-type (WT) littermates after infection with P. yoelii YM parasites. Inhibition of G protein signaling and P38 activity could affect cyclic AMP-responsive element promoter-driven luciferase signals and IFN-ß mRNA levels in HEK293T cells expressing the Olfr1386 gene, respectively. Screening of malaria parasite metabolites identified nicotinamide adenine dinucleotide (NAD) as a potential ligand for Olfr1386, and NAD could stimulate IFN-ß responses and phosphorylation of TBK1 and STAT1/2 in RAW264.7 cells. Additionally, parasite RNA (pRNA) could significantly increase Olfr1386 mRNA levels. This study links multiple Olfrs to host immune response pathways, identifies a candidate ligand for Olfr1386, and demonstrates the important roles of Olfr1386 in regulating type I interferon (IFN-I) responses during malaria parasite infections.

Partial order relation-based gene ontology embedding improves protein function prediction.

Protein annotation has long been a challenging task in computational biology. Gene Ontology (GO) has become one of the most popular frameworks to describe protein functions and their relationships. Prediction of a protein annotation with proper GO terms demands high-quality GO term representation learning, which aims to learn a low-dimensional dense vector representation with accompanying semantic meaning for each functional label, also known as embedding. However, existing GO term embedding methods, which mainly take into account ancestral co-occurrence information, have yet to capture the full topological information in the GO-directed acyclic graph (DAG). In this study, we propose a novel GO term representation learning method, PO2Vec, to utilize the partial order relationships to improve the GO term representations. Extensive evaluations show that PO2Vec achieves better outcomes than existing embedding methods in a variety of downstream biological tasks. Based on PO2Vec, we further developed a new protein function prediction method PO2GO, which demonstrates superior performance measured in multiple metrics and annotation specificity as well as few-shot prediction capability in the benchmarks. These results suggest that the high-quality representation of GO structure is critical for diverse biological tasks including computational protein annotation.

Bioremediation of mercury-polluted soil and water by the plant symbiotic fungus Metarhizium robertsii.

Fungi are central to every terrestrial and many aquatic ecosystems, but the mechanisms underlying fungal tolerance to mercury, a global pollutant, remain unknown. Here, we show that the plant symbiotic fungus Metarhizium robertsii degrades methylmercury and reduces divalent mercury, decreasing mercury accumulation in plants and greatly increasing their growth in contaminated soils. M. robertsii does this by demethylating methylmercury via a methylmercury demethylase (MMD) and using a mercury ion reductase (MIR) to reduce divalent mercury to volatile elemental mercury. M. robertsii can also remove methylmercury and divalent mercury from fresh and sea water even in the absence of added nutrients. Overexpression of MMD and MIR significantly improved the ability of M. robertsii to bioremediate soil and water contaminated with methylmercury and divalent mercury. MIR homologs, and thereby divalent mercury tolerance, are widespread in fungi. In contrast, MMD homologs were patchily distributed among the few plant associates and soil fungi that were also able to demethylate methylmercury. Phylogenetic analysis suggests that fungi could have acquired methylmercury demethylase genes from bacteria via two independent horizontal gene transfer events. Heterologous expression of MMD in fungi that lack MMD homologs enabled them to demethylate methylmercury. Our work reveals the mechanisms underlying mercury tolerance in fungi, and may provide a cheap and environmentally friendly means of cleaning up mercury pollution.

Heterozygous LRP1 deficiency causes developmental dysplasia of the hip by impairing triradiate chondrocytes differentiation due to inhibition of autophagy.

Developmental dysplasia of the hip (DDH) is one of the most common congenital skeletal malformations; however, its etiology remains unclear. Here, we conducted whole-exome sequencing in eight DDH families followed by targeted sequencing of 68 sporadic DDH patients. We identified likely pathogenic variants in the LRP1 (low-density lipoprotein receptor-related protein 1) gene in two families and seven unrelated patients. All patients harboring the LRP1 variants presented a typical DDH phenotype. The heterozygous Lrp1 knockout (KO) mouse (Lrp1+/-) showed phenotypes recapitulating the human DDH phenotypes, indicating Lrp1 loss of function causes DDH. Lrp1 knockin mice with a missense variant corresponding to a human variant identified in DDH (Lrp1R1783W) also presented DDH phenotypes, which were milder in heterozygotes and severer in homozygotes than those of the Lrp1 KO mouse. The timing of triradiate cartilage development was brought forward 1 or 2 wk earlier in the LRP-deficient mice, which leads to malformation of the acetabulum and femoral head. Furthermore, Lrp1 deficiency caused a significant decrease of chondrogenic ability in vitro. During the chondrogenic induction of mice bone marrow stem cells and ATDC5 (an inducible chondrogenic cell line), Lrp1 deficiency caused decreased autophagy levels with significant ß-catenin up-regulation and suppression of chondrocyte marker genes. The expression of chondrocyte markers was rescued by PNU-74654 (a ß-catenin antagonist) in an shRNA-Lrp1-expressed ATDC5 cell. Our study reveals a critical role of LRP1 in the etiology and pathogenesis of DDH, opening an avenue for its treatment.

Utilization of plant-derived sugars and lipids are coupled during colonization of rhizoplane and rhizosphere by the fungus Metarhizium robertsii.

Plant-derived sugars and lipids are key nutritional sources for plant associated fungi. However, the relationship between utilization of host-derived sugars and lipids during development of the symbiotic association remains unknown. Here we show that the fungus Metarhizium robertsii also needs plant-derived lipids to develop symbiotic relationship with plants. The fatty acid binding proteins FABP1 and FABP2 are important for utilization of plant-derived lipids as the deletion of Fabp1 and Fabp2 significantly reduced the ability of M. robertsii to colonize rhizoplane and rhizosphere of maize and Arabidopsis thaliana. Deleting Fabp1 and Fabp2 increased sugar utilization by upregulating six sugar transporters, and this explains why deleting the monosaccharide transporter gene Mst1, which plays an important role in utilization of plant-derived sugars, had no impact on the ability of the double-gene deletion mutant ΔFabp1::ΔFabp2 to colonize plant roots. FABP1 and FABP2 were also found in other plant-associated Metarhizium species, and they were highly expressed in the medium using the tomato root exudate as the sole carbon and nitrogen source, suggesting that they could be also important for these species to develop symbiotic relationship with plants. In conclusion, we discovered that utilization of plant-derived sugars and lipids are coupled during colonization of rhizoplane and rhizosphere by M. robertsii.

Construction and Validation of a Prognostic Model Based on Mitochondrial Genes in Prostate Cancer.

This study attempted to build a prostate cancer (PC) prognostic risk model with mitochondrial feature genes. PC-related MTGs were screened for Cox regression analyses, followed by establishing a prognostic model. Model validity was analyzed via survival analysis and receiver operating characteristic (ROC) curves, and model accuracy was validated in the GEO dataset. Combining risk score with clinical factors, the independence of the risk score was verified by using Cox analysis, followed by generating a nomogram. The Gleason score, microsatellite instability (MSI), immune microenvironment, and tumor mutation burden were analyzed in two risk groups. Finally, the prognostic feature genes were verified through a q-PCR test. Ten PC-associated MTGs were screened, and a prognostic model was built. Survival analysis and ROC curves illustrated that the model was a good predictor for the risk of PC. Cox regression analysis revealed that risk score acted as an independent prognostic factor. The Gleason score and MSI in the high-risk group were substantially higher than in the low-risk group. Levels of ESTIMATE Score, Immune Score, Stromal Score, immune cells, immune function, immune checkpoint, and immunopheno score of partial immune checkpoints in the high-risk group were significantly lower than in the low-risk group. Genes with the highest mutation frequencies in the two groups were SPOP, TTN, and TP53. The q-PCR results of the feature genes were consistent with the gene expression results in the database. The 10-gene model based on MTGs could accurately predict the prognosis of PC patients and their responses to immunotherapy.

[Intestinal absorption components and absorption characteristics of Wuwei Qingzhuo San by everted intestinal sac models].

This study investigated the absorption profile of Wuwei Qingzhuo San in different intestinal segments and the absorption characteristics of its alkaloids(piperine, piperanine, piperlonguminine, and dihydropiperlonguminine). The everted gut sac model was established, and the chemical components of Wuwei Qingzhuo San in different intestinal segments were detected by UPLC-Q-TOF-MS. The content of piperine, piperanine, piperlonguminine, and dihydropiperlonguminine in intestinal absorption fluid was determined by UPLC-Q-TRAP-MS and the absorption parameters were calculated. The absorption characteristics in different intestinal segments at different time were analyzed. As a result, 27, 27, 8, and 6 absorbent components from Wuwei Qingzhuo San were detected in the intestinal cyst fluid of jejunum, ileum, duodenum, and colon by UPLC-Q-TOF-MS technology, respectively. It was also found that piperine, piperanine, piperlonguminine, and dihydropiperlonguminine from Wuwei Qingzhuo San showed linear absorption in various intestinal segments, with r values exceeding 0.9. In terms of absorption content, the components were ranked as piperine>piperanine>dihydropiperlonguminine>piperlonguminine in various intestinal segments, but the absorption rate and mechanism of each component varied. The results demonstrate that the absorption of the components of Wuwei Qingzhuo San in different intestinal segments is selective and is not a simple semi-permeable membrane permeation process.

Recombinant rotavirus expressing the glycosylated S1 protein of SARS-CoV-2.

Reverse genetic systems have been used to introduce heterologous sequences into the rotavirus segmented double-stranded (ds)RNA genome, enabling the generation of recombinant viruses that express foreign proteins and possibly serve as vaccine vectors. Notably, insertion of SARS-CoV-2 sequences into the segment seven (NSP3) RNA of simian SA11 rotavirus was previously shown to result in the production of recombinant viruses that efficiently expressed the N-terminal domain (NTD) and the receptor-binding domain (RBD) of the S1 region of the SARS-CoV-2 spike protein. However, efforts to generate a similar recombinant (r) SA11 virus that efficiently expressed full-length S1 were less successful. In this study, we describe modifications to the S1-coding cassette inserted in the segment seven RNA that allowed recovery of second-generation rSA11 viruses that efficiently expressed the ~120-kDa S1 protein. The ~120-kDa S1 products were shown to be glycosylated, based on treatment with endoglycosidase H, which reduced the protein to a size of ~80 kDa. Co-pulldown assays demonstrated that the ~120-kDa S1 proteins had affinity for the human ACE2 receptor. Although all the second-generation rSA11 viruses expressed glycosylated S1 with affinity for the ACE receptor, only the S1 product of one virus (rSA11/S1f) was appropriately recognized by anti-S1 antibodies, suggesting the rSA11/S1f virus expressed an authentic form of S1. Compared to the other second-generation rSA11 viruses, the design of the rSA11/S1f was unique, encoding an S1 product that did not include an N-terminal FLAG tag. Probably due to the impact of FLAG tags upstream of the S1 signal peptides, the S1 products of the other viruses (rSA11/3fS1 and rSA11/3fS1-His) may have undergone defective glycosylation, impeding antibody binding. In summary, these results indicate that recombinant rotaviruses can serve as expression vectors of foreign glycosylated proteins, raising the possibility of generating rotavirus-based vaccines that can induce protective immune responses against enteric and mucosal viruses with glycosylated capsid components, including SARS-CoV-2.

Type III and Not Type I Interferons Efficiently Prevent the Spread of Rotavirus in Human Intestinal Epithelial Cells.

Rotavirus infects intestinal epithelial cells and is the leading cause of gastroenteritis in infants worldwide. Upon viral infection, intestinal cells produce type I and type III interferons (IFNs) to alert the tissue and promote an antiviral state. These two types of IFN bind to different receptors but induce similar pathways that stimulate the activation of interferon-stimulated genes (ISGs) to combat viral infection. In this work, we studied the spread of a fluorescent wild-type (WT) SA11 rotavirus in human colorectal cancer cells lacking specific interferon receptors and compared it to that of an NSP1 mutant rotavirus that cannot interfere with the host intrinsic innate immune response. We could show that the WT rotavirus efficiently blocks the production of type I IFNs but that type III IFNs are still produced, whereas the NSP1 mutant rotavirus allows the production of both. Interestingly, while both exogenously added type I and type III IFNs could efficiently protect cells against rotavirus infection, endogenous type III IFNs were found to be key to limit infection of human intestinal cells by rotavirus. By using a fluorescent reporter cell line to highlight the cells mounting an antiviral program, we could show that paracrine signaling driven by type III IFNs efficiently controls the spread of both WT and NSP1 mutant rotavirus. Our results strongly suggest that NSP1 efficiently blocks the type I IFN-mediated antiviral response; however, its restriction of the type III IFN-mediated one is not sufficient to prevent type III IFNs from partially inhibiting viral spread in intestinal epithelial cells. Additionally, our findings further highlight the importance of type III IFNs in controlling rotavirus infection, which could be exploited as antiviral therapeutic measures. IMPORTANCE Rotavirus is one of the most common causes of gastroenteritis worldwide. In developing countries, rotavirus infections lead to more than 200,000 deaths in infants and children. The intestinal epithelial cells lining the gastrointestinal tract combat rotavirus infection by two key antiviral compounds known as type I and III interferons. However, rotavirus has developed countermeasures to block the antiviral actions of the interferons. In this work, we evaluated the arms race between rotavirus and type I and III interferons. We determined that although rotavirus could block the induction of type I interferons, it was unable to block type III interferons. The ability of infected cells to produce and release type III interferons leads to the protection of the noninfected neighboring cells and the clearance of rotavirus infection from the epithelium. This suggests that type III interferons are key antiviral agents and could be used to help control rotavirus infections in children.

Testing for adaptive changes linked to range expansion following a single introduction of the fall webworm.

Adaptive evolution following colonization can affect the impact of invasive species. The fall webworm (FWW) invaded China 40 years ago through a single introduction event involving a severe bottleneck and subsequently diverged into two genetic groups. The well-recorded invasion history of FWW, coupled with a clear pattern of genetic divergence, provides an opportunity to investigate whether there is any sign of adaptive evolution following the invasion. Based on genome-wide SNPs, we identified genetically separated western and eastern groups of FWW and correlated spatial variation in SNPs with geographical and climatic factors. Geographical factors explained a similar proportion of the genetic variation across all populations compared with climatic factors. However, when the two population groups were analysed separately, environmental factors explained more variation than geographical factors. SNP outliers in populations of the western group had relatively stronger response to precipitation than temperature-related variables. Functional annotation of SNP outliers identified genes associated with insect cuticle protein potentially related to desiccation adaptation in the western group and genes associated with lipase biosynthesis potentially related to temperature adaptation in the eastern group. Our study suggests that invasive species may maintain the evolutionary potential to adapt to heterogeneous environments despite a single invasion event. The molecular data suggest that quantitative trait comparisons across environments would be worthwhile.

Methods for extending working distance using modified photonic crystal for near-field lithography.

Near-field lithography has evident advantages in fabricating super-resolution nano-patterns. However, the working distance (WD) is limited due to the exponential decay characteristic of the evanescent waves. Here, we proposed a novel photolithography method based on a modified photonic crystal (PC), where a defect layer is embedded into the all-dielectric multilayer structure. It is shown that this design can amend the photonic band gap and enhance the desired high-kwaves dramatically, then the WD in air conditions could be extended greatly, which would drastically relax the engineering challenges for introducing the near-field lithography into real-world manufacturing applications. Typically, deep subwavelength patterns with a half-pitch of 32 nm (i.e.,λ/6) could be formed in photoresist layer at an air WD of 100 nm. Moreover, it is revealed that diversified two-dimensional patterns could be produced with a single exposure using linear polarized light. The analyses indicate that this improved dielectric PC is applicable for near-field lithography to produce super-resolution periodic patterns with large WD, strong field intensity, and great uniformity.

From Feynman's ratchet to time crystalline molecular motors.

Cats have an instinctive ability to use the connection governing parallel transport in the space of shapes to land safely on their feet. Here, we argue that the concept of connection, which is extensively used in general relativity and other parts of theoretical physics, also explains the impressive performance of molecular motors by enabling molecules to evade the conclusions of Feynman's ratchet-and-pawl analysis. First, we demonstrate the emergence of directed rotational motion from shape changes, which is independent of angular momentum. Then, we computationally design knotted polyalanine molecules and demonstrate the organization of individual atom thermal vibrations into collective rotational motion, which is independent of angular momentum. The motion occurs effortlessly even in ambient water and can be further enhanced through spontaneous symmetry breaking, rendering the molecule an effective theory time crystal. Our findings can be experimentally verified via nuclear magnetic resonance measurements and hold practical potential for molecular motor design and engineering.

Exploring the therapeutic mechanisms of Sijunzi decoction in the treatment of sarcopenia: Key targets and signaling pathways.

Sarcopenia, an age-associated condition, negatively impacts the quality of life. This study investigates the mechanism of Sijunzi decoction (SJZD), a traditional Chinese formula, against sarcopenia. Active compounds and potential targets of SJZD for sarcopenia were gathered from databases. Hub targets were identified using protein-protein interaction networks, with GO and KEGG analyses suggesting potential pathways. Molecular docking was used to assess compound-target affinity. A lipopolysaccharide-induced sarcopenia rat model was used to verify the targets. Sijunzi decoction contains 92 compounds and 47 targets for sarcopenia. The top 10 hub targets comprise AKT1, ALB, INS, IL6, TNF, TP53, VEGFA, SIRT1, CAT and FOS. GO and KEGG analyses indicate involvement in steroid hormone response, vesicle lumen, receptor agonist activity, and FoxO and HIF-1 signaling pathways. Validation experiments showed that SJZD alleviates sarcopenia by downregulating SIRT1, IL-6, TNF and AKT1. Sijunzi decoction treats sarcopenia by targeting SIRT1, IL-6, TNF and AKT1, potentially involving FoxO and HIF-1 signaling pathways. This highlights SJZD's potential for sarcopenia treatment.

Exercise-Induced Tachycardia-Dependent Atrioventricular Block with Narrow QRS Complexes: A Case Report and Review of Literature.

Exercise-induced tachycardia-dependent atrioventricular block with a normal electrocardiogram at rest is rare. Herein, we present a case of a 65-year-old woman with exercise-related chest suppression and treadmill exercise test-induced second- and third-degree atrioventricular blocks with narrow QRS wave and normal resting electrocardiogram. High atrioventricular block leads to tissue and organ insufficiency, resulting in exercise intolerance, dyspnea, dizziness, and syncope. Ths diagnosis was exercise-induced high-degree atrioventricular block. For lack of effective medicines, the patient received a permanent dual chamber pacemaker to ensure atrioventricular sequential pacing during exercise. No exercise-related discomfort occurred during follow up.

Characteristics and Potential Challenges of Digital-Based Interventions for Children and Young People: Scoping Review.

BACKGROUND: Digital health technologies are becoming increasingly available to children and young people and their families. However, there are no scoping reviews that provide both an overview of the characteristics of digital interventions for children and young people and potential challenges to be considered when developing and implementing them. OBJECTIVE: This study aimed to systematically review scientific publications to identify the current characteristics and potential complications of digital interventions for children and young people. METHODS: This scoping review was conducted using the framework of Arksey and O'Malley and adheres to the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines for scoping reviews. A search of 5 databases (PubMed, Scopus, Embase, MEDLINE, and CINAHL) and Google Scholar was performed for eligible clinical trials published between January 1, 2018, and August 19, 2022. RESULTS: The initial search of the 5 databases yielded 3775 citations; duplicates and those not meeting the inclusion criteria were eliminated. In total, 34 articles were included in the final review and relevant information, such as the descriptive characteristics and potential challenges, were classified. Mental health (26/34, 76%) was the most common target for the digital intervention for children and young people, exceeding physical health (8/34, 24%) by more than 3 times. In addition, a substantial number of digital interventions were dedicated solely to children and young people. Digital interventions for children and young people were more likely to be delivered via computers (17/34, 50%) rather than smartphones (13/34, 38%). More than one-third of the studies (13/34, 38%) applied cognitive behavioral theory as the theory of digital interventions. The duration of the digital intervention for children and young people was more likely to vary depending on the target user rather than the target disease. Intervention components were classified into 5 categories: guidance, task and activity, reminder and monitoring, supportive feedback, and reward system. Potential challenges were subcategorized into ethical, interpersonal, and societal challenges. For ethical challenges, the consent of children and young people or caregivers, potential risk of adverse events, and data privacy issues were considered. For interpersonal challenges, the engagement of children and young people was affected by the preference or barrier of caregivers to participate in studies. For societal challenges, restricted ethnicity in recruitment, limited availability of digital technology, differences in internet use patterns between girls and boys, unified clinical settings, and language barriers were described. CONCLUSIONS: We identified potential challenges and provided suggestions about ethical, interpersonal, and societal aspects to consider when developing and deploying digital-based interventions for children and young people. Our findings provide a thorough overview of the published literature and may serve as a comprehensive, informative foundation for the development and implementation of digital-based interventions for children and young people.

An inactivating mutation in the vacuolar arginine exporter gene Vae results in culture degeneration in the fungus Metarhizium robertsii.

Culture degeneration usually results in great commercial losses in the economically important filamentous fungi, but the genetic causes of the degeneration remain elusive. In the fungus Metarhizium robertsii, we found that deletion of the vacuolar arginine exporter gene Vae caused culture degeneration. Compared to the WT strain, the mutant showed increased apoptosis, reactive oxygen species (ROS) level and mitochondrial membrane potential collapse, reduced conidial yield and abnormal lipid droplet formation. The extent of the degeneration in the mutant gradually increased over the successive subculturing, which eventually became irreversible; compared to the third subculture of the mutant, the seventh subculture showed a lower conidial yield and pathogenicity to insects, stronger apoptosis, higher ROS level and a smaller number of conidial lipid droplets. Incorporation of the genomic clone of Vae could not restore the WT phenotypes in the seventh subculture, but could in the third one. Loss-of-function in Vae resulted in vacuolar arginine accumulation and reduction in the cytosolic arginine. This downregulated the expression of the regulator CAG9 of G protein signalling pathway, which accounted for most of the phenotypic changes associated with the degeneration of the mutant. We identified a deleterious mutation that causes culture degeneration in a filamentous fugus.

Out of chaos: Phylogenomics of Asian Sonerileae.

Sonerileae is a diverse Melastomataceae lineage comprising ca. 1000 species in 44 genera, with >70% of genera and species distributed in Asia. Asian Sonerileae are taxonomically intractable with obscure generic circumscriptions. The backbone phylogeny of this group remains poorly resolved, possibly due to complexity caused by rapid species radiation in early and middle Miocene, which hampers further systematic study. Here, we used genome resequencing data to reconstruct the phylogeny of Asian Sonerileae. Three parallel datasets, viz. single-copy ortholog (SCO), genomic SNPs, and whole plastome, were assembled from genome resequencing data of 205 species for this purpose. Based on these genome-scale data, we provided the first well resolved phylogeny of Asian Sonerileae, with 34 major clades identified and 74% of the interclade relationships consistently resolved by both SCO and genomic data. Meanwhile, widespread phylogenetic discordance was detected among SCO gene trees as well as species trees reconstructed using different tree estimation methods (concatenation/site-based coalescent method/summary method) or different datasets (SCO/genomic/plastome). We explored sources of discordance using multiple approaches and found that the observed discordance in Asian Sonerileae was mainly caused by a combination of biased distribution of missing data, random noise from uninformative genes, incomplete lineage sorting, and hybridization/introgression. Exploration of these sources can enable us to generate hypotheses for future testing, which is the first step towards understanding the evolution of Asian Sonerileae. We also detected high levels of homoplasy for some characters traditionally used in taxonomy, which explains current chaotic generic delimitations. The backbone phylogeny of Asian Sonerileae revealed in this study offers a solid basis for future taxonomic revision at the generic level.