bluuebunny/crossref_metadata_2025 huggingface dataset:
Reproductive resource allocation correlates with successful global invasion of a mosquito species
Authors: Khorramnejad Ayda , Alfaro Claudia , Quaranta Stefano , Lozada-Chávez Alejandro Nabor , Gasmi Laila , Perdomo Hugo D. , Chiarelli Laurent Roberto , Bonizzoni Mariangela
Published: July 2024

The understanding of traits favoring biological invasions has been considered an essential step to predict which species would become successful invaders. Classical approaches test for differences between invasive vs. not invasive species and emphasize reproduction as a critical phenotype for successful establishment of an invasive species. However, cross-species comparisons underestimate intraspecies differences, which may be relevant in invasive species with highly genetically diverse populations. Here we capitalize on the well-characterized invasion history of the arboviral vectorAedes albopictus, which resulted in genetically-distinct native, old and invasive populations, and compared the reproductive capacity (fertility and fecundity), development (timing of egg development, oviposition patterns and egg hatching) and physiology (blood digestion and nutrient movement during oogenesis) across populations. We observed that invasive mosquitoes optimize their nutrient investment during development and oogenesis, which leads to increased egg production with respect to native and long adapted laboratory mosquitoes. This higher fecundity results from a delay in oogenesis and is accompanied by higher fertility. We further tested inheritance of reproductive traits via reciprocal crosses, which showed a higher fertility and fecundity in hybrids with respect to parental strains and a potential contribution of males to the reproductive success of invasive mosquitoes. Our results provide evidence that resource allocation during development and oogenesis influences the reproductive capacity ofAe. albopictusand manifests in population differences that correlate with their invasion success.

Significance Statement

In addition to being an essential process to ensure the survival of a species, reproduction is a key determinant for a species invasion success because it facilitates a species’ ability to establish in a new area. Reproduction is a complex phenotype that relies on intricate and timely interactions between genetic and physiological factors. Here we combined molecular, biochemical, and genetic approaches to show that efficient allocation of energetic resources during development and oogenesis fosters the reproductive output ofAe. albopictusmosquitoes and manifests as variation in the reproductive capacity of its geographic populations. These results are critical for predicting the invasion success of this species and tailoring effective control strategies.
Identification and characterization of thiamine analogues with antiplasmodial activity
Authors: Fathoni Imam , Ho Terence C. S. , Chan Alex H. Y. , Leeper Finian J. , Matuschewski Kai , Saliba Kevin J.
Published: July 2024

Thiamine is metabolized into thiamine pyrophosphate (TPP), an essential enzyme cofactor. Previous work has shown that oxythiamine, a thiamine analogue, is metabolized by thiamine pyrophosphokinase (TPK) into oxythiamine pyrophosphate (OxPP) within the malaria parasitePlasmodium falciparum, and then inhibits TPP-dependent enzymes, killing the parasitein vitroandin vivo. To identify a more potent antiplasmodial thiamine analogue, 11 commercially available compounds were tested againstP. falciparumandP. knowlesi. Five active compounds were identified, but only N3-pyridyl thiamine (N3PT), a potent transketolase inhibitor and candidate anticancer lead compound, was found to suppressP. falciparumproliferation with an IC50value 10-fold lower than that of oxythiamine. N3PT was active againstP. knowlesiand was >17 times less toxic to human fibroblast, as compared to oxythiamine. Increasing the extracellular thiamine concentration reduced the antiplasmodial activity of N3PT, consistent with N3PT competing with thiamine/TPP. A transgenicP. falciparumline overexpressing TPK was found to be hypersensitized to N3PT. Docking studies showed an almost identical binding mode in TPK between thiamine and N3PT. Furthermore, we show that [3H]thiamine accumulation, resulting from a combination of transport and metabolism, in isolated parasites is reduced by N3PT. Treatment ofP. berghei-infected mice with 200 mg/kg/day N3PT reduced their parasitemia, prolonged their time to malaria symptoms, and appeared to be non-toxic to mice. Collectively, our studies are consistent with N3PT competing with thiamine for TPK binding and inhibiting parasite proliferation by reducing TPP production, as well as being converted into a TPP antimetabolite that inhibits TPP-dependent enzymes.
Tubules, rods and spirals: diverse modes of SepF-FtsZ assembling
Authors: Choudhury Jagrity , Chaudhuri Barnali N.
Published: July 2024

Z-ring formation by FtsZ, the master assembler of divisome, is a key step in bacterial cell division. Both formation and membrane anchoring of the Z-ring requires assistance of a number of Z-ring binding proteins, such as FtsA, EzrA, SepF, SepH and ZipA. SepF participates in bundling and membrane anchoring of FtsZ in gram-positive bacteria. We reportin vitrobiophysical studies of the interactions between FtsZ and cytoplasmic component of cognate SepF from three different bacteria:Mycobacterium tuberculosis,Staphylococcus aureusandEnterococcus gallinarum.While the cytosolic domain of SepF fromM. tuberculosisis a dimer, those fromS. aureusandE. gallinarumpolymerize to form ring-like structures. Mycobacterial SepF helps in bundling of FtsZ filaments to form thick filaments and large spirals. On the other hand, ring-forming SepF from theFirmicutesbundle FtsZ into tubules.
Statistical Survey of Chemical and Geometric Patterns on Protein Surfaces as a Blueprint for Protein-mimicking Nanoparticles
Authors: McBride John M. , Koshevarnikov Aleksei , Siek Marta , Grzybowski Bartosz A. , Tlusty Tsvi
Published: July 2024

Despite recent breakthroughs in understanding how protein sequence relates to structure and function, considerably less attention has been paid to the general features of protein surfaces beyond those regions involved in binding and catalysis. This paper provides a systematic survey of the universe of protein surfaces and quantifies the sizes, shapes, and curvatures of the positively/negatively charged and hydrophobic/hydrophilic surface patches as well as correlations between such patches. It then compares these statistics with the metrics characterizing nanoparticles functionalized with ligands terminated with positively and negatively charged ligands. These particles are of particular interest because they are also surface-patchy and have been shown to exhibit both antibiotic and anticancer activities – via selective interactions against various cellular structures – prompting loose analogies to proteins. Our analyses support such analogies in several respects (e.g., patterns of charged protrusions and hydrophobic niches similar to those observed in proteins), although there are also significant differences. Looking forward, this work provides a blueprint for the rational design of synthetic nanoobjects with further enhanced mimicry of proteins’ surface properties.
Evaluating the Efficacy of AI-Based Interactive Assessments Using Large Language Models for Depression Screening
Authors: Jin Zheng , Bi Dandan , Hu Jiaxing , Zhao Kaibin
Published: July 2024

The evolution of language models, particularly the development of Large Language Models like ChatGPT, has opened new avenues for psychological assessment, potentially revolutionizing the rating scale methods that have been used for over a century. This study introduces a new Automated Assessment Paradigm (AAP), which aims to integrate natural language processing (NLP) techniques with traditional measurement methods. This integration enhances the accuracy and depth of mental health evaluations, while also addressing the acceptance and subjective experience of participants—areas that have not been extensively measured before. A pilot study was conducted with 32 participants, seven of whom were diagnosed with depression by licensed psychiatrists using the Clinical Interview Schedule-Revised (CIS-R). The participants completed the BDI-Fast Screen (BDI-FS) using a custom ChatGPT (GPTs) interface and the Chinese version of the PHQ-9 in a private setting. Following these assessments, participants also completed the Subjective Evaluation Scale. Spearman’s correlation analysis showed a high correlation between the total scores of the PHQ-9 and the BSI-FS-GPTs. The agreement of diagnoses between the two measures, as measured by Cohen’s kappa, was also significant. BSI-FS-GPTs diagnosis showed significantly higher agreement with the current diagnosis of depression. However, given the limited sample size of the pilot study, the AUC value of 1.00 and a sensitivity of 0.80 at a cutoff of 0.5, with zero false positive rate, likely overstate the classifier’s performance. Bayesian factors suggest that participants may feel more comfortable expressing their true feelings and opinions through this method. For ongoing follow-up research, a total sample size of approximately 104 participants, including about 26 diagnosed individuals, may be required to ensure the analysis maintains a necessary power of 0.80 and an alpha level of 0.05. Nonetheless, these findings provide a promising foundation for the ongoing validation of the new AAP in larger-scale studies, aiming to confirm its validity and reliability.
Health-Related Quality of Life Perception Among Older Persons with Non-Communicable Diseases in Primary Healthcare Facilities: A Qualitative Inquiry
Authors: Fiona Atim , Regina Ndagire , Nampima Chloe , Kiyinji Frank , Lwanira Catherine , Nanyonga Rose Clarke , Orach-Meza Faustino
Published: July 2024

Background

Unveiling the understanding of older persons with non-communicable diseases (NCDs) regarding health well-being is paramount and can translate to increased self-efficiency, independence, and enhanced well-being. However, little is known about older persons’ understanding of the concept of health-related quality of life (HRQoL) in Uganda. The study explored perceptions of older persons with NCDs on HRQoL in central Uganda.

Methods

This exploratory qualitative study design involved 23 participants recruited from selected Primary healthcare facilities in Central Uganda. Thematic analysis using an inductive approach generated themes that informed the study’s qualitative findings.

Results

The key themes that emerged from the study include holistic well-being, lifestyle modification, and financial stability. The key component of HRQoL that came out clearly from the study was the physical domain. There is a need to embrace a person-centered approach based on the perceptions of older persons on HRQoL, which has the potential to improve well-being and enhance a healthy aging journey.
Association of natural induced antibodies against select Group B streptococcus surface proteins and invasive disease in early infancy
Authors: Dzanibe Sonwabile , Izu Alane , Dangor Ziyaad , Kwatra Gaurav , Adrian Peter V. , Kimaro Mlacha Sheila Z. , Madhi Shabir A.
Published: July 2024

Background

Group Bstreptococcus(GBS) surface protein epitopes are potential targets for development of vaccines that could confer protection against invasive GBS disease (IGbsD) irrespective of the capsular serotype. The aim of this study was to determine the association of natural acquired mother-newborn GBS surface proteins specific serum IgG and IGbsD during early infancy (<90 days age).

Methods

Clinical GBS isolates from 81 women who delivered either infant with IGbsD (n=38) and healthy controls (n=43) were assessed for surface expression of proteins Sip, GBS0393 and GBS206 using flow cytometry. Serum IgG titres to Sip, GBS0393 and GBS206 surface proteins were measured in paired maternal-infant sera on multiplex Luminex platform to determine IgG titres associated with reduce risk of IGbsD caused by GBS strains expressing homotypic protein.

Results

Infant sera IgG GMT in IGbsD cases caused by strains expressing Sip and GBS2016 were lower (64 and 61 U/ml, respectively) compared to healthy controls born to women colonized by strains expressing the respective proteins (145 and 151 U/mL, respectively, p<0.01). Moreover, increasing infant antibody titres against Sip and GBS2016 were associated with ≥80% adjusted disease risk (ADR) reduction to GBS isolates expressing homotypic proteins. Among women colonized with GBS isolates expressing GBS2106, mothers of cases had lower GBS2016-specific IgG GMT (249 U/mL) compared to mothers of controls (163 U/mL, adj-p=0.023).

Conclusion

Increasing infant IgG titres to GBS2106 and Sip was associated with reduced IGbsD risk and therefore warranting further investigations as potential GBS vaccines and/or protein conjugants.
Impact analysis of infant antibiotic exposure on the burden of asthma: a simulation modeling study
Authors: Lee Tae Yoon , Petkau John , Saatchi Ariana , Marra Fawziah , Turvey Stuart E , Lishman Hannah , Patrick David M , Cragg Jacquelyn J , Johnson Kate M , Sadatsafavi Mohsen
Published: July 2024

Background

Infant antibiotic use is associated with increased risk of asthma. We examined the population impact of antibiotic exposure in the first year of life on the burden of pediatric asthma in British Columbia, Canada, using simulation modeling.

Methods

We performed a Bayesian meta-analysis of empirical studies to construct dose-response equations between antibiotic exposure in the first year of life and pediatric (<19 years of age) asthma. We used administrative health data to document trends in infant (< 1 year of age) antibiotic use in British Columbia during 2001 and 2018 (the study period). An independently developed microsimulation model of asthma was utilized to estimate asthma-related outcomes under three scenarios pertaining to the trends in antibiotic use during the study period: 1) observed trends, 2) flat trend in which the prescription rate remained at the 2001 value, and 3) intermediate trends midway between these two. We reported cumulative person-years with asthma, cumulative asthma incidence, and cumulative asthma exacerbations among the pediatric population during the study period.

Results

There were 773,160 live births during the study period, with an average antibiotic prescription rate of 523 per 1,000 infants in the first year of life. The prescription rate decreased by 71.5% during the study period. In Scenario 1, there were 1,982,861 person-years with asthma, 183,392 asthma incident cases, and 383,072 exacerbations. Had the antibiotic exposure remained at the 2001 values (Scenario 2), there would have been additional 37,213 person-years with asthma, 10,053 asthma incident cases, and 23,280 exacerbations. Had the decline been half of the observed trend (Scenario 3), there would have been additional 20,318 person-years with asthma, 5,486 asthma incident cases, and 12,728 exacerbations. At least 80% of the excess burden in each outcome was attributable to the younger pediatric population of <10 years of age.

Conclusions

The decline in infant antibiotic exposure has resulted in a substantial reduction in the burden of asthma in British Columbia. Such benefits should be considered when evaluating the value proposition of initiatives aimed at reducing unnecessary antibiotic exposure in early life.
Exposome-wide association study of cognition among older adults in the National Health and Nutrition Examination Survey
Authors: Middleton Lauren Y. M. , Walker Erika , Cockell Scarlet , Dou John , Nguyen Vy K. , Schrank Mitchell , Patel Chirag J. , Ware Erin B. , Colacino Justin A. , Park Sung Kyun , Bakulski Kelly M.
Published: July 2024

Cognitive impairment among older adults is a growing public health challenge and environmental chemicals may be modifiable risk factors. A wide array of chemicals has not yet been tested for association with cognition in an environment-wide association framework. In the US National Health and Nutrition Examination Survey (NHANES) 1999-2000 and 2011-2014 cross-sectional cycles, cognition was assessed using the Digit Symbol Substitution Test (DSST, scores 0-117) among participants aged 60 years and older. Concentrations of environmental chemicals measured in blood or urine were log2transformed and standardized. Chemicals with at least 50% of measures above the lower limit of detection were included (nchemicals=147, nclasses=14). We tested for associations between chemical concentrations and cognition using parallel survey-weighted multivariable linear regression models adjusted for age, sex, race/ethnicity, education, smoking status, fish consumption, cycle year, urinary creatinine, and cotinine. Participants with at least one chemical measurement (n=4,982) were mean age 69.8 years, 55.0% female, 78.2% non-Hispanic White, and 77.0% at least high school educated. The mean DSST score was 50.4 (standard deviation (SD)=17.4). In adjusted analyses, 5 of 147 exposures were associated with DSST at p-value<0.01. Notably, a SD increase in log2-scaled cotinine concentration was associated with 2.71 points lower DSST score (95% CI -3.69, -1.73). A SD increase in log2-scaled urinary tungsten concentration was associated with 1.34 points lower DSST score (95% CI -2.11, -0.56). Exposure to environmental chemicals, particularly heavy metals and tobacco smoke, may be modifiable factors for cognition among older adults.
Multicellular immune ecotypes within solid tumors predict real-world therapeutic benefits with immune checkpoint inhibitors
Authors: Wang Xuefeng , Li Tingyi , Eljilany Islam , Sukrithan Vineeth , Ratan Aakrosh , McCarter Martin , Carpten John , Colman Howard , Ikeguchi Alexandra P. , Puzanov Igor , Arnold Susanne , Churchman Michelle , Hwu Patrick , Rodriguez Paulo C. , Dalton William S. , Weiner George J. , Tarhini Ahmad A.
Published: July 2024

Background

Cancer initiation, progression, and immune evasion depend on the tumor microenvironment (TME). Thus, understanding the TME immune architecture is essential for understanding tumor metastasis and therapy response. This study aimed to create an immune cell states (CSs) atlas using bulk RNA-seq data enriched by eco-type analyses to resolve the complex immune architectures in the TME.

Methods

We employed EcoTyper, a machine-learning (ML) framework, to study the real-world prognostic significance of immune CSs and multicellular ecosystems, utilizing molecular data from 1,610 patients with multiple malignancies who underwent immune checkpoint inhibitor (ICI) therapy within the ORIEN Avatar cohort, a well-annotated real-world dataset.

Results

Our analysis revealed consistent ICI-specific prognostic TME carcinoma ecotypes (CEs) (including CE1, CE9, CE10) across our pan-cancer dataset, where CE1 being more lymphocyte-deficient and CE10 being more proinflammatory. Also, the analysis of specific immune CSs across different cancers showed consistent CD8+ and CD4+ T cell CS distribution patterns. Furthermore, survival analysis of the ORIEN ICI cohort demonstrated that ecotype CE9 is associated with the most favorable survival outcomes, while CE2 is linked to the least favorable outcomes. Notably, the melanoma-specific prognostic EcoTyper model confirmed that lower predicted risk scores are associated with improved survival and better response to immunotherapy. Finally, de novo discovery of ecotypes in the ORIEN ICI dataset identified Ecotype E3 as significantly associated with poorer survival outcomes.

Conclusion

Our findings offer important insights into refining the patient selection process for immunotherapy in real-world practice and guiding the creation of novel therapeutic strategies to target specific ecotypes within the TME.

Trial Registration:NCT04526730
Autoinhibition of cMyBP-C by its middle domains
Authors: Greenman A.C. , Sadler R.L. , Harris S.P.
Published: July 2024

Cardiac myosin binding protein-C (cMyBP-C) is a sarcomere regulatory protein consisting of 11 well-folded immunoglobulin-like (Ig-like) and fibronectin type-III domains with the individual domains numbered C0-C10. Despite progress in understanding the functions of the N’ and C’-terminal ends of the protein, our understanding of the functional effects of the middle domains (C3-C4-C5-C6-C7) is still limited. Here we aimed to determine the functional significance of the middle domains by replacing endogenous cMyBP-C with recombinant proteins with and without the middle domains using our “cut and paste” SpyC3 mouse model. Specifically, we deleted domains C3-C7 or substituted these domains with unrelated Ig-like domains from titin to behave as inert “spacer” domains. Replacement with the spacer constructs resulted in a significant increase in myofilament calcium sensitivity, an almost instantaneous redevelopment of tension after a slack re-stretch protocol, and altered stretch activation responses, suggesting that the middle domains are functionally relevant and normally exert inhibitory effects on force development. We also investigated the significance of a flexible linker between domains C4 and C5 and a unique 28 amino acid loop insertion in C5. Whereas deletion of the C5 loop had no effect on force, deletion of the linker between C4 and C5 had comparable effects to deletion of domains C3-C7. Taken together, these data indicate that the middle domains play an important role in limiting the activating effects of the C0-C2 domains and that the C4C5 linker contributes to these effects.

Significance Statement

The functional role of the middle domains of cardiac myosin binding protein-C (cMyBP-C) are poorly understood, in part due to technical challenges inherent toin vitromethods that have mainly been used to study recombinant N’-terminal domains in the absence of the whole protein. Here we overcome this barrier by using a “cut and paste” approach, selectively removing and replacing the middle domains of cMyBP-C in permeabilized cardiomyocytes. Substituting the middle domains with titin Ig-like domains resulted in a large increase in myofilament calcium sensitivity, almost instantaneous redevelopment of force, and altered response to rapid stretch. Deletion of only the C4-C5 linker (11 amino acids) qualitatively resulted in the same alterations in force mechanics, albeit to a lesser magnitude. We suggest that the middle domains directly affect the regulation of cardiac muscle function by inhibiting the activating effects of the N’-terminal domains of cMyBP-C.
AliMarko: A Novel Tool for Eukaryotic Virus Identification Using Expert-Guided Approach
Authors: Popov Nikolay , Evdokimova Anastasia , Sonets Ignat , Molchanova Maria , Panova Vera , Korneenko Elena , Manolov Alexander , Ilina Elena
Published: July 2024

Metagenomic sequencing is a valuable tool for studying viral diversity in biological samples. Analyzing this data is complex due to the high variability of viral genomes and their low representation in databases. We present the Alimarko pipeline, designed to streamline virus identification in metagenomic data. A key feature of our tool is the focus on the interpretability of findings: results are provided with tabular and visual information to help determine the confidence level in the identified viral sequences.

The pipeline employs two approaches for identifying viral sequences: mapping to reference genomes and de novo assembly followed by the application of Hidden Markov Models (HMM). Additionally, it includes a step for phylogenetic analysis, which constructs a phylogenetic tree to determine the evolutionary relationships with reference sequences. We also emphasize reducing false-positive results. Reads related to cellular organisms are computationally depleted, and the identified viral sequences are checked against a list of potential contaminants. The output is an HTML document containing visualizations and tabular information designed to assist researchers in making informed decisions about the presence of viruses. Using our pipeline for total RNA sequencing of bat feces, we identified a range of viruses and rapidly determined the validity and phylogenetic relationships of the findings to known sequences with the aid of reports generated by AliMarko.
Multiple cell types support productive infection and dynamic translocation of infectious Ebola virus to the surface of human skin
Authors: Messingham Kelly N. , Richards Paige T. , Fleck Anthony , Patel Radhika A. , Djurkovic Marija , Elliff Jonah , Connell Samuel , Crowe Tyler P. , Gonzalez J. Pablo Munoz , Gourronc Francoise , Dillard Jacob A. , Davey Robert A. , Klingelhutz Al , Shtanko Olena , Maury Wendy
Published: July 2024

Ebola virus (EBOV) within theFiloviridaefamily causes severe human disease. At late stages of infection, EBOV virions are found on the surface of patients’ skin; however, the permissive cell types within the skin and how infectious virus translocates to the apical skin surfaces is not known. Here, we describe a human transwell skin explant culture model and show that EBOV infection of human skin tissues via the basal media results in a time- and dose-dependent increase in infectious virus in dermal and epidermal tissue. Infectious virus was detected on the apical epidermal surface within 3 days, indicating that the virus propagates within and traffics through the tissue. In the dermis, EBOV-infected cells were of myeloid, endothelial and fibroblast origins, whereas keratinocytes harbored virus in the epidermis. Complementary studies showed that both purified skin fibroblasts and keratinocytes supported EBOV infection ex vivo and that both cell types required the phosphatidylserine receptor, Axl, and the endosomal protein, NPC1, for virus entry. Our experimental platform identified new susceptible cell types and demonstrated dynamic trafficking of EBOV virions that resulted in infectious virus on the skin surface; findings that may explain person-to-person transmission via skin contact.

Teaser

Using a human skin explant model, these studies identify and characterize skin cell populations that support Ebola virus infection.
Disentangling the regulatory response ofAgrobacterium tumefaciensCHLDO to glyphosate for engineering whole-cell phosphonate biosensors
Authors: Masotti Fiorella , Krink Nicolas , Lencina Nicolas , Gottig Natalia , Ottado Jorgelina , Nikel Pablo I.
Published: July 2024

Abstract

Phosphonates (PHTs), organic compounds with a stable C—P bond, are widely distributed in nature. Glyphosate (GP), a synthetic PHT, is extensively used in agriculture and has been linked to various human health issues and environmental damage. Given the prevalence of GP, developing cost-effective, on-site methods for GP detection is key for assessing pollution and reducing exposure risks. We adoptedAgrobacterium tumefaciensCHLDO, a natural GP degrader, as the source of genetic parts for constructing PHT biosensors. In this species, thephngene cluster, encoding the C—P lyase pathway, is regulated by the PhnF transcriptional repressor and is part of the Pho regulon. We selected thephnGpromoter, which displays a dose-dependent response to GP, to build a set of whole-cell biosensors. Through stepwise optimization of the transcriptional cascade, we created a biosensor capable of detecting GP in the 0.25-50 μM range in various samples, including soil and water.
Resolving DJ-1 Glyoxalase Catalysis Using Mix-and-Inject Serial Crystallography at a Synchrotron
Authors: Zielinski Kara A. , Dolamore Cole , Dalton Kevin M. , Smith Nathan , Termini John , Henning Robert , Srajer Vukica , Hekstra Doeke R. , Pollack Lois , Wilson Mark A.
Published: July 2024

DJ-1 (PARK7) is an intensively studied protein whose cytoprotective activities are dysregulated in multiple diseases. DJ-1 has been reported as having two distinct enzymatic activities in defense against reactive carbonyl species that are difficult to distinguish in conventional biochemical experiments. Here, we establish the mechanism of DJ-1 using a synchrotron-compatible version of mix-and-inject-serial crystallography (MISC), which was previously performed only at XFELs, to directly observe DJ-1 catalysis. We designed and used new diffusive mixers to collect time-resolved Laue diffraction data of DJ-1 catalysis at a pink beam synchrotron beamline. Analysis of structurally similar methylglyoxal-derived intermediates formed through the DJ-1 catalytic cycle shows that the enzyme catalyzes nearly two turnovers in the crystal and defines key aspects of its glyoxalase mechanism. In addition, DJ-1 shows allosteric communication between a distal site at the dimer interface and the active site that changes during catalysis. Our results rule out the widely cited deglycase mechanism for DJ-1 action and provide an explanation for how DJ-1 produces L-lactate with high chiral purity.
Preparation of oxygen-sensitive proteins for high-resolution cryoEM structure determination using (an)aerobic blot-free vitrification
Authors: Cook Brian D. , Narehood Sarah M. , McGuire Kelly L. , Li Yizhou , Tezcan F. Akif , Herzik Mark A.
Published: July 2024

High-quality grid preparation for single-particle cryogenic electron microscopy (cryoEM) remains a bottleneck for routinely obtaining high-resolution structures. The issues that arise from traditional grid preparation workflows are particularly exacerbated for oxygen-sensitive proteins, including metalloproteins, whereby oxygen-induced damage and alteration of oxidation states can result in protein inactivation, denaturation, and/or aggregation. Indeed, 99% of the current structures in the EMBD were prepared aerobically and limited successes for anaerobic cryoEM grid preparation exist. Current practices for anaerobic grid preparation involve a vitrification device located in an anoxic chamber, which presents significant challenges including temperature and humidity control, optimization of freezing conditions, costs for purchase and operation, as well as accessibility. Here, we present a streamlined approach that allows for the (an)aerobic vitrification of oxygen-sensitive proteins using an automated aerobic blot-free grid vitrification device – the SPT Labtech chameleon. This robust workflow allows for high-resolution structure determination of dynamic, oxygen-sensitive proteins, of varying complexity and molecular weight.
Power Outages: An Underappreciated Risk Factor for Children’s Carbon Monoxide Poisoning
Authors: Northrop Alexander J. , Do Vivian , Flores Nina M. , Wilner Lauren Blair , Sheffield Perry E. , Casey Joan A.
Published: July 2024

Children’s risk of exposure to carbon monoxide (CO) increases after disasters, likely due to improper generator use during power outages. Here, we evaluate the impact of outages on children’s CO-related emergency department (ED) visits in New York State (NYS). We leveraged power outage data spanning 2017-2020 from the NYS Department of Public Service for 1,865 power operating localities (i.e., communities) and defined all-size and large-scale power outage hours. All-size outage hours affected ≥1% of customers, and large-scale outage hours affected ≥20%. We identified CO poisoning using diagnostic codes among those aged <18 between 2017 and 2020 using the Statewide Planning and Research Cooperative System (SPARCS), an all-payer reporting system in NYS. We linked community power outage exposure to patients using the population-weighted centroid of their block group of residence. We estimated the impact of power outages on CO poisoning using a time-stratified case-crossover study design with conditional logistic regression, controlling for daily relative humidity, mean temperature, and total precipitation. Analyses were stratified by urban and rural communities. From 2017-2020, there were 917 pediatric CO poisoning ED visits in NYS. Most cases (83%) occurred in urban region of the state. We observed an association statewide between all-size and large-scale outages and CO ED visits on the index day and the following two days before a return to baseline on lag day 3. Four hours without power increased the odds of a pediatric CO poisoning ED visit by ≥50% for small-scale and ≥150% for large-scale outages, and associations were stronger in urban versus rural areas. While CO poisoning is a relatively rare cause of pediatric ED visits in NYS, it can be deadly and is also preventable. Expanded analyses of the health impacts of outages and advocacy for reliable energy access are needed to support children’s health in a changing climate
Estimating the impact of the RTS,S/AS01 malaria vaccine in Benin: A mathematical modelling study
Authors: Alohoutade Sena , Hounsell Rachel , Silal Sheetal
Published: July 2024

Background

Malaria constitutes a major public health burden in sub-Saharan Africa. It remains the primary health concern in health care institutions and the leading cause of death in children under five years of age. Since October 2021, the World Health Organization (WHO) has recommended the RTS,S/AS01 malaria vaccine in children in sub-Saharan Africa and other high burden areas wherePlasmodium falciparumtransmission is moderate to high. However, with the exception of Ghana, there is a scarcity of studies modelling the potential impact of the RTS,S/AS01 vaccine in the context of West Africa. This study addresses a gap in research by modelling the additional impact of malaria childhood vaccination onPlasmodium falciparummalaria transmission in Benin.

Methods

A compartmental mathematical model has been developed to estimate clinical and severe malaria cases averted in children under the age of five with the primary series (3 doses) of the RTS,S/AS01 malaria vaccine in Benin. Key features of the model include seasonality and the use of insecticide-treated bed nets. Over a period of 10 years, a number of scenarios of vaccine, coverage are simulated. These scenarios included vaccination introduction at different coverage levels to supplement the use of Insecticide-treated bed nets to assess the impact of the RTS,S/AS01 vaccine on malaria transmission in Benin.

Results

The model projected outcomes in terms of malaria burden alleviation by childhood vaccination in Benin. The combination of malaria childhood vaccination at a coverage equivalent to the national DTP3 and the current uptake of insecticide-treated bed nets is projected to lead to a 45% reduction in malaria cases and deaths among children under five years old in the country from 2025 to 2034. However, a decrease in the use of ITN is projected to lead to an increase in malaria burden, surpassing baseline levels despite vaccine introduction.

Conclusion

Child vaccination is expected to avert clinical and severe cases of malaria and is an additional tool to advance malaria control efforts in Benin.
High-dose vitamin D3in the treatment of complicated severe acute malnutrition in Pakistan: a double-blind randomised controlled trial (ViDiSAM)
Authors: Saleem Javeria , Zakar Rubeena , Butt Muhammad Salman , Kaleem Rameeza , Chaudhary Asif , Chandna Jaya , Jolliffe David A , Piper Joseph , Abbas Zaigham , Tang Jonathan C Y , Fraser William D , Freemantle Nick , Prendergast Andrew J , Martineau Adrian R
Published: July 2024

We have previously shown that high-dose vitamin D3improved weight gain and neurodevelopmental indices in children receiving standard therapy for uncomplicated severe acute malnutrition (SAM). We conducted a randomised placebo-controlled trial to determine whether two oral doses of 200,000 international units (IU) vitamin D3would benefit children aged 6-59 months receiving standard therapy for complicated SAM in Lahore, Pakistan. The primary outcome was mean weight-for-height or -length z-score (WHZ) at 2-month follow-up. Secondary efficacy outcomes were mean WHZ at 6-month follow-up and mean lean mass index, Malawi Development Assessment Tool (MDAT) scores and serum 25-hydroxyvitamin D (25[OH]D) concentrations at 2- and 6-month follow-up. 259 children were randomised (128 to vitamin D, 131 to placebo), of whom 251 (96.9%) contributed data to analysis of the primary outcome. At 2-month follow-up, participants allocated to vitamin D had significantly higher mean serum 25(OH)D concentrations than those allocated to placebo (adjusted mean difference [aMD] 100.0 nmol/L, 95% confidence interval [CI] 72.2 to 127.8 nmol/L). This was not associated with an inter-arm difference in mean WHZ (aMD 0.02, 95% CI −0.20 to 0.23), or in any other anthropometric or neurodevelopmental outcome assessed at 2- or 6-month follow-up overall. The intervention was safe. In conclusion, high-dose vitamin D3elevated mean serum 25(OH)D concentrations in children receiving standard therapy for complicated SAM in Pakistan, but did not influence any anthropometric or neurodevelopmental outcome studied. The trial was registered atClinicalTrials.govwith the identifierNCT04270643.
Eye Movement Variations in Indoor, Outdoor, and Reading Scenarios and their Implications for Myopia
Authors: Li Qi , Zhou Chao , Liu Tingting , Han Yingxiang , Wang Dajiang , Wang Xiaofei
Published: July 2024

Purpose

To quantitatively measure eye movement behaviors in indoor, outdoor, and reading scenarios to understand their potential link to myopia.

Methods

Forty-one healthy adult subjects freely viewed indoor and outdoor scenes and performed reading activities using virtual reality (VR). Eye movement data were recorded with the built-in eye tracker of the VR headset (HTC Vive Pro Eye). Gaze and fixation data were calculated and reported for eight regions of the visual field.

Results

Indoor scenes exhibited a more pronounced downward gaze than outdoor environments. Significant differences (p < 0.05) in gaze and fixation behaviors were observed between reading and other scenarios. In region 8 (peripheral inferior visual field), the median (1st quartile, 3rd quartile) number of gaze points were 816 (463, 1175), 1123 (743, 1497), and 1705 (966, 2382) for outdoor, indoor and reading scenarios, respectively. Similarly, fixation behavior counts were 4 (1, 9), 7 (1, 11), and 39 (22, 54), respectively.

Conclusions

Downward gaze and fixation behaviors are more prevalent in reading and indoor environments. Given that downwards eye movements can induce instantaneous axial elongation, our results suggested a potential biomechanical pathway for myopia progression through optic nerve traction and ocular tissue remodeling. This study underscores the need for further research to explore the specific role of eye movement behaviors in the progression of myopia, especially in real-life settings.
Integrated proteasomal and lysosomal activity shape mTOR-regulated proteome remodeling
Authors: Cui Danica S. , Webster Samantha M. , Davis Joseph H.
Published: July 2024

The crucial growth regulator mTOR is suppressed during nutrient limitation, which reduces protein synthesis and activates the ubiquitin-proteasome system (UPS) and lysosomal degradation pathways. Whereas these pathways have been extensively studied individually, their integrated dynamics, including the interplay between protein synthesis and degradation, and the coordination between lysosomal and UPS pathways, remain underexplored. Here, we couple stable isotope pulse-labeling and mass spectrometry to quantify and kinetically model proteome dynamics following mTOR inhibition in cultured human cells. Using a combination of genetics and pharmacological inhibitors, we identify proteins strictly degraded by one pathway, those that undergo multimodal degradation, and others that can flexibly access the proteasome or lysosome subject to the availability of either. Our data resource, comprised of ∼5.2 million proteomic measurements, reveals that the UPS and lysosomal pathway operate with disparate kinetics, and highlights the rapid nature of lysosomal degradation. Additionally, we observe that cells coordinate the synthesis and degradation of translation initiation and elongation factors, leading to preferential synthesis from key classes of mRNA transcripts. Taken together, this work illuminates the complex, integrated pathways influencing proteostasis when mTOR is inhibited, provides a rich resource detailing the kinetics of protein synthesis and degradation, and establishes a robust methodology for measuring proteome dynamics on a per-protein basis in the context of cellular stress.
Structural Dynamics of the Ubiquitin Specific Protease USP30 in Complex with a Cyanopyrrolidine-Containing Covalent Inhibitor
Authors: O’Brien Darragh P , Jones Hannah BL , Shi Yuqi , Guenther Franziska , Vendrell Iolanda , Viner Rosa , Brennan Paul E , Mead Emma , Zarganes-Tzitzikas Tryfon , Davis John B , Pinto-Fernández Adán , England Katherine S , Murphy Emma J , Turnbull Andrew P , Kessler Benedikt M
Published: July 2024

Inhibition of the mitochondrial deubiquitinating enzyme USP30 is neuroprotective and presents therapeutic opportunities for the treatment of idiopathic Parkinson’s Disease and mitophagy-related disorders. We have integrated structural and quantitative proteomics with biochemical assays to decipher the mode of action of covalent USP30 inhibition by a small molecule containing a cyanopyrrolidine reactive group,USP30-I-1. The inhibitor demonstrated high potency and selectivity for endogenous USP30 in neuroblastoma cells. Enzyme kinetics and Hydrogen Deuterium eXchange mass spectrometry (HDX-MS) infers that the inhibitor binds tightly to regions surrounding the USP30 catalytic cysteine and positions itself to form a binding pocket along the thumb and palm domains of the protein, thereby interfering its interaction with ubiquitin substrates. A comparison to a non-covalent USP30 inhibitor containing a benzosulfonamide scaffold revealed a slightly different binding mode closer to the active site Cys77, which may provide the molecular basis for improved selectivity towards USP30 against other members of the DUB enzyme family. Our results highlight advantages in developing covalent inhibitors, such asUSP30-I-1, for targeting USP30 as treatment of disorders with impaired mitophagy.
CryptosporidiumOocyst Wall Proteins are true oocyst wall proteins, with COWP8 functioning to hold the inner and outer layers of the oocyst wall together
Authors: Bacchetti Ross , Stevens Sarah , Lemgruber Leandro , Gonzalez Oliva Mariana Azevedo , Sands Emma M , Alexandrou Konstantinos , Tinti Michele , Robinson Lee , Hanna Jack C , Seizova Simona , Goddard Peyton , Vassalli Massimo , Pawlowic Mattie C
Published: July 2024

Cryptosporidiosis is a significant cause of diarrhoeal disease contributing to substantial morbidity and mortality for the immunocompromised and for young children, especially those who are malnourished. There are no vaccines available and no effective treatments for these patients. Another challenge is thatCryptosporidiaare waterborne and resistant to common water treatments including chlorination.Cryptosporidiaare transmitted as an oocyst that is made up of a hardy oocyst wall that protects four parasites. Little is understood about how the oocyst is constructed, its composition, and the how it resists chlorination. A family of predictedCryptosporidiumOocyst Wall Proteins (COWPs) was identified from the genome. Using a genetic approach, we confirm that all members of the COWP family localise to the oocyst wall. Our studies indicate that COWP2, 3 and 4 localise specifically to the oocyst “suture”, a zipper-like structure on the oocyst wall from which parasites emerge during infection. In parasites lacking COWP8, we observe that the inner and outer layers of the oocyst wall are no longer associated suggesting a role for COWP8 in oocyst wall morphology. Despite loss of COWP8, these transgenic parasites are viable, unchanged in mechanical strength, and retain resistance to chlorination. This work sets the foundation for future exploration ofCryptosporidiumtransmission.
EGFR does not directly interact with cortical actin: A SRRF’n’TIRF Study
Authors: Pandey Shambhavi , Wohland Thorsten
Published: July 2024

The epidermal growth factor receptor (EGFR) governs pivotal signaling pathways in cell proliferation and survival, with mutations implicated in numerous cancers. The organization of EGFR on the plasma membrane (PM) is influenced by the lipids and the cortical actin (CA) cytoskeleton. Despite the presence of a putative actin-binding domain (ABD) spanning 13 residues, a direct interaction between EGFR and CA has not been definitively established. While disrupting the cytoskeleton can impact EGFR behavior, suggesting a connection, the influence of the static actin cytoskeleton has been found to be indirect. Here, we investigate the potential interaction between EGFR and CA, as well as the extent to which CA regulates EGFR’s distribution on the PM using SRRF’n’TIRF, a spatiotemporal super-resolution microscopy technique that provides sub-100 nm resolution and ms-scale dynamics from the same dataset. To label CA, we constructed PMT-mEGFP-F-tractin, which combines an inner leaflet targeting domain PMT, fluorescent probe mEGFP, and the actin-binding protein F-tractin. In addition to EGFR-mEGFP, we included two control constructs: a) an ABD deletion mutant, EGFRΔABD-mEGFP serving as a negative control, and b) EGFR-mApple-F-tractin, where F-tractin is fused to the C-terminus of EGFR-mApple, serving as the positive control. We find that EGFR-mEGFP and EGFRΔABD-mEGFP show similar membrane dynamics, implying that EGFR-mEGFP dynamics and organization are independent of CA. EGFR dynamics show CA dependence when F-tractin is anchored to the cytoplasmic tail. Together, our results demonstrate that EGFR does not directly interact with the CA in its resting and activated state.

SIGNIFICANCE

SRRF’n’TIRF is a spatiotemporal super-resolution microscopy technique that allows for the investigation of plasma membrane-cytoskeleton interactions. We investigate how cortical actin (CA) influences the dynamic behavior and structural organization of EGFR, employing specific probe targeting CA structure and dynamics. Our results suggest that EGFR, whether in its resting or activated state, does not directly bind to or interact with the CA. Any influence of CA on EGFR is indirect through membrane modulating activities of CA.
A facile chemical strategy to synthesize precise AAV-protein conjugates for targeted gene delivery
Authors: Pham Quan , Glicksman Jake , Shahraeini Sadegh , Han Boyang , Jewel Delilah , Loynd Conor , Singha Roy Souyma Jyoti , Chatterjee Abhishek
Published: July 2024

The efficacy of current gene therapy approaches using adeno associated virus (AAV) vectors is limited by the poor control over their tissue tropism. Untargeted AAV vectors require high doses to achieve therapeutic efficacy, which is associated with toxic off-target impacts and increased therapeutic costs. The ability to reprogram existing AAV vectors to selectively transduce target tissues is essential to develop next-generation human gene therapies that are safer, more efficacious, and less expensive. Using selective and high-affinity antibodies and antibody-like proteins to retarget existing AAV vectors to bind novel cell-surface receptors offers an attractive and modular approach to reprogram their tropism. However, attaching these proteins onto the complex and delicate AAV capsids remains challenging. Here, we report a versatile chemical strategy to covalently attach recombinant proteins onto the capsid of AAV, using a combination of genetic code expansion and bioorthogonal conjugation chemistry. This method is efficient, and allows precise control over the site and stoichiometry of protein attachment onto the AAV capsid, enabling systematic optimization of the resulting conjugate. Using this approach, we generated conjugates of AAV2 with an anti-HER2 nanobody and a full-length anti-HER2 IgG, which show highly efficient and selective gene delivery into HER2+ cancer cells. Remarkably, the optimized AAV2-nanobody conjugate facilitated efficient transduction of HER2+ tumor xenograft in mice with little off-target gene expression, including in the liver. Programmable synthesis of AAV-protein conjugates using this method offers a promising new strategy to rationally engineer next-generation gene therapy vectors.