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Summary: Ovarian cancer is characterized by early metastatic spread. This study demonstrates that carcinoma-associated mesenchymal stromal cells (CA-MSCs) enhance metastasis by increasing tumor cell heterogeneity through mitochondrial donation. CA-MSC mitochondrial donation preferentially occurs in ovarian cancer cells with low levels of mitochondria (“mito poor”). CA-MSC mitochondrial donation rescues the phenotype of mito poor cells, restoring their proliferative capacity, resistance to chemotherapy, and cellular respiration. Receipt of CA-MSC-derived mitochondria induces tumor cell transcriptional changes leading to the secretion of ANGPTL3, which enhances the proliferation of tumor cells without CA-MSC mitochondria, thus amplifying the impact of mitochondrial transfer. Donated CA-MSC mitochondrial DNA persisted in recipient tumor cells for at least 14 days. CA-MSC mitochondrial donation occurs in vivo, enhancing tumor cell heterogeneity and decreasing mouse survival. Collectively, this work identifies CA-MSC mitochondrial transfer as a critical mediator of ovarian cancer cell survival, heterogeneity, and metastasis and presents a unique therapeutic target in ovarian cancer.

The magnitude and timing of extreme events such as debris and floodflows (collectively referred to as floodflows) in post-wildfire basins are difficult to measure and are even more difficult to predict. To address this challenge, a sensor ensemble consisting of noncontact, ground-based (near-field), Doppler velocity (velocity) and pulsed (stage or gage height) radars, rain gages, and a redundant radio communication network was leveraged to monitor flood wave velocities, to validate travel times, and to compliment observations from NEXRAD weather radar. The sensor ensemble (DEbris and Floodflow Early warNing System, DEFENS) was deployed in Waldo Canyon, Pike National Forest, Colorado, USA, which was burned entirely (100 percent burned) by the Waldo Canyon fire during the summer of 2012 (MTBS, 2020).Surface velocity, stage, and precipitation time series collected during the DEFENS deployment on 10 August 2015 were used to monitor and predict flood wave velocities and travel times as a function of stream discharge (discharge; streamflow). The 10 August 2015 event exhibited spatial and temporal variations in rainfall intensity and duration that resulted in a discharge equal to 5.01 cubic meters per second (m3/s). Discharge was estimated post-event using a slope-conveyance indirect discharge method and was verified using velocity radars and the probability concept algorithm. Mean flood wave velocities – represented by the kinematic celerity ck=2.619meterspersecond,m/s±0.556percent and dynamic celerity cd=3.533m/s±0.181percentandtheiruncertainties were computed. L-moments were computed to establish probability density functions (PDFs) and associated statistics for each of the at-a-section hydraulic parameters to serve as a workflow for implementing alert networks in hydrologically similar basins that lack data.Measured flood wave velocities and travel times agreed well with predicted values. Absolute percent differences between predicted and measured flood wave velocities ranged from 1.6 percent to 49 percent and varied with water slope, hydraulic radius, and depth. The kinematic celerity was a better predictor for steep slopes and wide flood plains associated with the Upper Waldo and Middle Waldo radar streamgages; whereas, the dynamic celerity was a better surrogate for shallow slopes and incised channels such as the Lower Waldo radar streamgage.The method demonstrates the potential extensibility of a post-wildfire warning system by (1) leveraging multiple systems (i.e., weather radar, near-field velocity and stage radars, and rain gages) for accurate and timely warnings of debris and floodflows, (2) establishing an order of operations to site, install, and operate near-field radars and conventional rain gages to record floodflows, forecast travel times, and document geomorphic change in this basin and hydrologically similar basins that lack data, and (3) communicating data operationally with the Colorado Department of Transportation engineering staff, National Weather Service forecasters, and emergency managers.

Dynamic changes in the endoplasmic reticulum (ER) morphology are central to maintaining cellular homeostasis. Microtubules (MT) facilitate the continuous remodeling of the ER network into sheets and tubules by coordinating with many ER-shaping protein complexes, although how this process is controlled by extracellular signals remains unknown. Here we report that TAK1, a kinase responsive to various growth factors and cytokines including TGF-β and TNF-α, triggers ER tubulation by activating αTAT1, an MT-acetylating enzyme that enhances ER-sliding. We show that this TAK1/αTAT1-dependent ER remodeling promotes cell survival by actively downregulating BOK, an ER membrane-associated proapoptotic effector. While BOK is normally protected from degradation when complexed with IP3R, it is rapidly degraded upon their dissociation during the ER sheets-to-tubules conversion. These findings demonstrate a distinct mechanism of ligand-induced ER remodeling and suggest that the TAK1/αTAT1 pathway may be a key target in ER stress and dysfunction.

Abstract Floral temperature is a flower characteristic that has the potential to impact the fitness of flowering plants and their pollinators. Likewise, the presence of floral temperature patterns, areas of contrasting temperature across the flower, can have similar impacts on the fitness of both mutualists. It is currently poorly understood how floral temperature changes under the influence of different weather conditions, and how floral traits may moderate these changes. The way that floral temperature changes with weather conditions will impact how stable floral temperatures are over time and their utility to plants and pollinators. The stability of floral temperature cues is likely to facilitate effective plant–pollinator interactions and play a role in the plant's reproductive success. We use thermal imaging to monitor how floral temperatures and temperature patterns of four plant species (Cistus ‘snow fire’ and ‘snow white’, Coreopsis verticillata and Geranium psilostemon) change with several weather variables (illumination, temperature; windspeed; cloud cover; humidity and pressure) during times that pollinators are active. All weather variables influenced floral temperature in one or more species. The directionality of these relationships was similar across species. In all species, light conditions (illumination) had the greatest influence on floral temperatures overall. Floral temperature and the extent to which flowers showed contrasting temperature patterns were influenced predominantly by light conditions. However, several weather variables had additional, lesser, influences. Furthermore, differences in floral traits, pigmentation and structure, likely resulted in differences in temperature responses to given conditions between species and different parts of the same flower. However, floral temperatures and contrasting temperature patterns that are sufficiently elevated for detection by pollinators were maintained across most conditions if flowers received moderate illumination. This suggests the presence of elevated floral temperature and contrasting temperature patterns are fairly constant and may have potential to influence plant–pollinator interactions across weather conditions.

Introduction Autism is a common neurodevelopmental condition with a complex genetic aetiology that includes contributions from monogenic and polygenic factors. Many autistic people have unmet healthcare needs that could be served by genomics-informed research and clinical trials. The primary aim of the European Autism GEnomics Registry (EAGER) is to establish a registry of participants with a diagnosis of autism or an associated rare genetic condition who have undergone whole-genome sequencing. The registry can facilitate recruitment for future clinical trials and research studies, based on genetic, clinical and phenotypic profiles, as well as participant preferences. The secondary aim of EAGER is to investigate the association between mental and physical health characteristics and participants’ genetic profiles.Methods and analysis EAGER is a European multisite cohort study and registry and is part of the AIMS-2-TRIALS consortium. EAGER was developed with input from the AIMS-2-TRIALS Autism Representatives and representatives from the rare genetic conditions community. 1500 participants with a diagnosis of autism or an associated rare genetic condition will be recruited at 13 sites across 8 countries. Participants will be given a blood or saliva sample for whole-genome sequencing and answer a series of online questionnaires. Participants may also consent to the study to access pre-existing clinical data. Participants will be added to the EAGER registry and data will be shared externally through established AIMS-2-TRIALS mechanisms.Ethics and dissemination To date, EAGER has received full ethical approval for 11 out of the 13 sites in the UK (REC 23/SC/0022), Germany (S-375/2023), Portugal (CE-085/2023), Spain (HCB/2023/0038, PIC-164-22), Sweden (Dnr 2023-06737-01), Ireland (230907) and Italy (CET_62/2023, CEL-IRCCS OASI/24-01-2024/EM01, EM 2024-13/1032 EAGER). Findings will be disseminated via scientific publications and conferences but also beyond to participants and the wider community (eg, the AIMS-2-TRIALS website, stakeholder meetings, newsletters).

The co-delivery of microRNAs (miRNAs) and protein-coding RNA presents an opportunity for a combined approach to gene expression and gene regulation for therapeutic applications. Protein delivery is established using long mRNA, self-, and trans-amplifying RNA (taRNA), whereas miRNA delivery typically uses short synthetic oligonucleotides rather than incorporating it as a precursor into long RNA. Although miRNA delivery into the cell cytoplasm using long genomes of RNA viruses has been described, concerns have remained regarding low processing efficiency. However, miRNA precursors can be released from long cytoplasmic alphaviral RNA by a cytoplasmic fraction of Drosha. taRNA, a promising vector platform for infectious disease vaccination, uses a nonreplicating mRNA expressing an alphaviral replicase to amplify a protein-coding short transreplicon-RNA (STR) in trans. To investigate the possibility of simultaneously delivering protein expression and gene silencing, we tested whether a taRNA system can carry and release functional miRNA to target cells. Here, we show that mature miRNA is released from STRs and silences specific targets in a replication-dependent manner for several days without compromising the expression of STR-encoded proteins. Our findings suggest that incorporating miRNAs into the taRNA vector platform has the potential for gene regulation alongside the expression of therapeutic genes.

Porous interpenetrated zirconium–organic frameworks (PIZOFs) are a class of Zr-based metal–organic frameworks (MOFs) which are composed of long, rod-like dicarboxylate linkers and Zr6O4(OH)4(O2C)12 nodes. Long oligoethylene glycol or aliphatic side chains are covalently attached to the linker molecules in the cases of PIZOF-10 and PIZOF-11, respectively. These side chains are supposedly highly mobile, thus mimicking a solvent environment. It is anticipated that such MOFs could be used as a solid catalyst – the MOF – with pore systems showing properties similar to a liquid reaction medium. To quantify the side-chain mobility, here we have applied different 1D and 2D NMR solid-state spectroscopic techniques like cross-polarization (CP) and dipolar-coupling chemical-shift correlation (DIPSHIFT) studies. The rather high 1H-13C CP efficiency observed for the CH2 groups of the side chains indicates that the long side chains are unexpectedly immobile or at least that their motions are strongly anisotropic. More detailed information about the mobility of the side chains was then obtained from DIPSHIFT experiments. Analytical expressions for elaborate data analysis are derived. These expressions are used to correlate order parameters and to slow motional rates with signals in indirect spectral dimensions, thus enabling the quantification of order parameters for the CH2 groups. The ends of the chains are rather mobile, whereas the carbon atoms close to the linker are more spatially restricted in mobility.

The major spliceosome mediates pre-mRNA splicing by recognizing the highly conserved sequences at the 5' and 3' splice sites and the branch point. More than 150 proteins participate in the splicing process and are organized in the spliceosomal A, B, and C complexes. FRA10AC1 is a peripheral protein of the spliceosomal C complex and its ortholog in the green alga facilitates recognition or interaction with splice sites. We identified biallelic pathogenic variants in FRA10AC1 in five individuals from three consanguineous families. The two unrelated Patients 1 and 2 with loss-of-function variants showed developmental delay, intellectual disability, and no speech, while three siblings with the c.494_496delAAG (p.Glu165del) variant had borderline to mild intellectual disability. All patients had microcephaly, hypoplasia or agenesis of the corpus callosum, growth retardation, and craniofacial dysmorphism. FRA10AC1 transcripts and proteins were drastically reduced or absent in fibroblasts of Patients 1 and 2. In a heterologous expression system, the p.Glu165del variant impacts intrinsic stability of FRA10AC1 but does not affect its nuclear localization. By co-immunoprecipitation, we found ectopically expressed HA-FRA10AC1 in complex with endogenous DGCR14, another component of the spliceosomal C complex, while the splice factors CHERP, NKAP, RED, and SF3B2 could not be co-immunoprecipitated. Using an in vitro splicing reporter assay, we did not obtain evidence for FRA10AC1 deficiency to suppress missplicing events caused by mutations in the highly conserved dinucleotides of 5' and 3' splice sites in an in vitro splicing assay in patient-derived fibroblasts. Our data highlight the importance of specific peripheral spliceosomal C complex proteins for neurodevelopment. It remains possible that FRA10AC1 may have other and/or additional cellular functions, such as coupling of transcription and splicing reactions.

Aim: The primary endothelial NADPH oxidase isoform 4 (NOX4) is notably induced during hypoxia, with emerging evidence suggesting its vasoprotective role through H2O2 production. Therefore, we aimed to elucidate NOX4′s significance in endothelial function under hypoxia. Methods: Human vessels, in addition to murine vessels from Nox4−/− mice, were explored. On a functional level, Mulvany myograph experiments were performed. To obtain mechanistical insights, human endothelial cells were cultured under hypoxia with inhibitors of hypoxia-inducible factors. Additionally, endothelial cells were cultured under combined hypoxia and laminar shear stress conditions. Results: In human occluded vessels, NOX4 expression strongly correlated with prostaglandin I2 synthase (PTGIS). Hypoxia significantly elevated NOX4 and PTGIS expression and activity in human endothelial cells. Inhibition of prolyl hydroxylase domain (PHD) enzymes, which stabilize hypoxia-inducible factors (HIFs), increased NOX4 and PTGIS expression even under normoxic conditions. NOX4 mRNA expression was reduced by HIF1a inhibition, while PTGIS mRNA expression was only affected by the inhibition of HIF2a under hypoxia. Endothelial function assessments revealed hypoxia-induced endothelial dysfunction in mesenteric arteries from wild-type mice. Mesenteric arteries from Nox4−/− mice exhibited an altered endothelial function under hypoxia, most prominent in the presence of cyclooxygenase inhibitor diclofenac to exclude the impact of prostacyclin. Restored protective laminar shear stress, as it might occur after thrombolysis, angioplasty, or stenting, attenuated the hypoxic response in endothelial cells, reducing HIF1a expression and its target NOX4 while enhancing eNOS expression. Conclusions: Hypoxia strongly induces NOX4 and PTGIS, with a close correlation between both factors in occluded, hypoxic human vessels. This relationship ensured endothelium-dependent vasodilation under hypoxic conditions. Protective laminar blood flow restores eNOS expression and mitigates the hypoxic response on NOX4 and PTGIS.

Voltage-controlled oscillators (VCOs) can be an excellent means of converting a magnitude into a readable value. However, their design becomes a real challenge for power-and-area-constrained applications, especially when a linear response is required. This paper presents a VCO for smart dust systems fabricated by 65 nm technology. It is designed to minimize leakage, limit high peak currents and provide an output whose frequency variation is linear with the input voltage, while allowing rail-to-rail input range swing. The oscillator occupies 592 μm2, operates in a frequency range from 43 to 53 Hz and consumes a maximum average power of 210 pW at a supply voltage of 1 V and 4 pW at 0.3 V. In addition, the proposed VCO exhibits a quasi-linear response of frequency vs. supply voltage and temperature, allowing easy temperature compensation with complementary to absolute temperature (CTAT) voltage.

Background/Objectives: Enzyme-linked immunosorbent assays (ELISAs) have been used to measure anti-human-papillomavirus (HPV) immunoglobulin IgG. The goal of this study was to evaluate the reproducibility of ELISAs measuring different HPV immunoglobulin isotypes, IgG1, 2, 3, and 4, IgA, and IgM, against HPV16. Methods: Seventy-two serum samples collected from participants in the Costa Rica HPV Vaccine Trial (CVT) and immunized with bivalent HPV vaccine (2vHPV) were used for reproducibility assessment. IgG2 and IgG4 levels were too low to be detected. Levels of IgG1, IgG3, IgA, and IgM were measured, and the data were used to calculate intraclass correlation coefficients (ICCs) and coefficients of variation (CVs). Results: CVs were assessed between technicians (12.8–22.7%) and across days (6.2–30.6%). The overall CVs ranged from 7.7–31.1%. IgM ELISA showed higher CVs (15.8–31.1%) than IgG1, IgG3, and IgA (6.2–22.7%). All ICC values were >98.7%. IgG3 was detected in all samples, while IgG1 and IgA had >86.3% detectability and IgM had 62.1% detectability. Pearson correlational analyses between different antibodies all showed significant correlations (p ≤ 0.001), except when comparing IgGs or IgA to IgM (p = 0.29–0.53). Conclusions: Our data showed that these ELISAs are reproducible and detect isotype antibodies to HPV16 L1 across a range of concentrations in 2vHPV-vaccinated participants.

BackgroundWe aimed to define the clinical and variant spectrum and to provide novel molecular insights into the DHX30-associated neurodevelopmental disorder.MethodsClinical and genetic data from affected individuals were collected through Facebook-based family support group, GeneMatcher, and our network of collaborators. We investigated the impact of novel missense variants with respect to ATPase and helicase activity, stress granule (SG) formation, global translation, and their effect on embryonic development in zebrafish. SG formation was additionally analyzed in CRISPR/Cas9-mediated DHX30-deficient HEK293T and zebrafish models, along with in vivo behavioral assays.ResultsWe identified 25 previously unreported individuals, ten of whom carry novel variants, two of which are recurrent, and provide evidence of gonadal mosaicism in one family. All 19 individuals harboring heterozygous missense variants within helicase core motifs (HCMs) have global developmental delay, intellectual disability, severe speech impairment, and gait abnormalities. These variants impair the ATPase and helicase activity of DHX30, trigger SG formation, interfere with global translation, and cause developmental defects in a zebrafish model. Notably, 4 individuals harboring heterozygous variants resulting either in haploinsufficiency or truncated proteins presented with a milder clinical course, similar to an individual harboring a de novo mosaic HCM missense variant. Functionally, we established DHX30 as an ATP-dependent RNA helicase and as an evolutionary conserved factor in SG assembly. Based on the clinical course, the variant location, and type we establish two distinct clinical subtypes. DHX30 loss-of-function variants cause a milder phenotype whereas a severe phenotype is caused by HCM missense variants that, in addition to the loss of ATPase and helicase activity, lead to a detrimental gain-of-function with respect to SG formation. Behavioral characterization of dhx30-deficient zebrafish revealed altered sleep-wake activity and social interaction, partially resembling the human phenotype.ConclusionsOur study highlights the usefulness of social media to define novel Mendelian disorders and exemplifies how functional analyses accompanied by clinical and genetic findings can define clinically distinct subtypes for ultra-rare disorders. Such approaches require close interdisciplinary collaboration between families/legal representatives of the affected individuals, clinicians, molecular genetics diagnostic laboratories, and research laboratories.