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Biology subjects

Brown, E. A.

Publications and source records attributed to Brown, E. A..

12 recordsLinked to original sources

Lean breast adipocytes secrete an oxylipin that suppresses breast cancer via ferroptosis

Obesity is predicted to become the largest modifiable risk factor for breast cancer in postmenopausal women, yet the mechanisms underlying this association are unclear. We identified a novel role for the endogenous oxylipin 9S-HODE, secreted by lean adipocytes, to induce ferroptosis in breast cancer cells while sparing normal breast epithelial cells. Obese adipocytes fail to secrete 9S-HODE, suggesting that the loss of ferroptosis induction significantly contributes to the acceleration of obesity-associated breast cancer. Consequently, the inhibition of ferroptosis accelerates breast cancer in lean, but not obese, mice. Further, 9S-HODE inhibits the growth of patient-derived breast cancer organoids, and supplementing 9S-HODE into tumors in obese mice is sufficient to reduce tumor burden, underscoring its potential as a therapeutic agent.

cancer biology↗

Controlled decorin delivery from injectable microgels promotes scarless vocal fold repair

Vocal fold (VF) scarring is a leading cause of poor voice, yet no therapies exist to prevent its progression. Current treatments, such as intracordal steroid injections, offer limited efficacy and carry significant off-target toxicities. To identify targeted anti-scarring strategies, we performed transcriptomics of human VF myofibroblasts, the cellular drivers of VF scarring, and identified the proteoglycan decorin (DCN) as downregulated in activated myofibroblasts. We also show a time-dependent decrease in DCN during fibrotic wound healing in a preclinical rat model of VF scarring. Administration of DCN suppressed VF myofibroblast activation by reducing pro-fibrotic gene expression, -smooth muscle actin (-SMA) levels, and cell contractility. DCN was encapsulated in hyaluronic acid microgels for sustained protein release for 3-4 weeks. In a rat model of VF scarring, DCN-loaded microgels prevented hallmark features of scarring, including collagen deposition and myofibroblast activation. These findings highlight DCN as a promising therapeutic and provide a sustained delivery platform with translational potential against VF scarring. TeaserControlled decorin drug delivery from hyaluronic acid microgels attenuates myofibroblast activation to prevent vocal fold scarring, offering a translational strategy to improve post-injury voice outcomes.

bioengineering↗

Dissociation of the mTOR protein interaction network following neuronal activation is altered by Shank3 mutation

The mechanistic target of Rapamycin (mTOR) kinase pathway plays critical roles in neuronal function and synaptic plasticity, and its dysfunction is implicated in numerous neurological and psychiatric disorders. Traditional linear models depict mTOR signaling as a sequential phosphorylation cascade, but accumulating evidence supports a model that includes signaling through dynamic protein-protein interaction networks. To examine how neuronal mTOR signaling discriminates between distinct stimuli, we quantified phosphorylation events and protein co-association networks in primary mouse cortical neurons. Unexpectedly, neuronal mTOR activation by IGF or glutamate triggered dissociation--rather than the anticipated assembly--of protein complexes involving mTOR complex1 (TORC1), mTOR complex 2 (TORC2), and translational machinery, distinguishing neurons from proliferative cells. Applying in vitro homeostatic scaling paradigms revealed distinct combinatorial encoding of synaptic scaling direction: both up- and down-scaling induced dissociation of translational complexes, but downscaling uniquely included dissociation of upstream pathway regulators. Cortical neurons from Shank3B knockout mice, modeling autism-associated Phelan-McDermid Syndrome, displayed baseline hyperactivation of the mTOR network, which reduced the dynamic range of network responses to homeostatic scaling and pharmacological inhibition. These findings reveal that neuronal mTOR signaling employs stimulus-specific combinations of dissociative protein interaction modules to encode opposing forms of synaptic plasticity.

biochemistry↗

finsyncR, an R package to synchronize 27 years of fish and invertebrate data across the United States

The United States government collects some of the most comprehensive data on freshwater systems in existence, but barriers exist for new users to apply these data without institutional guidance. finsyncR (fish and invertebrate synchronizer in R) is an R package that streamlines the process of acquiring, processing, and integrating fish and macroinvertebrate datasets collected in streams and rivers, increasing access to cleaned data and ensuring straightforward application of the data. The data sources are the US Environmental Protection Agencys National River and Streams Assessment and US Geological Surveys BioData. The resulting datasets span 27 years (1993 to 2019) and include 8,115 sites, 15,169 sampling events, 963 macroinvertebrate genera, and 687 fish species. We document common challenges to working with these data including understanding sampling designs, harmonizing taxonomy, calculating densities and standardized abundances, accounting for differences in sampling effort, and considering improvements in taxonomic identifications. We anticipate this package will spur research exploring changes in fish and macroinvertebrate communities across space and time and the causes and consequences of those changes.

ecology↗

Three linked opposing regulatory variants under selection associate with IVD

While genome-wide association studies (GWAS) and selection scans identify genomic loci driving human phenotypic diversity, functional validation is required to discover the variant(s) responsible. We dissected the IVD locus, implicated by selection statistics, multiple GWAS, and clinical genetics as important to function and fitness. We combined luciferase assays, CRISPR/Cas9 genome-editing, massively parallel reporter assays (MPRA), and bashing of regulatory loci. We identified three regulatory variants, including an indel, that may underpin GWAS signals for pulmonary fibrosis and testosterone, and that are linked on a positively selected haplotype in the Japanese population. These regulatory variants exhibit synergistic and opposing effects on IVD expression experimentally. Alleles at these variants lie on a haplotype tagged by the variant most strongly associated with IVD expression and metabolites, but with no functional evidence itself. This work demonstrates how comprehensive functional investigation and multiple technologies are needed to discover the true genetic drivers of phenotypic diversity.

genetics↗

In vitro evolution predicts emerging CoV-2 mutations with high affinity for ACE2 and cross-species binding

Emerging SARS CoV-2 variants are creating major challenges in the ongoing Covid-19 pandemic. Predicting CoV-2 mutations that increase transmissibility or immune evasion would be extremely valuable in development of broad-acting therapeutics and vaccines and prioritising viral monitoring and containment. Using in vitro evolution, we identify a double mutation in CoV-2 receptor binding domain (RBD) that increases affinity for ACE2 almost 20-fold. We determine the mutant:ACE2 structure to reveal the binding mechanism and show the main affinity driver, Q498H, boosts binding of other RBD variants. We find this mutation incompatible with the common N501Y mutation, but N501Y variants can acquire Q498R to access a similar bonding network and affinity gain. We show Q498H, and Q498R plus N501Y, enable variants to bind rat ACE2 with high affinity. These mutations are now emerging in CoV-2 variants, such as the Omicron variant, where they would be expected to drive increased human-to-human and cross-species transmission.

biochemistry↗

Prior dengue immunity enhances Zika virus infection of the maternal-fetal interface in rhesus macaques

Concerns have arisen that pre-existing immunity to dengue virus (DENV) could enhance Zika virus (ZIKV) disease, due to the homology between ZIKV and DENV and the observation of antibody-dependent enhancement (ADE) among DENV serotypes. To date, no study has examined the impact of pre-existing DENV immunity on ZIKV pathogenesis during pregnancy in a translational non-human primate model. Here we show that prior DENV-2 exposure enhanced ZIKV infection of maternal-fetal interface tissues in macaques. However, pre-existing DENV immunity had no detectable impact on ZIKV replication kinetics in maternal plasma, and all pregnancies progressed to term without adverse outcomes or gross fetal abnormalities detectable at delivery. Understanding the risks of ADE to pregnant women worldwide is critical as vaccines against DENV and ZIKV are developed and licensed and as DENV and ZIKV continue to circulate.

microbiology↗

African-lineage Zika virus replication dynamics and maternal-fetal interface infection in pregnant rhesus macaques

Following the Zika virus (ZIKV) outbreak in the Americas, ZIKV was causally associated with microcephaly and a range of neurological and developmental symptoms, termed congenital Zika syndrome (CZS). The isolates responsible for this outbreak belonged to the Asian lineage of ZIKV. However, in-vitro and in-vivo studies assessing the pathogenesis of African-lineage ZIKV demonstrated that African-lineage isolates often replicated to high titer and caused more severe pathology than Asian-lineage isolates. To date, the pathogenesis of African-lineage ZIKV in a translational model, particularly during pregnancy, has not been rigorously characterized. Here we infected four pregnant rhesus macaques with a low-passage strain of African-lineage ZIKV and compared its pathogenesis to a cohort of four pregnant rhesus macaques infected with an Asian-lineage isolate and a cohort of mock-infected controls. Viral replication kinetics were not significantly different between the two experimental groups and both groups developed robust neutralizing antibody titers above levels considered to be protective. There was no evidence of significant fetal head growth restriction or gross fetal harm at delivery in either group. However, a significantly higher burden of ZIKV vRNA was found in maternal-fetal interface tissues in the macaques exposed to an African-lineage isolate. Our findings suggest that ZIKV isolates of any genetic lineage pose a threat to women and their infants. IMPORTANCEZIKV was first identified over 70 years ago in Africa, but most of our knowledge of ZIKV is based on studies of the distinct Asian genetic lineage, which caused the outbreak in the Americas in 2015-16. In its most recent update, the WHO stated that improved understanding of African-lineage pathogenesis during pregnancy must be a priority. Recent detection of African-lineage isolates in Brazil underscores the need to understand the impact of these viruses. Here we provide the first comprehensive assessment of African-lineage ZIKV infection during pregnancy in a translational non-human primate model. We show African-lineage isolates replicate with similar kinetics to Asian-lineage isolates and are capable of infecting the placenta. However, there was no evidence of more severe outcomes with African-lineage isolates. Our results highlight both the threat that African-lineage ZIKV poses to women and their infants and the need for future epidemiological and translational in-vivo studies with African-lineage ZIKV.

microbiology↗

Activity-dependent changes in synaptic protein complex composition are consistent in different detergents despite differential solubility

At the post-synaptic density (PSD), large protein complexes dynamically form and dissociate in response to synaptic activity, comprising the biophysical basis for learning and memory. The use of detergents to both isolate the PSD fraction and release its membrane-associated proteins complicates studies of these activity-dependent protein interaction networks, because detergents can simultaneously disrupt the very interactions under study. Despite widespread recognition that different detergents yield different experimental results, the effect of detergent on activity-dependent synaptic protein complexes has not been rigorously examined. Here, we characterize the effect of three detergents commonly used to study synaptic proteins on activity-dependent protein interactions. We first demonstrate that SynGAP-containing interactions are more abundant in 1% Deoxycholate (DOC), while Shank-, Homer-and mGluR5-containing interactions are more abundant in 1% NP-40 or Triton. All interactions were detected preferentially in high molecular weight (HMW) complexes generated by size exclusion chromatography, although the detergent-specific abundance of proteins in HMW fractions did not correlate with the abundance of detected interactions. Activity-dependent changes in protein complexes were consistent across detergent types, suggesting that detergents do not isolate distinct protein pools with unique behaviors. However, detection of activity-dependent changes is more or less feasible in different detergents due to baseline solubility. Collectively, our results demonstrate that detergents affect the solubility of individual proteins, but activity-dependent changes in protein interactions, when detectable, are consistent across detergent types.

neuroscience↗

An emergent Ebola virus nucleoprotein variant influences virion budding, oligomerization, transcription and replication

For highly pathogenic viruses, reporter assays that can be rapidly performed are critically needed to identify potentially functional mutations for further study under maximal containment (e.g., biosafety level 4 [BSL-4]). The Ebola virus nucleoprotein (NP) plays multiple essential roles during the viral life cycle, yet few tools exist to study the protein under BSL-2 or equivalent containment. Therefore, we adapted reporter assays to measure NP oligomerization and virion-like particle (VLP) production in live cells and further measure transcription and replication using established minigenome assays. As a proof-of-concept, we examined the NP-R111C substitution, which emerged during the 2013-2016 Western African Ebola virus disease epidemic and rose to high frequency. NP-R111C slightly increased NP oligomerization and VLP budding but slightly decreased transcription and replication. By contrast, a synthetic charge-reversal mutant, NP-R111E, greatly increased oligomerization but abrogated transcription and replication. These results are intriguing in light of recent structures of NP oligomers, which reveal that the neighboring residue, K110, forms a salt bridge with E349 on adjacent NP molecules. By developing and utilizing multiple reporter assays, we find that the NP-111 position mediates a complex interplay between NPs roles in protein structure, virion budding, and transcription and replication.

microbiology↗

Untargeted Mass Spectrometry-Based Metabolomics Tracks Molecular Changes in Raw and Processed Foods and Beverages

A major aspect of our daily lives is the need to acquire, store and prepare our food. Storage and preparation can have drastic effects on the compositional chemistry of our foods, but we have a limited understanding of the temporal nature of processes such as storage, spoilage, fermentation and brewing on the chemistry of the foods we eat. Here, we performed a temporal analysis of the chemical changes in foods during common household preparations using untargeted mass spectrometry and novel data analysis approaches. Common treatments of foods such as home fermentation of yogurt, brewing of tea, spoilage of meats and ripening of tomatoes altered the chemical makeup through time, through both chemical and biological processes. For example, brewing tea altered its composition by increasing the diversity of molecules, but this change was halted after 4 min of brewing. The results indicate that this is largely due to differential extraction of the material from the tea and not modification of the molecules during the brewing process. This is in contrast to the preparation of yogurt from milk, spoilage of meat and the ripening of tomatoes where biological transformations directly altered the foods molecular composition. Comprehensive assessment of chemical changes using multivariate statistics showed the varied impacts of the different food treatments, while analysis of individual chemical changes show specific alterations of chemical families in the different food types. The methods developed here represent novel approaches to studying the changes in food chemistry that can reveal global alterations in chemical profiles and specific transformations at the chemical level.\n\nO_LSTHighlightsC_LSTO_LIWe created a reference data set for tomato, milk to yogurt, tea, coffee, turkey and beef.\nC_LIO_LIWe show that normal preparation and handling affects the molecular make-up.\nC_LIO_LITea preparation is largely driven by differential extraction.\nC_LIO_LIFormation of yogurt involves chemical transformations.\nC_LIO_LIThe majority of meat molecules are not altered in 5 days at room temperature.\nC_LI

biochemistry↗

FGF4 Retrogene On CFA12 Is Responsible For Chondrodystrophy And Intervertebral Disc Disease In Dogs

Chondrodystrophy in dogs is defined by dysplastic, shortened long bones and premature degeneration and calcification of intervertebral discs. Independent genome-wide association analyses for skeletal dysplasia (short limbs) within a single breed (pBonferroni=0.0072) and intervertebral disc disease (IVDD) across breeds (pBonferroni=4.02x10-10) both identified a significant association to the same region on CFA12. Whole genome sequencing identified a highly expressed FGF4 retrogene within this shared region. The FGF4 retrogene segregated with limb length and had an odds ratio of 51.23 (95% CI = 46.69, 56.20) for IVDD. Long bone length in dogs is a unique example of multiple disease-causing retrocopies of the same parental gene in a mammalian species. FGF signaling abnormalities have been associated with skeletal dysplasia in humans, and our findings present opportunities for both selective elimination of a medically and financially devastating disease in dogs and further understanding of the ever-growing complexity of retrogene biology.

genetics↗