Biology subjects

Lee, S. A.

Publications and source records attributed to Lee, S. A..

At least 19 recordsLinked to original sources

bioRxiv2026

Prenatal diet buffers infant epigenetic changes linked to pollution and transient wheeze

Prenatal air pollution exposure is associated with childhood asthma, particularly among biological males. The mechanisms remain unclear, but may involve lasting epigenetic changes, such DNA methylation (DNAm), that occur during gestation in response to oxidative stress and inflammation. Higher maternal intake of "protective" micronutrients, like antioxidants, could buffer pollution-induced oxidative stress and inflammation to mitigate potentially adverse DNAm differences contributing to asthma. Using data from 515 CANDLE participants, we examined associations between prenatal NO2, PM2.5, and PM10 and cord blood DNAm, evaluated DNAm mediation of pollution associations with childhood wheeze phenotypes (transient, persistent, and late-onset), and assessed buffering of DNAm by maternal polyunsaturated fatty acid, vitamin C, or folate intake, and overall diet quality measured by the Alternative Healthy Eating Index-Pregnancy (AHEI-P). We identified 19, seven, and five regional DNAm differences associated NO2, PM2.5, and PM10. Mediation analyses suggested a role for HLA-DPA1/DPB1 DNAm in NO2 and PM2.5 associations with transient wheeze. To assess buffering, we fit pollutant-by-diet interaction models, defining buffering as an interaction opposite in sign to the main pollutant effect. One or more micronutrients or AHEI-P attenuated pollutant effects at 16 of 19 NO2-associated DMRs, including HLA-DPA1/DPB1, and all PM2.5- and PM10-associated DMRs. However, attenuation of HLA-DPA1/DPB1 DNAm did not significantly reduce the indirect effect of NO2 on transient wheeze. In sex-stratified analyses, biological males exhibited lower PM2.5-associated DNAm in SERPINB9, a gene linked to lung function. These findings suggest prenatal air pollution alters DNAm, which may contribute to transient wheeze, with some differences partially buffered by maternal diet. Significance StatementPrenatal air pollution exposure contributes to child wheeze and asthma, potentially through the oxidative stress response and subsequent changes to infant DNA methylomes. Here, we used data from the CANDLE cohort to identify cord blood DNAm differences associated with NO2, PM2.5, or PM10. We examined if any alterations mediated the relationship between prenatal air pollution exposures and transient, persistent, or late-onset wheeze at age 4 to 6 years. Some of these DNAm differences appeared to be at least partially buffered by maternal micronutrients and/or overall diet quality.

genomics↗

bioRxiv2025

CRISPRi-based functional genomic screening identifies genes essential for CH4-dependent growth in the methanotroph Methylococcus capsulatus Bath.

Aerobic methanotrophic bacteria are the primary organisms that consume atmospheric methane (CH4) and have potential to mitigate the climate-active gas. However, a limited understanding of the genetic determinants of methanotrophy hinders the development of biotechnologies leveraging these unique microbes. Here, we developed and optimized a CRISPR interference (CRISPRi) system to enable functional genomic screening in methanotrophic bacteria. We built a genome-wide single guide RNA (sgRNA) library in the industrial methanotroph, Methylococcus capsulatus, consisting of [~]45,000 unique sgRNAs mediating inducible, CRISPRi-dependent transcriptional repression. A selective screen during growth on CH4 identified 233 genes whose transcription repression resulted in a fitness defect and repression of 13 genes associated with a fitness advantage. Enrichment analysis of the 233 putative essential genes linked many of the encoded proteins with critical cellular processes like ribosome biosynthesis, translation, transcription, and other central biosynthetic metabolism, highlighting the utility of CRISPRi for functional genetic screening in methanotrophs. The CRISPRi screen uncovered that 50 ppb Co2+ in methanotroph Nitrate Mineral Salts medium is growth inhibitory and decreasing Co2+ to < 10 ppb improved biomass productivity from CH4. Collectively, our results show that the CRISPRi system and sgRNA library developed here can be used for facile gene-function analyses and genomic screening to identify novel genetic determinants of methanotrophy and isolation of improved methanotroph biocatalysts.

microbiology↗

bioRxiv2025

NF-κB dependent gene expression and plasma IL-1β, TNFα and GCSF drive transcriptomic diversity and CD4:CD8 ratio in people with HIV on ART.

Despite antiretroviral therapy (ART), people with HIV (PWH) on ART experience higher rates of morbidity and mortality vs. age-matched HIV negative controls, which may be driven by chronic inflammation due to persistent virus. We performed bulk RNA sequencing (RNA-seq) on peripheral CD4+ T cells, as well as quantified plasma immune marker levels from 154 PWH on ART to identify host immune signatures associated with immune recovery (CD4:CD8) and HIV persistence (cell-associated HIV DNA and RNA). Using a novel dimension reduction tool - Pairwise Controlled Manifold Approximation (PaCMAP), we defined three distinct participant transcriptomic clusters. We found that these three clusters were largely defined by differential expression of genes regulated by the transcription factor NF-{kappa}B. While clustering was not associated with HIV reservoir size, we observed an association with CD4:CD8 ratio, a marker of immune recovery and prognostic factor for mortality in PWH on ART. Furthermore, distinct patterns of plasma IL-1{beta}, TNF- and GCSF were also strongly associated with the clusters, suggesting that these immune markers play a key role in CD4+ T cell transcriptomic diversity and immune recovery in PWH on ART. These findings reveal novel subgroups of PWH on ART with distinct immunological characteristics, and define a transcriptional signature associated with clinically significant immune parameters for PWH. A deeper understanding of these subgroups could advance clinical strategies to treat HIV-associated immune dysfunction.

microbiology↗

bioRxiv2024

Aperiodic neural excitation of the prefrontal cortex offsets age-related decrease in hippocampal theta activity for spatial memory maintenance

Currently, there is a critical gap in age-related electrophysiological changes in the human brain and how they are correlated with individual decline and maintenance of spatial cognitive function. To characterize these complex neurocognitive changes using direct intracranial recordings, we isolated periodic band power from the aperiodic spectral slope from iEEG power spectra of 69 presurgical epilepsy patients between 19 and 61 years of age while they performed a virtual spatial navigation task. We found a flattening of aperiodic spectral slope in the prefrontal cortex, but also observed a steepening in the hippocampus, suggestive of region-specific changes in excitatory/inhibitory balance across aging. The hippocampus showed pronounced changes in periodic (oscillatory) activity, including a decrease in theta power that correlated with impaired spatial memory, potentially due to changes in the cholinergic system. Interestingly, individuals with the flatter spectral slope in DLPFC showed preserved performance despite lower hippocampal theta, indicating a potential compensatory mechanism for cognitive maintenance. These findings provide new evidence that individual age-related cognitive decline can be predicted by changes in hippocampal theta oscillation, in combination with concomitant prefrontal compensatory mechanisms.

neuroscience↗

bioRxiv2023

Detection of Blood Volume Reduction and Vasoconstriction Following Focused Ultrasound Blood-Brain Barrier Opening Using Ultrasound Flow Imaging

Microbubble-mediated focused ultrasound (FUS) offers a non-invasive treatment for transient and localized blood-brain barrier (BBB) opening for drug delivery or immunostimulation. It is known that FUS-induced BBB opening is accompanied by blood flow changes, vasoconstriction, and vasodilation, as validated by optical microscopy through a cranial window. In this study, we introduce a novel method for quantifying vascular changes after FUS-induced BBB opening by employing ultrasound flow imaging in mice. We acquired pre-FUS and post-FUS ultrasound flow images with the same microbubble concentration in the brain. Contrast-enhanced power Doppler (CEPD) images and ultrasound localization microscopy images were obtained to evaluate changes in cerebral blood volume and vessel diameter at the sonicated region of the brain. Our findings demonstrate that FUS leads to a reduction in blood volume at the treated region, with vasoconstriction being more dominant than vasodilation. Furthermore, we show that transcranial CEPD can detect local blood reduction following FUS, which spatially coincides with the edema region identified in T2-weighted MRI. Our findings suggest that ultrasound flow imaging has the potential to serve as a cost-effective and immediate monitoring tool for evaluating the safety and efficacy of FUS-induced BBB opening.

bioengineering↗

bioRxiv2023

Stereoscopic RNA Analysis of Transcriptional and Post-transcriptional Regulation

Regulation of gene expression takes place at multiple stages of RNA synthesis, processing, and decay. The dynamics of RNA processing is an essential layer of RNA regulation, required for a deeper understanding of gene expression. Here, we present a streamlined analysis combining nascent RNA sequencing and poly(A) tail length sequencing methods, named Stereoscopic Analysis of Transcriptome (STOAT). Using this analysis, we were able to redefine known and unknown transcripts with high precision, and quantitatively assess RNA expression and stability. We also investigated poly(A) tail processing and their linkage to post-transcriptional features in different cell lines and identified the diversity of microRNA networks in RNA stability and opposing effects of HuR-TTP on poly(A) processing among human cell lines. This method can effectively measure promoter activity, RNA synthesis, poly(A) processing, stability, and decay, providing a comprehensive perspective of the dynamic transcriptome as well as discovering diverse transcript isoforms. MOTIVATIONRNA sequencing (RNA-seq) enables transcriptome-wide mapping and quantitative analysis, but it only provides a static view of the transcriptome. Also, due to its lack of standardization between sequencing platforms and read depth, it compromises reproducibility, and the cost remains prohibitive. To improve these limitations, we are presenting the minimal combination of two methods, nascent RNA sequencing and poly(A) tail length sequencing, which can effectively measure promoter activity, RNA synthesis, abundance, poly(A) processing, stability, and decay, and can discover diverse undefined transcript isoforms. This streamlined analysis of transcription rate and polyadenylation status will provide a comprehensive perspective of the dynamic transcriptome.

genomics↗

bioRxiv2023

Brain Representations of Affective Valence and Intensity in Sustained Pleasure and Pain

Pleasure and pain are two opposites that compete and influence each other, implying the existence of brain systems that integrate them to generate modality-general affective experiences. Here, we examined the brains general affective codes (i.e., affective valence and intensity) across sustained pleasure and pain through an fMRI experiment (n = 58). We found that the distinct sub-populations of voxels within the ventromedial and lateral prefrontal cortices, the orbitofrontal cortex, the anterior insula, and the amygdala were involved in decoding affective valence versus intensity, which was replicated in an independent test dataset (n = 62). The affective valence and intensity models were connected to distinct large-scale brain networks--the intensity model to the ventral attention network and the valence model to the limbic and default mode networks. Overall, this study identified the brain representations of affective valence and intensity across pleasure and pain, promoting the systems-level understanding of human affective experiences.

neuroscience↗

bioRxiv2023

FUS Facilitates Gene Body Export from Nuclear Condensates

Membraneless nuclear condensates are an emerging model of transcriptional regulation that involves spatially clustered enhancers and genes forming transcription condensates (TCs). We used a physical isolation approach and DNA/nascent RNA analysis to investigate the global organization of the genome in TCs. Comparative analysis of the two revealed the dynamic nature of the genome, with stable localization of promoters, dynamic recruitment of enhancers, and dynamic export of gene bodies out of TCs upon gene activation. Our findings also suggest that the RNA binding protein FUS undergoes decondensation upon nascent RNA binding, facilitating gene body export out of TCs.

molecular biology↗

bioRxiv2023

Few-shot learning: temporal scaling in behavioral and dopaminergic learning

Learning the causes of rewards is necessary for survival. Thus, it is critical to understand the mechanisms of such a vital biological process. Cue-reward learning is controlled by mesolimbic dopamine and improves with spacing of cue-reward pairings. However, whether a mathematical rule governs such improvements in learning rate, and if so, whether a unifying mechanism captures this rule and dopamine dynamics during learning remain unknown. Here, we investigate the behavioral, algorithmic, and dopaminergic mechanisms governing cuereward learning rate. Across a range of conditions in mice, we show a strong, mathematically proportional relationship between both behavioral and dopaminergic learning rates and the duration between rewards. Due to this relationship, removing up to 19 out of 20 cue-reward pairings over a fixed duration has no influence on overall learning. These findings are explained by a dopamine-based model of retrospective learning, thereby providing a unified account of the biological mechanisms of learning.

neuroscience↗

bioRxiv2023

Long COVID manifests with T cell dysregulation, inflammation, and an uncoordinated adaptive immune response to SARS-CoV-2

Long COVID (LC), a type of post-acute sequelae of SARS-CoV-2 infection (PASC), occurs after at least 10% of SARS-CoV-2 infections, yet its etiology remains poorly understood. Here, we used multiple "omics" assays (CyTOF, RNAseq/scRNAseq, Olink) and serology to deeply characterize both global and SARS-CoV-2-specific immunity from blood of individuals with clear LC and non-LC clinical trajectories, 8 months following infection and prior to receipt of any SARS-CoV-2 vaccine. Our analysis focused on deep phenotyping of T cells, which play important roles in immunity against SARS-CoV-2 yet may also contribute to COVID-19 pathogenesis. Our findings demonstrate that individuals with LC exhibit systemic inflammation and immune dysregulation. This is evidenced by global differences in T cell subset distribution in ways that imply ongoing immune responses, as well as by sex-specific perturbations in cytolytic subsets. Individuals with LC harbored increased frequencies of CD4+ T cells poised to migrate to inflamed tissues, and exhausted SARS-CoV-2-specific CD8+ T cells. They also harbored significantly higher levels of SARS-CoV-2 antibodies, and in contrast to non-LC individuals, exhibited a mis-coordination between their SARS-CoV-2-specific T and B cell responses. RNAseq/scRNAseq and Olink analyses similarly revealed immune dysregulatory mechanisms, along with non-immune associated perturbations, in individuals with LC. Collectively, our data suggest that proper crosstalk between the humoral and cellular arms of adaptive immunity has broken down in LC, and that this, perhaps in the context of persistent virus, leads to the immune dysregulation, inflammation, and clinical symptoms associated with this debilitating condition.

immunology↗

bioRxiv2023

Differences in expression of tumor suppressor, innate immune, inflammasome, and potassium/gap junction channel host genes significantly predict viral reservoir size during treated HIV infection

The major barrier to an HIV cure is the persistence of infected cells that evade host immune surveillance despite effective antiretroviral therapy (ART). Most prior host genetic HIV studies have focused on identifying DNA polymorphisms (e.g., CCR5{Delta}32, MHC class I alleles) associated with viral load among untreated "elite controllers" (~1% of HIV+ individuals who are able to control virus without ART). However, there have been few studies evaluating host genetic predictors of viral control for the majority of people living with HIV (PLWH) on ART. We performed host RNA sequencing and HIV reservoir quantification (total DNA, unspliced RNA, intact DNA) from peripheral CD4+ T cells from 191 HIV+ ART-suppressed non-controllers. Multivariate models included covariates for timing of ART initiation, nadir CD4+ count, age, sex, and ancestry. Lower HIV total DNA (an estimate of the total reservoir) was associated with upregulation of tumor suppressor genes NBL1 (q=0.012) and P3H3 (q=0.012). Higher HIV unspliced RNA (an estimate of residual HIV transcription) was associated with downregulation of several host genes involving inflammasome (IL1A, CSF3, TNFAIP5, TNFAIP6, TNFAIP9, CXCL3, CXCL10) and innate immune (TLR7) signaling, as well as novel associations with potassium (KCNJ2) and gap junction (GJB2) channels, all q<0.05. Gene set enrichment analyses identified significant associations with TLR4/microbial translocation (q=0.006), IL-1{beta}/NRLP3 inflammasome (q=0.008), and IL-10 (q=0.037) signaling. HIV intact DNA (an estimate of the "replication-competent" reservoir) demonstrated trends with thrombin degradation (PLGLB1) and glucose metabolism (AGL) genes, but data were (HIV intact DNA detected in only 42% of participants). Our findings demonstrate that among treated PLWH, that inflammation, innate immune responses, bacterial translocation, and tumor suppression/cell proliferation host signaling play a key role in the maintenance of the HIV reservoir during ART. Further data are needed to validate these findings, including functional genomic studies, and expanded epidemiologic studies in female, non-European cohorts. Author SummaryAlthough lifelong HIV antiretroviral therapy (ART) suppresses virus, the major barrier to an HIV cure is the persistence of infected cells that evade host immune surveillance despite effective ART, "the HIV reservoir." HIV eradication strategies have focused on eliminating residual virus to allow for HIV remission, but HIV cure trials to date have thus far failed to show a clinically meaningful reduction in the HIV reservoir. There is an urgent need for a better understanding of the host-viral dynamics during ART suppression to identify potential novel therapeutic targets for HIV cure. This is the first epidemiologic host gene expression study to demonstrate a significant link between HIV reservoir size and several well-known immunologic pathways (e.g., IL-1{beta}, TLR7, TNF- signaling pathways), as well as novel associations with potassium and gap junction channels (Kir2.1, connexin 26). Further data are needed to validate these findings, including functional genomic studies and expanded epidemiologic studies in female, non-European cohorts.

genetics↗

bioRxiv2022

Epidermal basal domains organization highlights skin robustness to environmental exposure

Adult interfollicular epidermis (IFE) renewal is likely orchestrated by physiological demands of its complex tissue architecture comprising spatial and cellular heterogeneity. Mouse tail and back skin display two kinds of basal IFE spatial domains that regenerate at different rates. Here we elucidate the molecular and cellular states of basal IFE domains by marker expression and single cell transcriptomics in mouse and human skin. We uncover two paths of basal cell differentiation that reflect in part the IFE spatial domain organization. We unravel previously unrecognized similarities between mouse tail IFE basal domains defined as scales and interscales versus human rete ridges and inter-ridges, respectively. Second, our basal IFE transcriptomics and gene targeting in mice provide evidence supporting a physiological role of IFE domains: adaptation to differential UV exposure. We identify Sox6 as a novel UV-induced and interscale/inter-ridge basal IFE-domain transcription factor, important for IFE proliferation and survival. The spatial, cellular, and molecular organization of IFE basal domains underscores skin adaptation to environmental exposure and its unusual robustness in adult homeostasis. Synopsis O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/481662v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@1565ee8org.highwire.dtl.DTLVardef@170f3e1org.highwire.dtl.DTLVardef@9b67adorg.highwire.dtl.DTLVardef@b3ba40_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2021

Deep Phenotypic Analysis of Blood and Lymphoid T and NK Cells from HIV+ Controllers and Non-Controllers

T and natural killer (NK) cells are effector cells with key roles in anti-HIV immunity, including in lymphoid tissues, the major site of HIV persistence. In this study, we used 42-parameter CyTOF to conduct deep phenotyping of paired blood- and lymph node (LN)-derived T and NK cells from three groups of HIV+ aviremic individuals: elite controllers, and antiretroviral therapy (ART)-suppressed individuals who had started therapy during chronic vs. acute infection, the latter of which is associated with better outcomes. We found that acute-treated individuals are enriched for specific subsets of T and NK cells, including blood-derived CD56-CD16+ NK cells previously associated with HIV control, and LN-derived CD4+ T follicular helper cells with heightened expansion potential. An in-depth comparison of the features of the cells from blood vs. LNs of individuals from our cohort revealed that T cells from blood were more activated than those from LNs. By contrast, LNs were enriched for follicle-homing CXCR5+ CD8+ T cells, which expressed increased levels of inhibitory receptors and markers of survival and proliferation as compared to their CXCR5-counterparts. In addition, a subset of memory-like CD56brightTCF1+ NK cells was enriched in LNs relative to blood. These results together suggest unique T and NK cell features in acute-treated individuals, and highlight the importance of examining effector cells not only in blood but also the lymphoid tissue compartment, where the reservoir mostly persists, and where these cells take on distinct phenotypic features.

immunology↗

bioRxiv2021

mRNA vaccine-induced SARS-CoV-2-specific T cells recognize B.1.1.7 and B.1.351 variants but differ in longevity and homing properties depending on prior infection status

While mRNA vaccines are proving highly efficacious against SARS-CoV-2, it is important to determine how booster doses and prior infection influence the immune defense they elicit, and whether they protect against variants. Focusing on the T cell response, we conducted a longitudinal study of infection-naive and COVID-19 convalescent donors before vaccination and after their first and second vaccine doses, using a high-parameter CyTOF analysis to phenotype their SARS-CoV-2-specific T cells. Vaccine-elicited spike-specific T cells responded similarly to stimulation by spike epitopes from the ancestral, B.1.1.7 and B.1.351 variant strains, both in terms of cell numbers and phenotypes. In infection-naive individuals, the second dose boosted the quantity and altered the phenotypic properties of SARS-CoV-2-specific T cells, while in convalescents the second dose changed neither. Spike-specific T cells from convalescent vaccinees differed strikingly from those of infection-naive vaccinees, with phenotypic features suggesting superior long-term persistence and ability to home to the respiratory tract including the nasopharynx. These results provide reassurance that vaccine-elicited T cells respond robustly to emerging viral variants, confirm that convalescents may not need a second vaccine dose, and suggest that vaccinated convalescents may have more persistent nasopharynx-homing SARS-CoV-2-specific T cells compared to their infection-naive counterparts.

immunology↗

bioRxiv2020

Lack of susceptibility of poultry to SARS-CoV-2 and MERS-CoV

Chickens, turkeys, ducks, quail and geese were challenged with SARS-CoV-2 or MERS-CoV. No disease was observed, no virus replication was detected, and antibodies were not detected in serum. Neither virus replicated in embryonating chickens eggs. Poultry are unlikely to serve a role in the maintenance of either virus.

microbiology↗

bioRxiv2020

Release sites are positioned to activate NMDA receptors

Neurotransmitter is released synchronously and asynchronously following an action potential. The release sites of these two phases are segregated within an active zone, with asynchronous release sites enriched near the center. Here we demonstrate that synchronous and asynchronous release sites are aligned with AMPA receptor and NMDA receptor clusters, respectively. Computational simulations indicate that this spatial and temporal arrangement of release ensures maximal membrane depolarization through AMPA receptors, alleviating the pore-blocking magnesium leading to greater activation of NMDA receptors. Together, these results suggest that release sites are organized to efficiently activate NMDA receptors.

neuroscience↗

bioRxiv2020

Automated Annotation of Cell Identities in Dense Cellular Images

Although identifying cell names in dense image stacks is critical in analyzing functional whole-brain data enabling comparison across experiments, unbiased identification is very difficult, and relies heavily on researchers experiences. Here we present a probabilistic-graphical-model framework, CRF_ID, based on Conditional Random Fields, for unbiased and automated cell identification. CRF_ID focuses on maximizing intrinsic similarity between shapes. Compared to existing methods, CRF_ID achieves higher accuracy on simulated and ground-truth experimental datasets, and better robustness against challenging noise conditions common in experimental data. CRF_ID can further boost accuracy by building atlases from annotated data in highly computationally efficient manner, and by easily adding new features (e.g. from new strains). We demonstrate cell annotation in C. elegans images across strains, animal orientations, and tasks including gene-expression localization, multi-cellular and whole-brain functional imaging experiments. Together, these successes demonstrate that unbiased cell annotation can facilitate biological discovery, and this approach may be valuable to annotation tasks for other systems.

neuroscience↗

bioRxiv2020

No detectable alloreactive transcriptional responses during donor-multiplexed single-cell RNA sequencing of peripheral blood mononuclear cells

Single-cell RNA sequencing (scRNA-seq) provides high-dimensional measurement of transcript counts in individual cells. However, high assay costs limit the study of large numbers of samples. Sample multiplexing technologies such as antibody hashing and MULTI-seq use sample-specific sequence tags to enable individual samples (e.g., different patients) to be sequenced in a pooled format before downstream computational demultiplexing. Critically, no study to date has evaluated whether the mixing of samples from different donors in this manner results in significant changes in gene expression resulting from alloreactivity (i.e., response to non-self immune antigens). The ability to demonstrate minimal to no alloreactivity is crucial to avoid confounded data analyses, particularly for cross-sectional studies evaluating changes in immunologic gene signatures. Here, we compared the expression profiles of peripheral blood mononuclear cells (PBMCs) from a single donor with and without pooling with PBMCs isolated from other donors with different blood types. We find that there was no evidence of alloreactivity in the multiplexed samples following three distinct multiplexing workflows (antibody hashing, MULTI-seq, and in silico genotyping using souporcell). Moreover, we identified biases amongst antibody hashing sample classification results in this particular experimental system, as well as gene expression signatures linked to PBMC preparation method (e.g., Ficoll-Paque density gradient centrifugation with or without apheresis using Trima filtration).

genomics↗