Biology subjects

Hong, T.

Publications and source records attributed to Hong, T..

At least 19 recordsLinked to original sources

bioRxiv2026

Integrative Clinical-Molecular Modeling Identifies LRRN4CL as a Determinant of Structural and Functional Myocardial Improvement

BackgroundMechanical ventricular unloading and systemic circulatory support with left ventricular assist devices (LVADs) enable myocardial recovery in a subset of advanced heart failure (HF) patients, but predictors and mechanisms of recovery are not well understood. Integrating clinical and molecular data may improve identification of patients most likely to recover and uncover biologically relevant targets in HF. MethodsWe collected and analyzed left ventricular apical myocardial tissue and clinical data from 208 patients undergoing LVAD implantation across five centers. Pre-implant transcriptomic profiles (22,373 mRNA transcripts) were integrated with 59 clinical variables using supervised machine learning with repeated cross-validation to identify and prioritize features associated with myocardial recovery, defined as a binary outcome based on improvement in left ventricular ejection fraction (LVEF [≥]40%) and left ventricular end-diastolic diameter (LVEDD [≤]5.9 cm). We also modeled functional (LVEF) and structural (LVEDD) improvement as a continuous outcome without any predefined LVEF and LVEDD pathological thresholds. Feature prioritization was followed by validation in human myocardial tissue and mechanistic interrogation in human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). ResultsIntegrative models achieved modest discrimination for myocardial recovery as a binary categorical outcome (maximum mean cross-validated area under the curve 0.73{+/-}0.15), identifying clinical features such as HF duration, LVEDD, HF pharmacologic therapy, and device configuration. Leucine-rich repeat neuronal 4C-like (LRRN4CL), measured in human myocardium, consistently emerged as a top transcriptomic predictor across both binary and continuous metric models (functional and structural). Higher pre-LVAD LRRN4CL expression was associated with reduced likelihood of myocardial recovery and localized primarily to cardiomyocytes. In iPSC-CMs, LRRN4CL overexpression localized to the sarcoplasmic reticulum, induced transcriptional remodeling characterized by suppression of contractile pathways and activation of stress programs, impaired calcium handling, impaired contraction-relaxation kinetics, and diminished mitochondrial respiratory reserve capacity. ConclusionsIntegration of clinical and myocardial transcriptomic data identifies LRRN4CL as a novel marker associated with impaired myocardial recovery following LVAD-mediated ventricular unloading and systemic circulatory support. These findings move beyond predictive modeling, linking integrative computational discovery to cardiomyocyte dysfunction and providing a translational framework for biologically informed risk stratification and therapeutic targeting for myocardial recovery. CLINICAL PERSPECTIVEO_ST_ABSWhat Is New?C_ST_ABSO_LIIntegrative clinical and myocardial transcriptomic modeling identifies LRRN4CL as a novel molecular determinant of structural and functional changes after LVAD-mediated ventricular unloading and enhanced systemic circulatory support. C_LIO_LIElevated LRRN4CL expression is associated with adverse remodeling signatures, impaired calcium handling, and stress responses in human iPSC-derived cardiomyocytes. C_LIO_LIExperimental overexpression of LRRN4CL directly disrupts calcium cycling, contractile performance, and mitochondrial respiration linking molecular signature to functional phenotype. C_LI What Are the Clinical Implications?O_LIIdentification of LRRN4CL as a marker associated with impaired myocardial recovery supports future efforts toward biologically informed risk stratification for patients undergoing LVAD therapy. C_LIO_LILRRN4CL as a marker of cardiac improvement potential may extend beyond advanced HF to earlier stage disease patients and inform prognosis, risk stratification, and response to medical therapies. C_LIO_LIThese findings highlight LRRN4CL-associated pathways as potential therapeutic targets and demonstrate how integrative clinical-transcriptomic approaches can move beyond clinical prediction toward identification of new biologically precise therapeutic targets in HF following a bedside to bench and back approach. C_LI

bioinformatics↗

bioRxiv2026

Neural correlates of combinatorial reasoning in prefrontal cortex

Complex economic decisions are often combinatorial: they require individuals to select from many alternatives under strict constraints on time, resources, and energy. Combinatorial reasoning is the cognitive process that enables decision makers to construct and evaluate multiple potential solutions in the face of these challenges, but its underlying neural mechanisms are not known. Using a combinatorial optimization paradigm, we show that rhesus macaques adopt distinct reasoning strategies and track the combinatorial upper bound--the best achievable outcome given the options examined so far. We recorded single units from the dorsolateral prefrontal cortex (DLPFC) - a structure central to higher cognition and complex behaviors - while the animals deliberated. Single DLPFC neurons encoded and updated the upper bound in real time, and the neuronal activity scaled with computational requirements. These findings uncover a neural correlate for combinatorial reasoning and reveal how the brain supports complex economic decisions.

neuroscience↗

bioRxiv2026

Regulation of BCL11A DNA binding and expression in human erythrocyte precursor HUDEP-2 cells

BCL11A is a transcription factor crucial for neurodevelopment and hematopoiesis. It regulates the developmental switch from fetal hemoglobin (HbF) to adult hemoglobin and is a major therapeutic target for sickle cell disease and {beta}-thalassemia. BCL11A exists in multiple isoforms, including the L isoform (containing a single two-finger ZF2-3 DNA-binding domain) and the XL isoform (containing two arrays: the two-finger ZF2-3 and the three-finger ZF4-6). We used three approaches to investigate BCL11A functions. First, we examined DNA recognition by BCL11A, which preferentially binds the short 6-bp DNA motif TGNCCA. ZF4-6 recognizes all four variants of this motif with distinct strand-specific interactions: TGTCCA on the top strand, TG(A/C)CCA on the complementary strand, and the palindromic TGGCCA on either strand. ZF2-3 also binds TGTCCA from the top strand, featuring a unique thymine interaction by ZF2 Phe388. Motif multiplicity within BCL11A binding sites may promote BCL11A oligomerization by enabling multiple ZF arrays to engage DNA simultaneously. Second, we treated HUDEP-2 cells (which express adult hemoglobin) with inhibitors targeting three epigenetic silencing marks - DNA methylation, histone H3 lysine 9 methylation or H3 lysine 27 methylation. All treatments, individually or in combination, increased HbF expression to varying degrees. Notably, FTX6058 markedly reduced BCL11A transcription and translation (likely via effects on LIN28B), while EML741 caused a partial reduction. Third, we screened 213 pomalidomide- and lenalidomide-derived compounds and quantified proportions of HbF+ cells by flow cytometry. Effects of four compounds were analyzed by protein mass spectrometry. Although BCL11A levels themselves were unchanged, all four compounds selectively decreased levels of known pomalidomide targets, with consistently decreased levels of the zinc-finger proteins IKZF1 and ZFP91. Together, our studies clarify how BCL11A recognizes DNA, how its expression can be modulated epigenetically, and how small-molecule degraders influence its regulatory network, providing new avenues for HbF reactivation therapies.

biochemistry↗

bioRxiv2026

Direct interactions of CEACAM1 and CD36 with LPS and each other

Micellar lipopolysaccharide (mLPS), a major shed product of gram-negative bacteria in the intestine, may result in TLR4-mediated sepsis in the circulation. CEACAM1 and CD36, expressed on both epithelia and immune cells, regulate TLR4 signaling and inflammation, suggesting a shared role in innate immunity. Furthermore, CEACAM1 associates with CD36 on hepatocytes, regulating lipid storage and bile acids (BAs) in the liver, where LPS detoxification occurs. Using E. coli mLPS-Ra as a model and soluble forms sCEACAM1 or sCD36, we assessed their direct binding to mLPS by SPR, SEC, and TEM. SPR derived steady state affinity constants (KD= 1.47 x 10-6 M and KD= 2.23 x 10-6 M) were obtained for mLPS-Ra binding to sCEACAM1 and sCD36, respectively. mLPS-Ra binding to sCEACAM1 and sCD36 was reduced by bile acids in order of sodium deoxycholate > sodium cholate. mLPS exhibited dose dependent binding to sCEACAM and sCD36 by SEC and the isolated complexes formed well defined micelles on TEM. A KD of 5.28 x 10-8 M was obtained for sCEACAM1 binding to immobilized sCD36. Antibody-based-proximity ligation assays (PLA) demonstrated the association of the ectodomains of CEACAM1 and CD36 on hepatic (P< 0.001) and HEK cells (P< 0.018), while biotin-based PLA demonstrated association of the cytoplasmic domains of CEACAM1 and a CD36-BioID2 fusion protein in HEK cells. Alpha Fold predicted CD36 binding in cis to CEACAM1 in a membrane model, in agreement with the results of the PLAs. Based on the SPR, SEC and TEM binding studies, CEACAM1 and CD36 share a cooperative mLPS binding function distinct from their regulation of TLR4 binding.

biochemistry↗

bioRxiv2026

EMTscore infers divergent EMT pathways from omics data and enables rapid screening for correlated gene sets

SummaryQuantitative analyses of epithelial-mesenchymal transition (EMT) have been widely used in several areas of biomedical sciences due to its importance in development and cancer progression, but its multi-contextual nature requires standardization and implementation of gene set scoring methods beyond capacities of conventional tools. We developed EMTscore, a package that provides an efficient implementation of unbiased scoring methods for multiple EMT pathways using individual single-cell or bulk omics data, and the package allows rapid screening for relationships between EMT and other cellular processes. Availability and ImplementationEMTscore is available from GitHub https://github.com/wenmm/EMTscore under the GNU General Public License, and it will be deposited to Zenodo upon acceptance. It is also under review at Bioconductor. ContactTian Hong (hong@utdallas.edu)

bioinformatics↗

bioRxiv2025

Multi-stem-loop constrained library enables accelerated SELEX for aptamers with superior kinetics and affinity

AbstractAptamers are single-stranded nucleic acids with ligand-binding capacity as cost-effective alternatives to antibodies. However, their utility is often undermined by the inherent conformational instability of single-stranded nucleic acids, which can compromise binding kinetics and final affinity. To overcome this critical limitation, we synthesized a novel primary library incorporating multiple inlaid stem-loop structures to enhance molecular stability and rigidity. This structural constraint dramatically accelerated the selection process, allowing us to isolate aptamer candidates with superior affinity in just three rounds of SELEX (Systematic Evolution of Ligands by EXponential enrichment), a significant reduction compared to conventional methodologies. Critically, the resulting aptamer demonstrated higher affinity and faster binding kinetics compared to previously reported sequences, which directly validate our hypothesis that structural stabilization leads to high-performance aptamers. This methodology, which integrated rational conformational constraint with a high-throughput selection process, offered a generalizable strategy to efficiently select aptamers with excellent kinetic and thermodynamic performance for demanding applications such as continuous, real-time ligand detection.

biochemistry↗

bioRxiv2025

The Pdgfd-Pdgfrb axis orchestrates tumor-nerve crosstalk in pancreatic cancer

Nerves are an integral component of the tumor microenvironment, contributing to cancer progression, metastasis, morbidity, and mortality. In pancreatic ductal adenocarcinoma (PDAC), worse clinical outcomes are associated with perineural invasion (PNI), a process by which cancer cells surround and invade nerves. Here, we employed whole-transcriptome and single-cell spatial transcriptomics to identify candidate tumor-nerve interactions that promote PNI. We discovered that Pdgfd signaling promotes key features of nerve invasion. Mechanistically, Pdgfd stimulated cancer cell invasiveness, neurite outgrowth, and direct physical engagement with glia. Pharmacological blockade of this axis reduced each of these processes in vitro as well as PNI in vivo. Thus, Pdgfd-Pdgfrb signaling mediates PNI by coordinating multifaceted cancer-neuron-glia interactions and represents a promising therapeutic strategy aimed at disrupting harmful cancer-nerve crosstalk.

cancer biology↗

bioRxiv2025

Distinct malignant cell states and myeloid glutamate signaling associated with aggressive pancreatic neuroendocrine tumors

Pancreatic neuroendocrine tumors (PNET) are rare malignancies of the endocrine pancreas with diverse clinical outcomes. While some PNETs are indolent, others are aggressive and metastasize quickly. However, clinically-relevant molecular stratification for PNET to predict outcomes and guide therapeutic decision-making is limited. Thus, there is an urgent need to understand the molecular heterogeneity of PNETs to refine prognostication and discover novel therapeutic vulnerabilities. We performed single-nucleus RNA sequencing on resected primary and metastatic PNETs (n = 20), including two PNETs with neoadjuvant treatment. We inferred gene expression programs (GEPs) of malignant and non-malignant cells and investigated associations with clinical outcomes. Next, we inferred interactions in the tumor microenvironment (TME) and performed transwell assays for functional validation. Finally, we explored genomic and transcriptomic evolution in a unique case study of an untreated primary PNET with two asynchronous hepatic metastases. A malignant GEP enriched for neural/synaptic signaling genes was associated with worse overall survival, broad chromosomal loss of heterozygosity, and alternative lengthening of telomeres. Another malignant GEP enriched for VEGF signaling increased throughout metastatic progression in our case study. We found that macrophage-derived glutamate drives polarization towards an immunosuppressive phenotype and activates the MAPK/ERK pathway in malignant cells to increase migratory capacity. This study provides a detailed single-nucleus transcriptomic classification of malignant, stromal, and immune cell types and states in PNETs, their interactions in the TME, and associations with clinical outcomes. The refined molecular taxonomy of PNET may guide the development of more efficacious biomarkers and therapeutic strategies.

cancer biology↗

bioRxiv2025

A continuum reaction-diffusion model for spread of gene silencing in chromosomal inactivation

Regulation of gene silencing in large regions of chromosomes is crucial for development and disease progression, and there has been an increasing interest in using it for new therapeutics. One example of massive gene silencing is X chromosome inactivation (XCI), a process essential for dosage compensation of X-linked genes. During XCI, most genes in the X chromosome are inactivated following the transcription of XIST, an X-linked long noncoding RNA. Recent experiments with transgenes showed that the spread of gene silencing can be induced by XIST transcription in cis, but the spread is restricted in space. The mechanism of controlling the spread remains unclear. In this work, we develop a continuum reaction-diffusion model that elucidates chromosomal inactivation through a bistable system governed by a regulatory network for XIST-mediated gene silencing. We find that the spread of XIST can be tuned by known negative feedback loops regulating its synthesis and degradation, and that the spread of gene silencing is controlled by a wave-pinning mechanism in which both global regulation of silencing complex and local variations of histone modifications can play crucial roles. In addition, we integrate the discrete three-dimensional arrangement of the X chromosome and autosomes into this continuous model. We use a 3D chromosome structure inferred from experimental data and our modeling framework to show the spatiotemporal regulation for spread of gene silencing. Our method enables the investigation for the inactivation dynamics of large regions of chromosomes with varying degrees of the spread of gene silencing. Our model provides mechanistic insights that quantitatively relate gene regulatory networks to tunability and stability of chromosomal inactivation. Author SummaryPrecise control of gene expression is a fundamental process in biology and turning off large parts of chromosomes is both common in many species and important for development and diseases. A well-known example of chromosomal scale gene silencing is X chromosome inactivation (XCI), which helps balancing gene activity between sexes. XCI is governed by the production of an RNA called XIST, leading to most genes on one X chromosome being turned off. Experiments have shown that XIST can trigger nearby genes to turn off in natural and engineered contexts, but this effect only spreads to specific regions. The exact mechanism for this limit is not very well understood. In this study, the authors created a mathematical model to explain how XIST spreads and controls gene activity. They found that feedback systems involving XIST regulations and chromatin modifications synergize and determine how far the inactivation spreads, through a process similar to a wave that gets "pinned" in place. They also included the 3D shape of chromosomes in their model to better understand how gene silencing happens over time and space. This model helps to quantitatively describe chromosome inactivation with gene regulatory networks.

systems biology↗

bioRxiv2025

Peroxisome proliferator-activated receptor gamma (PPARG)-mediated myocardial salvage in acute myocardial infarction managed with left ventricular unloading and coronary reperfusion

Ischemic heart disease and acute myocardial infarction (AMI) is a leading cause of morbidity and mortality. Improvements have been made in coronary interventions to restore blood flow, but ischemia/reperfusion (I/R) injury significantly impacts clinical outcomes. We previously reported that activation of percutaneous mechanical unloading of the left ventricle (LV) with a transvalvular axial-flow device simultaneously with reperfusion improves myocardial salvage. However, the underlying mechanisms, potential adjuvant pharmacological interventions and the timing of the use of LV unloading as a cardioprotective approach in AMI are not well understood. This study investigated a) the mechanisms associated with improved myocardial salvage, b) a pharmacological intervention, and c) the timing of LV unloading. Following 90 minutes of ischemia, adult swine were subjected to reperfusion alone, simultaneous unloading with reperfusion, upfront unloading with delayed reperfusion, upfront reperfusion with delayed unloading, or reperfusion with concurrent use of esmolol and milrinone. Compared to controls, the simultaneous group had a 47% increase in myocardial salvage following AMI. This was associated with increased expression of neutrophil degranulation, macrophage activation, iNOS signaling, wound healing, and PPAR signaling. From these pathways, PPARG (peroxisome proliferator-activated receptor gamma) emerged as a potential cardioprotective gene that was uniquely overexpressed in the simultaneously unloaded and reperfused myocardium. Next, we showed PPARG agonism with rosiglitazone reduces mitochondrial oxygen demand in cardiomyocytes and in vivo, improves myocardial salvage following I/R injury in C57BL6/J mice. Thiazolidinediones (TZDs), such as rosiglitazone could be investigated as therapies combined with simultaneous LV unloading and coronary interventions to mitigate reperfusion injury. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=129 SRC="FIGDIR/small/637726v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1c4d6e6org.highwire.dtl.DTLVardef@1f8d064org.highwire.dtl.DTLVardef@396eddorg.highwire.dtl.DTLVardef@9d932f_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2025

Intercellular signaling stabilizes single-cell level phenotypic transitions and accelerates the reestablishment of equilibrium of heterogeneous cancer cell populations

BackgroundCancer cells within tumors exhibit a wide range of phenotypic states driven by non-genetic mechanisms in addition to extensively studied genetic alterations. Conversions among cancer cell states can result in intratumoral heterogeneity which contributes to metastasis and development of drug resistance. However, mechanisms underlying the initiation and/or maintenance of such phenotypic plasticity are poorly understood. In particular, the role of intercellular communications in phenotypic plasticity remains elusive. MethodsIn this study, we employ a multiscale inference-based approach using single-cell RNA sequencing (scRNA-seq) data to explore how intercellular interactions influence phenotypic dynamics of cancer cells, particularly cancers undergoing epithelial-mesenchymal transition. In addition, we use mathematical models based on our data-driven findings to interrogate the roles of intercellular communications at the cell populations from the viewpoint of dynamical systems. ResultsOur inference approach reveals that signaling interactions between cancerous cells in small cell lung cancer (SCLC) result in the reinforcement of the phenotypic transition in single cells and the maintenance of population-level intratumoral heterogeneity. Additionally, we find a recurring signaling pattern across multiple types of cancer in which the mesenchymal-like subtypes utilize signals from other subtypes to support its new phenotype, further promoting the intratumoral heterogeneity. Our models show that inter-subtype communication both accelerates the development of heterogeneous tumor populations and confers robustness to their steady state phenotypic compositions. ConclusionsOur work highlights the critical role of intercellular signaling in sustaining intratumoral heterogeneity, and our approach of computational analysis of scRNA-seq data can infer inter- and intra-cellular signaling networks in a holistic manner.

systems biology↗

bioRxiv2024

Cell Type Specific Enhancers for Dorsolateral Prefrontal Cortex.

The dorsolateral prefrontal cortex (DLPFC) is crucial to primate cognitive functions, but a paucity of cell type specific tools limits studies of DLPFC neurocomputational principles. Therefore, we set out to identify enhancers that fit inside Adeno-Associated Virus (AAV) vectors and that elicited functional, cell type specific gene expression in the non-human primate (NHP) DLPFC. We used single nucleus RNA-Seq and ATAC-Seq from rhesus macaque tissue samples to define DLPFC cell types and their associated open chromatin regions (OCRs). We trained machine learning (ML) models to recognize the unique regulatory grammar associated with each DLPFC neuron type, performed in silico screening of all OCRs, and identified candidate enhancers most likely to elicit cell type specific transgene expression in each neuron type. For layer 3 pyramidal neurons (L3PNs) and layer 5 extratelencephalic neurons (L5ETs), we cloned the top twelve identified candidates into AAVs and injected them into NHP DLPFC. In situ observation of enhancer-driven expression revealed the best performers, RMacL3-01 and RMacL5ET-01. We validated RMacL3-01 and RMacL5ET-01 using one-at-a-time injections in NHP DLPFC. RMacL3-01 restricted GFP expression to pyramidal neurons in layers 2 and 3, whereas RMacL5ET-01 restricted expression to POU3F1+ neurons in layer 5. RMacL3-01 elicited functional levels of channelrhodopsin expression that enabled optical activation of single- and multi-unit activity in NHP DLFPC. Together, these results and resources establish a solid foundation to study cell type specific principles of primate cognitive functions.

neuroscience↗

bioRxiv2024

Rearrangement of 3D genome organization in breast cancer epithelial - mesenchymal transition and metastasis organotropism

Breast cancer cells exhibit organotropism during metastasis, showing preferential homing to certain organs such as bone, lung, liver, and brain. One potential explanation for this organotropic behavior is that cancer cells gain properties that enable thriving in certain microenvironments. Such specific metastatic traits may arise from gene regulation at the primary tumor site. Spatial genome organization plays a crucial role in oncogenic transformation and progression, but the extent to which chromosome architecture contributes to organ-specific metastatic traits is unclear. This work characterizes chromosome architecture changes associated with organotropic metastatic traits. By comparing a collection of genomic data from different subtypes of localized and lung metastatic breast cancer cells with both normal and cancerous lung cells, we find important trends of genomic reorganization. The most striking differences in 3D genome compartments segregate cell types according to their epithelial vs. mesenchymal status. This EMT compartment signature occurs at genomic regions distinct from transcription-defined EMT signatures, suggesting a separate layer of regulation. Specifically querying organotropism, we find 3D genome changes consistent with adaptations needed to survive in a new microenvironment, with lung metastatic breast cells exhibiting compartment switch signatures that shift the genome architecture to a lung cell-like conformation and brain metastatic prostate cancer cells showing compartment shifts toward a brain-like state. TCGA patient data reveals gene expression changes concordant with these organ-permissive compartment changes. These results suggest that genome architecture provides an additional level of cell fate specification informing organotropism and enabling survival at the metastatic site. SIGNIFICANCEComputational analysis of a cohort of cancer cell lines reveals some 3D genome spatial compartment changes are associated with transitions in cancer cell state that relate to metastasis (EMT) and others reflect the characteristics of the metastatic organ context.

cancer biology↗

bioRxiv2024

Using Genomic Tools to Predict Antimicrobial Resistance and Markers in Clinical Bacterial Samples

Lysogenic bacteriophages can integrate their genome into the bacterial chromosome in the form of a prophage and can promote genetic transfer between bacterial strains in vitro. However, the contribution of lysogenic phages to the incidence of antimicrobial resistance (AMR) in clinical settings is poorly understood. Here, in a set of 186 clinical isolates of Pseudomonas aeruginosa collected from respiratory cultures from 82 patients with cystic fibrosis (CF), we evaluate the links between prophage counts and both genomic and phenotypic resistance to six anti-pseudomonal antibiotics: tobramycin, colistin, ciprofloxacin, meropenem, aztreonam, and piperacillin-tazobactam. We identified 239 different prophages in total. We find that P. aeruginosa isolates contain on average 3.06 +/- 1.84 (SD) predicted prophages. We find no significant association between the number of prophages per isolate and the minimum inhibitory concentration (MIC) for any of these antibiotics. We then investigate the relationship between particular prophages and AMR. We identify a single lysogenic phage associated with phenotypic resistance to the antibiotic tobramycin and, consistent with this association, we observe that AMR genes associated with resistance to tobramycin are more likely to be found when this prophage is present. However we find that they are not encoded directly on prophage sequences. Additionally, we identify a single prophage statistically associated with ciprofloxacin resistance but do not identify any genes associated with ciprofloxacin phenotypic resistance. These findings suggest that prophages are only infrequently associated with the AMR genes in clinical isolates of P. aeruginosa. ImportanceAntibiotic-resistant infections of Pseudomonas aeruginosa, a leading pathogen in patients with Cystic Fibrosis (CF) are a global health threat. While lysogenic bacteriophages are known to facilitate horizontal gene transfer, their role in promoting antibiotic resistance in clinical settings remains poorly understood. In our analysis of 186 clinical isolates of P. aeruginosa from CF patients, we find that prophage abundance does not predict phenotypic resistance to key antibiotics but that specific prophages are infrequently associated with tobramycin resistance genes. In addition, we do not find antimicrobial resistance (AMR) genes encoded directly on prophages. These results highlight that while phages can be associated with AMR, phage-mediated AMR transfer may be rare in clinical isolates and difficult to identify. This work is important for future efforts on mitigating AMR in Cystic Fibrosis and other vulnerable populations affected by Pseudomonas aeruginosa infections and advances our understanding of bacterial-phage dynamics in clinical infections.

microbiology↗

bioRxiv2024

Hypothalamus-specific NSCs derived from hPSCs ameliorate age-associated dysfunction upon transplantation into aged mouse hypothalamus

The hypothalamus is the brain region that regulates systemic body metabolism and multiple brain functions. The adult hypothalamus harbors neural stem/precursor cell (NSC)-like cells. Along with age-related body changes, the hypothalamic NSC (htNSC) population declines, indicating the potential of htNSC replacement as an anti-aging strategy. Here, we developed protocol to generate htNSCs from human pluripotent stem cells (hPSCs). Implanting the hPSC-derived htNSCs into the hypothalamus of aged mice ameliorated age-related declines in metabolic fitness, physical capacity, and cognitive function. Mechanistically, these anti-aging effects were mediated by inter-tissue communication: enhanced neuronal activity in the htNSC-transplanted hypothalamus stimulated adipose tissues to produce and release the anti-aging molecule eNAMPT into systemic circulation via the sympathetic nervous system. Concurrently, the aged inflammatory environment in the hypothalamus was alleviated by peripheral anti-aging signals. Collectively, our findings support the potential of anti- or healthy aging therapies by targeting hypothalamus.

cell biology↗

bioRxiv2024

The subcommissural organ regulates brain development via secreted peptides

The subcommissural organ (SCO) is a gland located at the entrance of the aqueduct of Sylvius in the brain. It exists in species as distantly related as amphioxus and humans, but its function is largely unknown. To explore its function, we compared transcriptomes of SCO and non-SCO brain regions and found three genes, Sspo, Car3, and Spdef, that are highly expressed in the SCO. Mouse strains expressing Cre recombinase from endogenous promoter/enhancer elements of these genes were used to genetically ablate SCO cells during embryonic development, resulting in severe hydrocephalus and defects in neuronal migration and development of neuronal axons and dendrites. Unbiased peptidomic analysis revealed enrichment of three SCO-derived peptides, namely thymosin beta 4, thymosin beta 10, and NP24, and their reintroduction into SCO-ablated brain ventricles substantially rescued developmental defects. Together, these data identify a critical role for the SCO in brain development.

neuroscience↗

bioRxiv2023

The Arp2/3 complex promotes periodic removal of Pak1-mediated negative feedback to facilitate anticorrelated Cdc42 oscillations.

The conserved GTPase Cdc42 is a major regulator of polarized growth in most eukaryotes. Cdc42 periodically cycles between active and inactive states at sites of polarized growth. These periodic cycles are caused by positive feedback and time-delayed negative feedback loops. In the bipolar yeast S. pombe, both growing ends must regulate Cdc42 activity. At each cell end, Cdc42 activity recruits the Pak1 kinase which prevents further Cdc42 activation thus establishing negative feedback. It is unclear how Cdc42 activation returns to the end after Pak1-dependent negative feedback. Using genetic and chemical perturbations, we find that disrupting branched actin-mediated endocytosis disables Cdc42 reactivation at the cell ends. With our experimental data and mathematical models, we show that endocytosis-dependent Pak1 removal from the cell ends allows the Cdc42 activator Scd1 to return to that end to enable reactivation of Cdc42. Moreover, we show that Pak1 elicits its own removal via activation of endocytosis. In agreement with these observations, our model and experimental data show that in each oscillatory cycle, Cdc42 activation increases followed by an increase in Pak1 recruitment at that end. These findings provide a deeper insight into the self-organization of Cdc42 regulation and reveal previously unknown feedback with endocytosis in the establishment of cell polarity.

cell biology↗

bioRxiv2023

An updated model of shoot apical meristem regulation by ERECTA family and CLAVATA3 signaling pathways

The shoot apical meristem (SAM) gives rise to above-ground organs. The size of the SAM is relatively constant due to the balance of stem cell replenishment versus cell recruitment into developing organs. In angiosperms, the transcription factor WUSCHEL (WUS) promotes stem cell identity in the central zone of the SAM. WUS forms a negative feedback loop with a signaling pathway activated by CLAVATA3 (CLV3). In the periphery of the SAM, the ERECTA family (ERf) receptors promote cell differentiation and constrain the expression of WUS and CLV3. Here, we show that four ligands of ERfs redundantly inhibit CLV3 and WUS expression. Transcriptome analysis confirmed that WUS and CLV3 are the main targets of ERf signaling and uncovered several new ones. Analysis of promoter reporters indicated that in the vegetative meristem, the WUS expression domain mostly overlapped with the CLV3 domain and did not shift along the apical-basal axis in clv3. A 3D mathematical model reproduced the experimentally observed CLV3 and WUS expression patterns with fewer assumptions than earlier models. Based on these findings, we propose that CLV3 regulates cellular levels of WUS expression through autocrine signaling, while ERfs regulate WUS spatial expression, preventing its encroachment into the peripheral zone. One Sentence SummaryThrough autocrine signaling, CLV3 regulates the level of WUS expression in the vegetative SAM but not its location, while ERfs regulate the WUS spatial pattern, preventing its expansion into the peripheral zone.

plant biology↗