Biology subjects

Zhou, L.

Publications and source records attributed to Zhou, L..

At least 91 records · Page 5Linked to original sources

bioRxiv2025

α7 nicotinic acetylcholine receptors regulate radial glia fate in the developing human cortex

Prenatal nicotine exposure impairs fetal cortical grey matter volume, but the precise cellular mechanisms remain poorly understood. This study elucidates the role of nicotinic acetylcholine receptors (nAChRs) in progenitor cells and radial glia (RG) during human cortical development. We identify two nAChR subunits--CHRNA7 and the human-specific CHRFAM7A--expressed in SOX2+ progenitors and neurons, with CHRFAM7A particularly enriched along RG endfeet. nAChR activation in organotypic slices and dissociated cultures increases RG proliferation while decreasing neuronal differentiation, whereas nAChR knockdown reduces RG and increases neurons. Single-cell RNA sequencing reveals that nicotine exposure downregulates key genes in excitatory neurons (ENs), with CHRNA7 or CHRFAM7A selectively modulating these changes, suggesting an evolutionary divergence in regulatory pathways. Furthermore, we identify YAP1 as a critical downstream effector of nAChR signaling, and inhibiting YAP1 reverses nicotine-induced phenotypic alterations in oRG cells, highlighting its role in nicotine-induced neurodevelopmental pathophysiology.

neuroscience↗

bioRxiv2025

Mosaic H3K9me3 at BREACHes predicts synaptic gene expression associated with fragile X syndrome cognitive severity

Diseases vary in clinical presentation across individuals despite the same molecular diagnosis. In fragile X syndrome (FXS), mutation-length expansion of a CGG short tandem repeat (STR) in FMR1 causes reduced gene expression and FMRP loss. Nevertheless, FMR1 and FMRP are limited predictors of adaptive functioning and cognition in FXS patients, suggesting that molecular correlates of clinical measures would add diagnostic value. We recently uncovered Megabase-scale domains of heterochromatin (BREACHes) in FXS patient-derived iPSCs. Here, we identify BREACHes in FXS brain tissue (N=4) and absent from sex/age-matched neurotypical controls (N=4). BREACHes span >250 genes and exhibit patient-specific H3K9me3 variation. Using N=4 FXS iPSC lines and N=7 single-cell isogenic FXS iPSC subclones, we observe a strong correlation between inter-sample H3K9me3 variation and heterogeneous BREACH gene repression. We demonstrate improved prediction of cognitive metrics in FXS patients with an additive model of blood FMRP and mRNA levels of H3K9me3-mosaic, but not H3K9me3-invariant, BREACH genes. Our results highlight the utility of H3K9me3 variation at BREACHes for identifying genes associated with FXS clinical metrics.

genomics↗

bioRxiv2025

Distinct forms of amyloid-β moderate sleep duration through NAD+-linked redox metabolism in Alzheimer's disease.

Sleep disruptions precede a clinical diagnosis of Alzheimers disease (AD) by several years. However, how AD pathologies affect sleep remains unclear. Here, we integrate epidemiological data with insights from Drosophila models of AD to investigate how AD progression could be linked to sleep disruption. We found that individuals with a high risk of AD report a shorter sleep duration than do those with a clinical diagnosis of AD. We showed that the expression of different forms of amyloid-{beta} in flies can replicate these variations in sleep duration. Analysis of the metabolome and proteome of these flies revealed distinct changes in NAD+-linked redox metabolism and levels of hyperkinetic (Hk), a redox-sensing sleep homeostat. We showed that the genetic upregulation of Hk is neuroprotective and increased KCNAB2 expression in the brain decreases AD risk in humans. Overall, our data provides a new mechanism linking the disruption of NAD+-linked redox sensing and sleep disruption in AD.

neuroscience↗

bioRxiv2025

CILIATED CELL DOMAINS WITH LOCALLY COORDINATED CILIARY MOTION GENERATE A MOSAIC OF MICROFLOWS IN THE BRAIN'S LATERAL VENTRICLES

Circulation of cerebrospinal fluid (CSF) through the brains ventricles is essential for maintaining brain homeostasis and supporting neurogenesis. CSF flow is supported by the structural polarization of multiciliated cells, which align with the flow direction. However, it remains unclear how the organization of tissue-wide polarity across the ciliary epithelium comprised of thousands of cells, determines the trajectory of the flow and efficient distribution of the CSF. Here, we used new approaches to analyze the organization of translational polarity across extensive areas of the lateral ventricular wall. We also used live imaging to examine cilia motion, flow trajectories, and ciliary beat frequency (CBF) in live preparations of ventricles. In addition to the primary flow running across the ventricular wall from the posterior area to the anterior (P-A), we found multiple local microflows with both direct and curved trajectories that deviate from the mainstream P-A direction. Our results suggest that the ciliated epithelium in the lateral ventricles varies in the alignment of ciliated cell translational polarity: whereas in the narrow dorsal area translational polarity is aligned with the direction of the mainstream flow, in the periphery of the mainstream it is organized into distinct cell clusters with locally aligned polarity vectors. We posit that the cluster organization of the multiciliated ependymal cells underpins the generation of a complex mosaic of flows, with the local microflows facilitating the wide spreading of the CSF across the ependyma. We demonstrate that nNOS is involved in the control of translational polarity, cluster organization, microflows, and CBF in the ependyma. SignificanceThe flow of cerebrospinal fluid (CSF) is crucial for brain homeostasis. This flow is driven by the coordinated beating of cilia on thousands of ciliated cells. Planar polarity vector of the ciliated cells is aligned with the flow direction in the areas of the ependyma underlying the mainstream flow (1). However, the overall alignment of planar cell polarity and flow across the whole tissue is unclear. Here we report a discovery of complex flow patterns over the ependyma, consisting of numerous microflows in the periphery of the mainstream flow. Using new approaches, we found that planar polarity of the ependymal ciliated cells on the periphery of the mainstream flow aligns only locally, indicating a clustered organization of the ependyma that supports various flow directions.

cell biology↗

bioRxiv2025

LDB1 regulates gene expression and chromatin structure in pluripotency and lineage differentiation

Chromatin organization is a pivotal factor in stem cell pluripotency and differentiation. However, the role of enhancer looping protein LDB1 in stem cells has not been explored. We generated Ldb1(-/-) embryonic stem cells (ESC) using CRISPR/Cas9 editing and observed a reduction in key stem cell factors SOX2 and KLF4 upon LDB1 loss. Embryoid bodies (EB) derived from Ldb1(-/-) ESC displayed reduced expression of lineage-specific markers and impaired ability to undergo terminal differentiation to erythroblasts. Differential gene expression, including of the Lin28-mediated self-renewal pathway genes, was observed between WT and Ldb1(-/-) ESC and EB but was most pronounced after differentiation to erythroblasts. LDB1 occupied super enhancers, including those of pluripotency genes, in ESC together with pluripotency factors. LDB1 loss resulted in globally decreased chromatin accessibility in ESC and EB. Conditional LDB1-deficient mice displayed reduced hematopoietic stem cell markers on bone marrow cells, and dysregulation of the Lin28 pathway. Thus, LDB1 function is critical for ESC and EB development and becomes progressively more important during differentiation to erythroblasts.

developmental biology↗

bioRxiv2024

The membrane-associated ubiquitin ligase MARCHF8 promotes cancer immune evasion by degrading MHC class I proteins

The loss of major histocompatibility complex class I (MHC-I) molecules has been proposed as a mechanism by which cancer cells evade tumor-specific T cells in immune checkpoint inhibitor (ICI)-refractory patients. Nevertheless, the mechanism by which cancer cells downregulate MHC-I is poorly understood. We report here that membrane-associated RING-CH-type finger 8 (MARCHF8), upregulated by human papillomavirus (HPV), ubiquitinates and degrades MHC-I proteins in HPV-positive head and neck cancer (HPV+ HNC). MARCHF8 knockdown restores MHC-I levels on HPV+ HNC cells. We further reveal that Marchf8 knockout significantly suppresses tumor growth and increases the infiltration of natural killer (NK) and T cells in the tumor microenvironment (TME). Furthermore, Marchf8 knockout markedly increases crosstalk between the cytotoxic NK cells and CD8+ T cells with macrophages and enhances the tumor cell-killing activity of CD8+ T cells. CD8+ T cell depletion in mice abrogates Marchf8 knockout-driven tumor suppression and T cell infiltration. Interestingly, Marchf8 knockout, in combination with anti-PD-1 treatment, synergistically suppresses tumor growth in mice bearing ICI-refractory tumors. Taken together, our finding suggests that MARCHF8 could be a promising target for novel immunotherapy for HPV+ HNC patients. One Sentence SummaryTargeting MARCHF8 restores MHC-I proteins, induces antitumor CD8+ T cell activity, and suppresses the growth of ICI-refractory tumors.

cancer biology↗

bioRxiv2024

Generation of a biosafety mouse model infection for SARS-CoV-2 replicon delivery particles

Animal models are essential for understanding the pathogenesis of SARS-CoV-2 and for developing therapeutic strategies. Replicon delivery particles (RDPs) were a component of trans-complementary systems for SARS-CoV-2, which is a safe and convenient tool in researching SARS-CoV-2 in animal biosafety level-II laboratory (ABSL-2). Here, we constructed a mouse model that conditional expressing SARS-CoV-2 N on the background of the K18-hACE2 KI mice. The SARS-CoV-2 N with flanked loxP-stop-loxP sequence was under the CAG promoter, and this cassette was knocked into the Tiger locus of mouse by CRISPR-Cas9 (K18-hACE2-N KI). By mating K18-hACE2-N KI mice with Cre tool mice, the offspring can express SARS-CoV-2 N (Cre-N-hACE2 KI) systemically and in a tissue-specific manner. Cre-N-hACE2 KI exhibited susceptibility to the SARS-CoV-2 {Delta}N-GFP/HBiT infection. The viral loads in lung exhibited a mountain-like trend, peaking at 4 days post-infection, and lung injuries can be observed. Overall, we demonstrated a mouse model infection for SARS-CoV-2 {Delta}N-GFP/HBiT to understand SARS-CoV-2 pathogenesis.

microbiology↗

bioRxiv2024

Pangenome Reveals Gene Content Variations and Structural Variants Contributing to Pig Characteristics

Pigs are among the most essential sources of high-quality protein in human diets. Structural variants (SVs) are a major source of genetic variants associated with diverse traits and evolutionary events. However, the current linear reference genome of pigs limits the presentation of position information for SVs. In this study, we generated a pangenome of pigs and a genome variation map of 599 deep-sequenced genomes across Eurasia. Moreover, a section-wide gene repertoire was constructed, which indicated that core genes were more evolutionarily conserved than variable genes. Subsequently, we identified 546,137 SVs, their enrichment regions, and relationships with genomic features and found significant divergence across Eurasian pigs. More importantly, the pangenome-detected SVs could complement heritability estimates and genome-wide association studies based only on single nucleotide polymorphisms. Among the SVs shaped by selection, we identified an insertion in the promoter region of the TBX19 gene, which may be related to the development, growth, and timidity traits of Asian pigs and may affect the gene expression. Our constructed pig pangenome and the identified SVs provide rich resources for future functional genomic research on pigs.

genomics↗

bioRxiv2024

Fate plasticity of interneuron specification

The generation of neuronal subtypes in the mammalian central nervous system is driven by competing genetic programs. The medial ganglionic eminence (MGE) gives rise to two major cortical interneuron (cIN) populations, marked by Somatostatin (Sst) and Parvalbumin (Pvalb), which develop on different timelines. The extent to which external signals influence these identities remains poorly understood. Pvalb-positive cINs are particularly important for regulating cortical circuits through strong perisomatic inhibition, yet they have been difficult to model in vitro. Here we investigated the role of the environment in shaping and maintaining Pvalb cINs. We grafted mouse MGE progenitors into a variety of 2D and 3D co-culture models, including mouse and human cortical, MGE, and thalamic systems with dissociated cells, organoids, organotypic cultures, and conditioned media. Across models, we observed distinct proportions of Sst- and Pvalb-positive cIN descendants. Strikingly, grafting MGE progenitors into 3D human, but not mouse, corticogenesis models led to efficient, non-autonomous differentiation of Pvalb-positive cINs. This differentiation was characterized by upregulation of Pvalb maturation markers, downregulation of Sst-specific markers, and the formation of perineuronal nets. Furthermore, lineage-traced postmitotic Sst-positive cINs, when grafted onto human cortical models, also upregulated Pvalb expression. These results reveal an unexpected level of fate plasticity in MGE-derived cINs, demonstrating that their identities can be dynamically shaped by the surrounding environment.

neuroscience↗

bioRxiv2024

Mechanism of NPC1L1 mediated 27-hydroxycholesterol metabolisms in the occurrence and development of osteoporosis

Cholesterol metabolism is closely related to the occurrence and development of osteoporosis, but the exact mechanism remains unclear. Niemann-Pick C1-like 1 (NPC1L1) is one of the key cholesterol transporter proteins, however, there are few reports on the functions of NPC1L1 besides regulating cholesterol transport, let alone bone homeostasis. Our preliminary research indicated that NPC1L1 may play a negative regulatory role in osteogenic differentiation. In this study, using in vitro osteogenic differentiation experiment and mouse osteoporosis model, we showed here that NPC1L1 expression was downregulated during osteogenesis, and NPC1L1 knockdown significantly enhanced osteogenic differentiation ability of osteoblasts and delayed progress of osteoporosis. Mechanistically, through RNA sequencing, NPC1L1 was found regulate cholesterol metabolism rather than transportation. It increased 27-Hydroxycholesterol (27-OHC) level through activating 27-hydroxylase (Cyp27a1), resulting in 27-OHC accumulation in osteoblasts and inhibition of osteogenesis. Moreover, C/EBP was proved mediated NPC1L1 promotes production of 27-OHC by Cyp27a1. These findings reveal that NPC1L1 inhibits osteogenesis and promotes osteoporosis via regulate cholesterol metabolism. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/615783v1_ufig1.gif" ALT="Figure 1"> View larger version (59K): org.highwire.dtl.DTLVardef@11a62bforg.highwire.dtl.DTLVardef@148fb9aorg.highwire.dtl.DTLVardef@7e486eorg.highwire.dtl.DTLVardef@c53116_HPS_FORMAT_FIGEXP M_FIG NPC1L1 inhibits osteogenic differentiation and promotes the progression of OP through the C/EBP/Cyp27a1/27-OHC axis. C_FIG

molecular biology↗

bioRxiv2024

Combination of Imipridone ONC201 or ONC206 with Temozolomide and Radiotherapy in triple ITR therapy reduces intracranial tumor burden and prolongs survival in an orthotopic wild-type IDH GBM mouse model

Glioblastoma remains the most lethal common primary brain tumor in adults with limited therapeutic options. TIC10/ONC201, a first-in-class imipridone we discovered, achieved meaningful therapeutic effects in phase I/II trials in patients with diffuse gliomas harboring H3K27M mutations, and currently the drug is in randomized phase III testing (ACTION trial; NCT05580562). ONC201 targets mitochondrial protease ClpP to disrupt oxidative phosphorylation and trigger the integrated stress response (ISR), TRAIL/DR5, and tumor cell death. We hypothesized that ONC201 and its analogue ONC206 synergize with temozolomide (TMZ) and ionizing radiation (IR), standard-of-care glioblastoma therapies. ONC201 enhances TMZ or IR-induced apoptosis, and cytotoxicity. ClpP-silencing suppresses ONC201-induced cytotoxicity but not TMZ or RT. Both ONC201 and ONC206 reduce expression of TMZ-resistance mediator MGMT. Suppression of MGMT protein was observed in H3K27M-mutated DIPG cell lines following treatment with ONC201 or ONC206 with or without TMZ. Cytokine profiling indicates distinct ONC201 alterations relative to TMZ suggesting distinct anti-tumor immune mechanisms. Triple IR+TMZ+ONC201 (ITR) therapy prolongs median survival to 123 days with a tail on survival curve (3-of-7 mice alive beyond 200-days) in an orthotopic U251 GBM model versus ONC201 (44-days; p=0.000197), IR (63-days; p=0.0012), TMZ (78-days; p=0.0354), ONC201+IR (55-days; p=0.0004), ONC201+TMZ (80-days; p=0.0041) and IR+TMZ (103-days; p>0.05). By 231-days, the only surviving mice were in IRT group. Our results support investigation of ONC201/ONC206 in combination with TMZ and IR (ITR) in GBM or H3K27M mutated diffuse glioma therapy.

cancer biology↗

bioRxiv2024

Precision-dependent modulation of social attention

Social attention, guided by cues like gaze direction, is crucial for effective social interactions. However, how dynamic environmental context modulates this process remains unclear. Integrating a hierarchical Bayesian model with fMRI, this study investigated how individuals adjusted attention based on the predictions about cue validity (CV). Thirty-three participants performed a modified Posner location-cueing task with varying CV. Behaviorally, individuals allocation of social attention was finely tuned to the precision (inverse variance) of CV predictions, with the predictions being updated by precision-weighted prediction errors (PEs) about the occurrence of target locations. Neuroimaging results revealed that the interaction between allocation of social attention and CV influenced activity in regions involved in spatial attention and/or social perception, such as the temporoparietal junction (TPJ), frontal eye field (FEF), superior temporal sulcus (STS), and inferior parietal sulcus (IPS). Precision-weighted PEs about target locations specifically modulated activity in the TPJ, STS, and primary visual cortex (V1), underscoring their roles in refining attentional predictions. Dynamic causal modeling (DCM) further demonstrated that enhanced absolute precision-weighted PEs about target locations strengthened the effective connectivity from V1 and STS to TPJ, emphasizing their roles in conveying residual error signals from low-level sensory areas to high-level critical attention areas. These findings elucidated how the precision of contextual predictions dynamically modulated social attention, offering insights into the computational and neurocognitive mechanisms of context-dependent social attention.

neuroscience↗

bioRxiv2024

Engineering a Cell-Based Orthogonal Ubiquitin Transfer Cascade for Profiling the Substrates of RBR E3 Parkin

The E3 ubiquitin (UB) ligase Parkin utilizes a Ring-Between-Ring (RBR) domain to mediate the transfer of UB to its substrates to regulate diverse cellular functions, including mitochondrial quality control, cell cycle progression, metabolism programming, and the establishment of synaptic functions. Mutations affecting the E3 ligase activity of Parkin are associated with cancer and Parkinsons disease (PD). An essential role of Parkin is to synthesize UB chains on the surface of damaged mitochondria to initiate mitophagy. Still, it is not clear how Parkin carries out other biological functions through the ubiquitination of its downstream targets in the cell. We hypothesized that a comprehensive substrate profile of Parkin would facilitate the discovery of ubiquitination pathways underpinning its multifaceted roles in cell regulation and reveal mechanistic linkages between Parkin malfunction and disease development. Here, we used phage display to assemble an orthogonal ubiquitin transfer (OUT) cascade of Parkin that can exclusively deliver an engineered UB mutant (xUB) to Parkin and its substrates in living cells. We then generated a substrate profile of Parkin by purifying xUB-conjugated proteins from cells and identifying them by proteomics. The OUT screen identified Parkin substrates involved in DNA replication, protein translation, intracellular protein transport, and rhythmic regulation. Based on previous literature implicating alterations in membrane vesicle trafficking in PD, we verified Parkin-catalyzed ubiquitination of Rab GTPases (Rab1a, Rab5a, Rab5c, Rab7a, Rab8a, Rab10, an Rab13) as well as CDK5, with reconstituted ubiquitination reactions in vitro and in cells. We also found chemical-induced stimulation of mitophagy enhanced Parkin-mediated ubiquitination of Rab proteins. These findings demonstrate that the OUT cascade of Parkin can serve as an empowering tool for identifying Parkin substrates to elucidate its cellular functions.

biochemistry↗

bioRxiv2024

Dysregulated Airway Host Defense in Hyper IgE Syndrome due to STAT3 Mutations

RationaleHyper IgE syndrome (STAT3-HIES), also known as Jobs syndrome, is a rare immunodeficiency disease typically caused by dominant-negative STAT3 mutations. STAT3-HIES syndrome is characterized by chronic pulmonary infection and inflammation, suggesting impairment of pulmonary innate host defense. ObjectivesTo identify airway epithelial host defense defects consequent to STAT3 mutations that, in addition to reported mutant STAT3 immunologic abnormalities, produce pulmonary infection. MethodsSTAT3-HIES sputum was evaluated for biochemical/biophysical properties. STAT3-HIES excised lungs were harvested for histology; bronchial brush samples were collected for RNA sequencing and in vitro culture. A STAT3-HIES-specific mutation (R382W), expressed by lentiviruses, and a STAT3 knockout, generated by CRISPR/Cas9, were maintained in normal human bronchial epithelia under basal or inflammatory (IL1{beta}) conditions. Effects of STAT3 deficiency on transcriptomics, and epithelial ion channel, secretory, antimicrobial, and ciliary functions were assessed. Measurements and Main ResultsMucus concentrations and viscoelasticity were increased in STAT3-HIES sputum. STAT3-HIES excised lungs exhibited mucus obstruction and elevated IL1{beta} expression. STAT3 deficiency impaired CFTR-dependent fluid and mucin secretion, inhibited expression of antimicrobial peptides, cytokines, and chemokines, and acidified airway surface liquid at baseline and post-IL1{beta} exposure in vitro. Notably, mutant STAT3 suppressed IL1R1 expression. STAT3 mutations also inhibited ciliogenesis in vivo and impaired mucociliary transport in vitro, a process mediated via HES6 suppression. Administration of a {gamma}-secretase inhibitor increased HES6 expression and improved ciliogenesis in STAT3 R382W mutant cells. ConclusionsSTAT3 dysfunction leads to multi-component defects in airway epithelial innate defense, which, in conjunction with STAT3-HIES immune deficiency, contributes to chronic pulmonary infection.

cell biology↗

bioRxiv2024

Synergistic combination therapy with ONC201 or ONC206, Enzalutamide and Darolutamide in preclinical studies of castration-resistant prostate cancer

Androgen receptor (AR) signaling plays a primary role in prostate cancer progression. Non-steroidal anti- androgens (NSAA) including enzalutamide, and apalutamide have been used to treat patients with advanced disease. However, patients with metastatic castration-resistant prostate cancer (mCPRC) develop resistance, resulting in limited overall survival benefit. Darolutamide is a novel next-generation androgen receptor- signaling inhibitor that is FDA approved for non-metastatic castration resistant prostate cancer (nmCRPC). Imipridone ONC201/TIC10 is first-in-class small molecule that activates the integrated stress response (ISR) and upregulates TNF-related apoptosis-inducing ligand (TRAIL). Our study investigates ISR and AR signaling in anti-tumor efficacy with ONC201 and enzalutamide or darolutamide against mCRPC cells. mCRPC cell lines 22RV1, LNCaP, DU145 and PC3 were treated with ONC201, darolutamide, and enzalutamide as single agents or in combinations. Combinations of ONC201 and darolutamide or enzalutamide demonstrated synergistic effects in mCRPC cells. Combinations of ONC201 and darolutamide or enzalutamide reduced PSA levels in LNCaP cells and induced of ATF4 in both LNCaP and 22RV1 cell lines. Darolutamide synergized with ONC201 regardless of AR status or castration sensitivity in vitro. Flow cytometric analysis showed increased intra-tumoral NK cells in mice treated with ONC201 and combination of ONC201 and darolutamide. Trends of increased TRAIL activation within NK cells were also observed in treatment groups. ONC201 and darolutamide demonstrated anti-tumor effects in vivo in the 22RV1 CRPC model. Our results prompt further translational and clinical studies with imipridones ONC201 or ONC201 in combination with enzalutamide or darolutamide for treatment of castrate resistant advanced or metastatic prostate cancer.

cancer biology↗

bioRxiv2024

MYC and p53 alterations cooperate through VEGF signaling to repress cytotoxic T cell and immunotherapy responses in prostate cancer

Patients with castration-resistant prostate cancer (CRPC) are generally unresponsive to tumor targeted and immunotherapies. Whether genetic alterations acquired during the evolution of CRPC impact immune and immunotherapy responses is largely unknown. Using our innovative electroporation-based mouse models, we generated distinct genetic subtypes of CRPC found in patients and uncovered unique immune microenvironments. Specifically, mouse and human prostate tumors with MYC amplification and p53 disruption had weak cytotoxic lymphocyte infiltration and an overall dismal prognosis. MYC and p53 cooperated to induce tumor intrinsic secretion of VEGF, which by signaling through VEGFR2 expressed on CD8+ T cells, could directly inhibit T cell activity. Targeting VEGF-VEGFR2 signaling in vivo led to CD8+ T cell-mediated tumor and metastasis growth suppression and significantly increased overall survival in MYC and p53 altered CPRC. VEGFR2 blockade also led to induction of PD-L1, and in combination with PD-L1 immune checkpoint blockade produced anti-tumor efficacy in multiple preclinical CRPC mouse models. Thus, our results identify a genetic mechanism of immune suppression through VEGF signaling in prostate cancer that can be targeted to reactivate immune and immunotherapy responses in an aggressive subtype of CRPC. SignificanceThough immune checkpoint blockade (ICB) therapies can achieve curative responses in many treatment-refractory cancers, they have limited efficacy in CRPC. Here we identify a genetic mechanism by which VEGF contributes to T cell suppression, and demonstrate that VEGFR2 blockade can potentiate the effects of PD-L1 ICB to immunologically treat CRPC.

cancer biology↗

bioRxiv2024

Synchronous assembly of peptide anisosome

Biomolecular condensates, formed by liquid-liquid phase separation (LLPS) of proteins or the complex of protein and nucleic acids, play key roles in regulating physiological events in biological system. However, the formation of mono-component yet inhomogeneous condensates is limited, and the underlying mechanism remains unclear. Here, we report the symmetrical core-shell structural biomolecular condensates formed through the LLPS by programming a tetra-peptide library. Mechanistic studies reveal that the tryptophan (W) is critical for the formation of core-shell structure because of its stronger homotypical {pi}-{pi} interaction compared with other amino acids, which endow us to modulate the droplets from core-shell to homogeneous structures by encoding the amino acid composition. Using molecular dynamics (MD) simulation and molecular engineering, we find that the inner core of LLPS is composed of dynamic and reversible fibers surrounded by liquid-like shells, resulting in a stable core-shell LLPS. Furthermore, we could control the multiphasic droplet formation by an intrinsic redox reaction or post-translational modification of peptide through phosphorylation, which facilitates the rational design of synthetic LLPS with various applications on demand.

bioengineering↗

bioRxiv2024

Activation of NLRP3 inflammasome/pyroptosis by protease inhibitor atazanavir at high concentrations is mediated through mitochondrial dysfunction

Protease inhibitor (PI)-based antiretroviral therapy increases CD4 and CD8 cell counts through anti-apoptosis mechanism. However, there are emerging reports that PIs could have pro-apoptotic effects; thus, PIs may have biphasic effect on apoptosis. We hypothesized that PI-induced apoptosis may be mediated through PI-induced mitochondrial dysfunction resulting in increased production of reactive oxygen species (ROS). To test this hypothesis, we used human T lymphoblastoid cell line (CEM) cultured with increasing concentrations of atazanavir (ATV). We assessed mitochondrial function--i.e., cell growth, apoptosis, mitochondrial membrane potential ({Delta}{Psi}), ROS, and electron transport chain (ETC) proteins. Apoptosis pathway genes were interrogated using the Human Apoptosis RT{superscript 2} Profiler PCR Array kit (QIAGEN), followed by quantitative PCR for validation. CEM cells treated with 15, 22.5, and 30 {micro}M of ATV resulted in significant reduction in cell growth and increased apoptosis. Further, high concentrations of ATV resulted in decreased mitochondrial {Delta}{Psi}, increased ROS production, and decreased protein expression of ETC I, II, III, and V. The following apoptosis pathway genes--caspase-1, BCL2A1, TP73 and TNFRSF1B--were differentially expressed. Caspase-1 is known to play a role in inducing NLRP3 inflammasome/pyroptosis pathway. We validated this with qPCR of the genes in the NLRP3 inflammasome pathway. Of note, NLRP3 inhibitor MCC950 and caspase-1 inhibitor YVAD reversed mitochondrial dysfunction and cell death. Our findings suggest that ATV-induced cell death is through the NLRP3 inflammasome/pyroptosis pathway. ATV-induced mitochondrial dysfunction plays an important role in the regulation of ATV-activated NLRP3 inflammasome pathway.

pharmacology and toxicology↗