Biology subjects

Cui, J.

Publications and source records attributed to Cui, J..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

Multimodal spatial transcriptomics determines repeat expansion, huntingtin aggregation, and selective cortical neuron loss in Huntington Disease

Huntingtons disease (HD) is caused by CAG expansion in HTT, yet how somatic repeat instability and huntingtin aggregation relate to selective cell loss in the human brain remains unclear. We have developed a multimodal spatial transcriptomics approach that enables defining transcriptional programs with subcellular resolution, somatic CAG repeat lengths, and six other pathology marks including huntingtin aggregates in every cell of intact brain sections. Imaging 428,173 cells in HD cortex revealed selective vulnerability: L5-6 NP and L6b deep-layer excitatory neurons undergo >50% loss, closely linked to very large (>380{+/-}55) somatic expansions. Intranuclear aggregation was most prevalent at intermediate somatic repeat expansion (220-300 CAGs) and was accompanied by broader transcriptional changes. In contrast, chandelier and somatostatin+ inhibitory interneurons are lost despite only modest repeat expansion or aggregation. These data provide a comprehensive resource and establish a broadly applicable framework for connecting repeat expansion and protein pathology across diverse cell types. Short Bullet pointsO_LIDevelopment of a novel, multimodal spatial transcriptomics platform enables definition of RNA transcriptomes with subcellular resolution, somatic repeat expansions, and protein accumulation in cells within tissue sections C_LIO_LIGeneration of two complementary datasets for HD and control cortex: 428,173 cells to quantify comprehensively cell-type vulnerability, and a deeper multimodal dataset in which CAG repeat expansion, huntingtin aggregation, six additional pathological readouts, and expression of 1,128 genes were measured in 185,721 cells, providing a comprehensive resource for the neurodegeneration community. C_LIO_LIDeep layer excitatory neuron loss (L5-6 NP and L6b) was associated with very large somatic CAG expansions (>380{+/-}55), while selective inhibitory neuron loss (chandelier and somatostatin interneurons) occurred with modest CAG repeat expansion or huntingtin intranuclear aggregation C_LIO_LIIntranuclear aggregation, not somatic repeat expansion, was more predictive of transcriptional changes, including chromatin remodeling and RNA export factors C_LI

neuroscience↗

bioRxiv2025

Mouse Cardiac Regeneration Database (MCAREDB): a single-cell atlas of neonatal cardiac regeneration in mice

BackgroundThe adult mammalian heart exhibits minimal regenerative capacity, rendering most major cardiac injuries irreversible. In contrast, neonatal mice retain the ability to regenerate cardiac tissue for a brief period after birth. Although single-cell RNA sequencing (scRNA-seq) has deepened our understanding of this regenerative window, existing datasets remain dispersed across studies, hindering integrative and comparative analyses. Methods and ResultsTo address this gap, we developed the Mouse CArdiac REgeneration DataBase (MCAREDB), a freely accessible, web-based resource that consolidates scRNA-seq datasets from multiple mouse cardiac regeneration studies. MCAREDB enables cell type-specific exploration of differential gene expression and pathway enrichment across key biological contexts, including neonatal versus postnatal stages and sham versus myocardial infarction surgeries. Core features include an interactive genome browser with epigenetic tracks, comparative gene expression visualization, pathway enrichment analysis, and bulk data download options. ConclusionBy integrating diverse datasets into a unified and interactive framework, MCAREDB lowers technical barriers to data exploration and accelerates hypothesis generation in cardiac regeneration research. The resource is freely available at https://MCAREDB.org:3305.

bioinformatics↗

bioRxiv2025

Linking MSMEG_1353 to lipid metabolism and envelope integrity in Mycolicibacterium smegmatis

Mycobacterium tuberculosis (Mtb) poses a significant global health burden. Rv0647c, an essential Mtb cell wall protein, is a potential drug target. We studied its Mycolicibacterium smegmatis (MSMEG) homologue, MSMEG_1353, using a CRISPRi conditional knockdown. MSMEG_1353 depletion increased cell width and volume, delayed log-phase initiation, slowed aggregation, reduced biofilm formation, and heightened susceptibility to antibiotics and sodium dodecyl sulfate (SDS). AlphaFold, sequence alignment, and UNIPROT analyses suggest MSMEG_1353 functions as a protein kinase, with conserved residues in intermediate high-confidence regions potentially forming an ATP-binding site. Investigating its role in cell envelope biosynthesis, mass spectrometry revealed elevated levels of mycolic acid biosynthesis proteins upon MSMEG_1353 knockdown. RT-qPCR confirmed upregulation of the fabD-acpM-kasA-KasB-accD6 operon, encoding key mycolic acid synthesis enzymes. Notably, a strong negative correlation with MSMEG_0911, the predominant isocitrate lyase, important in the glyoxylate cycle, was observed via both MS and RT-qPCR. Collectively, MSMEG_1353 deficiency compromises cell wall integrity, likely due to altered lipid composition resulting from dysregulated mycolic acid biosynthesis and lipid metabolism. These findings support the development of models explaining MSMEG_1353s involvement in these pathways.

microbiology↗

bioRxiv2025

Targeting Putrescine Homeostasis with Lactiplantibacillus plantarum PS128 to Mitigate Autism-like Behaviors in Fmr1 KO Mice

BackgroundAutism spectrum disorder (ASD) is biologically heterogeneous and has limited mechanism-informed interventions targeting core behavioral symptoms. Emerging evidence suggests that gut microbial metabolism shapes neurobehavioral outcomes, yet the specific metabolic pathways linking gut ecology to brain function remain incompletely understood. One candidate pathway is polyamine metabolism, which links microbial amino acid metabolism to host regulation and represents a plausible but underexplored contributor to ASD-related phenotypes. Notably, the psychobiotic Lactiplantibacillus plantarum PS128 has been reported to improve social behaviors in individuals with ASD, although the molecular basis for these effects is unclear. In this study, we used Fragile X mental retardation 1 knockout (Fmr1 KO) mice, a well-established ASD model, to investigate microbiota-dependent metabolic mechanisms underlying autism-like behaviors. ResultsWe found that autism-like behaviors in Fmr1 KO mice were associated with gut dysbiosis, impaired intestinal barrier integrity, disruption of the arginine-ornithine-polyamine pathway, and elevated putrescine in the prefrontal cortex (PFC). Supplementation with PS128 remodeled the gut microbiota, reduced inflammation- and disease-associated taxa, improved intestinal structure and permeability, and restored polyamine homeostasis. Targeted metabolomics revealed an increased PFC-to-serum putrescine ratio in KO mice, which was normalized following PS128 intervention. This correction was accompanied by reduced expression of polyamine transporters in the PFC, including ATP13A family members. Causal experiments supported a functional role for putrescine, as peripheral elevation of putrescine induced autism-like behaviors in wild-type mice, whereas pharmacological inhibition of putrescine synthesis ameliorated behavioral deficits in Fmr1 KO mice. ConclusionsThese findings identify putrescine metabolic dysregulation as a key contributor to autism-like phenotypes and support the existence of an ASD subtype defined by disruption of the arginine-ornithine-polyamine axis. Integrated multi-omics and causal perturbation analyses support a model in which microbiota-targeted intervention rebalances systemic polyamine regulation along the gut-brain axis, thereby improving ASD-relevant behaviors. Our work provides mechanistic evidence linking microbial metabolism to neurochemical homeostasis and highlights the translational potential of metabolic stratification in ASD. Graphical AbstractWe identify dysregulated microbiota-driven polyamine homeostasis as a causal contributor to autism-like behaviors in Fmr1 knockout mice. A psychobiotic intervention restores gut-brain metabolic balance and rescues social and repetitive behavioral deficits through coordinated microbial and host transport regulation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/694366v2_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@e1e780org.highwire.dtl.DTLVardef@125a3baorg.highwire.dtl.DTLVardef@201228org.highwire.dtl.DTLVardef@1880650_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Immune-Derived THBS1-CD47 Axis Induces Cellular Senescence and Suppresses Osteogenesis in Diabetic Periosteum

Type 2 diabetes mellitus (T2DM) weakens bone repair and increases fracture risk, but how the periosteal environment contributes to this problem is not fully understood. We found that diabetic mice showed thinning of the periosteum, reduced bone mass, and poor function of bone-forming cells. Using single-cell RNA sequencing, we discovered that immune cells and bone-lineage cells interact more strongly in diabetes through the THBS1-CD47 pathway. THBS1 was mainly produced by macrophages, while its receptor CD47 was abundant in osteogenic cells. Laboratory experiments showed that THBS1 blocked bone gene activity, increased inflammation and cell death, and damaged mitochondria, leading to higher oxidative stress. Knocking down CD47 reversed these effects, restoring bone cell activity and energy balance. In diabetic fracture models, blocking THBS1 improved callus formation and bone healing. These findings identify THBS1-CD47 as a key driver of periosteal dysfunction in T2DM and highlight it as a potential target to improve skeletal repair in diabetic patients.

cell biology↗

bioRxiv2025

Dynamic interactions in human amygdala subregions support hierarchical processing of fearful faces

Fearful-expression recognition is critical for adaptive responses to potential threats and relies on both rapid threat detection and fine-grained face encoding. Yet how human amygdala subregions differentially support these distinct cognitive components remains unclear. Here, we recorded intracranial EEG from lateral and medial amygdala in epilepsy patients performing an emotional face-matching task and combined multivariate decoding, time-frequency and directed-connectivity analyses with intracranial stimulation. The lateral amygdala exhibited early fear-specific responses, characterized by higher decoding accuracy and increased theta/alpha-band (4-12 Hz) power, and transmitted this fear-related information to the medial amygdala, which showed delayed and sustained activation. By contrast, the medial amygdala encoded face-specific information at later stages in the 2-16 Hz band with superior decoding accuracy and then relayed this information back to the lateral amygdala. Intracranial stimulation produced a double dissociation in behavior, with lateral amygdala stimulation disrupting fear detection, whereas medial amygdala stimulation selectively accelerated neutral-face recognition. Together, these findings reveal a temporal hierarchy in the human amygdala, whereby dynamic bidirectional interactions between subregions implement distinct components of fearful-expression recognition, providing a circuit-level framework for understanding social threat processing.

neuroscience↗

bioRxiv2025

Structural Basis for Iterative Methylation by a Cobalamin-dependent Radical S-Adenosylmethionine Enzyme in Cystobactamids Biosynthesis

Cystobactamids are non-ribosomal peptide natural products that function as DNA gyrase inhibitors, exhibiting significant antibacterial activity. They are isolated from Cystobacter sp. Cbv34 and contain various alkoxy groups on para-aminobenzoic acid moieties, which are believed to play a crucial role in antibacterial functions. The alkoxy groups are generated by iterative methylations on a methoxy group by the cobalamin (Cbl)-dependent radical S-adenosylmethionine (SAM) enzyme CysS. CysS catalyzes up to three methylations to give ethoxy, isopropoxy, sec-butoxy, and tert-butoxy groups. For each methylation, CysS uses a ping-pong mechanism in which two molecules of SAM are consumed. One SAM is used to methylate cob(I)alamin, while another generates a 5'-deoxyadenosyl 5'-radical to initiate substrate methylation. However, little is known about how the enzyme promotes both Cbl methylation and iterative substrate methylation, which occur by polar SN2 and radical processes, respectively. Here, we report three X-ray crystal structures of a homolog of CysS from Corallococcus sp. CA054B. Two were determined in the presence of methoxy- and ethoxy-containing substrates, showing how CysS accommodates substrates and products during iterative methylation. The third structure, determined in the absence of a substrate, exhibits structural changes that reorient the SAMs conformation to allow for the methylation of cob(I)alamin.

biochemistry↗

bioRxiv2025

Persistent trade-offs balance competition and colonization across centuries

Microbial competition drives rapid adaptation, often forcing organisms to specialize in new ecological niches. Adaptations that improve competitive ability can reduce performance in other environments creating trade-offs. Whether such trade-offs persist in nature--or are eroded as lineages adapt through compensatory changes--remains largely unknown. Here we show that a trade-off between competitive ability and host colonization has been stably maintained in natural Pseudomonas populations for centuries. Wild plant-pathogenic Pseudomonas compete using tailocins--phage-derived molecular weapons that bind to specific cell-surface receptors. Genomic surveys and functional assays reveal that the most broadly lethal tailocins remain rare--while the tailocins production increases competitive killing, it also compromises plant colonization. We determine that the polymorphisms behind this trade-off are not transient -- historical genomes spanning two centuries show that the trade-off has been maintained for at least 10-10 generations. Our results demonstrate that, in natural populations, a trade-off between competition and pathogenicity is fundamental and not easily overcome. SignificanceWhen a microbe colonizes a host, it must both establish infection and outcompete other organisms. Short-term experiments show that gains in competitive ability can reduce colonization, creating trade-offs, but whether microbes resolve these conflicts over long evolutionary timescales is unknown. We show that a trade-off between competitive killing and host colonization has been stably maintained for centuries in natural Pseudomonas populations infecting Arabidopsis thaliana. Tailocins--phage-derived weapons--provide strong competitive advantages, yet their production reduces colonization success, explaining why the most broadly lethal variants remain rare. Genomic surveys and historical genomes spanning two centuries reveal that the polymorphisms underlying this trade-off have persisted across 10-10 generations. Understanding such long-lived constraints can inform antimicrobial strategies that exploit evolutionary trade-offs.

evolutionary biology↗

bioRxiv2025

The mycobacterial ATP burst is a lysis artifact and serves as an assay for drug-induced cell wall damage

Antibiotics targeting the mycobacterial cell wall are a cornerstone of tuberculosis treatment. Understanding how these drugs facilitate bacterial killing has important implications for identifying novel therapeutic targets and optimizing drug combinations. Previous studies on mycobacteria using the BacTiter-Glo luminescence assay have reported increased ATP levels after treatment with cell wall inhibitors such as isoniazid, a phenomenon referred to as an ATP burst. This phenomenon has been proposed to contribute to drug-induced killing. In this study, we demonstrate that the ATP burst is not a biological response but rather an experimental artifact resulting from enhanced cell lysis induced by cell wall-targeting drugs. The addition of a bead-beating lysis step prior to luminescence measurement abolishes the ATP burst, enabling a more reliable assessment of intracellular ATP levels with this assay. Our findings thus establish a direct link between the ATP burst and the canonical mechanism of action of cell wall inhibitors. We further demonstrate the utility of this approach as a functional readout for identifying compounds that disrupt the mycobacterial cell wall and for screening synergistic or antagonistic interactions with known cell wall-targeting agents. Together, our results elucidate the mechanistic basis of the ATP burst and provide a valuable tool for antimycobacterial drug discovery and development.

microbiology↗

bioRxiv2025

Pan-cancer analysis reveals that tumor microbiomes generate specific epitopes through transcriptional reprogramming

The therapeutic promise of tumor microbial peptides faces a critical safety paradox: tumors and peritumor tissues harbor compositionally similar microbiomes, yet safe immunotherapy demands tumor-specific targeting. We performed comprehensive transcriptomic analysis of microbial communities from 1,868 tumor and peritumor samples across 11 cancer types. Although 88.5% of microbial species are present in both tissue types, tumor and peritumor tissues exhibit profound transcriptional divergence: only 34.1% of microbial genes are expressed in both tissue types, and merely 18.2% of predicted HLA-binding peptides overlap. This transcriptional divergence represents microbial transcriptional reprogramming (MTR) in tumor tissues, through which microbes activate distinct genomic regions while adapting to tumor microenvironmental pressures, thereby generating tumor-specific peptide repertoires. Using Microbial Transcriptional Reprogramming Index (MiTRI) to quantify MTR, we found that higher MiTRI values correlate with expanded immunogenic peptide repertoires. Mass spectrometry-based immunopeptidomics confirmed that 25% of naturally presented microbial peptides originate from MTR regions. Crucially, in microsatellite-stable colorectal cancer, neo-adjuvant therapy responders exhibited significantly higher MiTRI values. T cell receptors (TCRs) recognized MTR-derived epitopes at 2.10-fold higher rates than genome-wide epitopes, accounting for 79.1% of clonally expanded TCRs. These findings establish MTR as the principle that ensures the tumor-specificity and safety of micorbiome-directed immunotherapy.

cancer biology↗

bioRxiv2025

Neutrophil extracellular traps-mediated thrombosis drive pyrrolizidine alkaloid-induced hepatic sinusoidal obstruction syndrome

Hepatic sinusoidal obstruction syndrome (HSOS) caused by pyrrolizidine alkaloid (PA) is a life-threatening disease with limited treatments, yet the initiating vascular event remains undefined. Neutrophil extracellular traps (NETs), as key driver of immunothrombosis in various thromboinflammatory diseases, may represent a critical yet uncharacterized component in the pathogenesis of PA-HSOS. Rats and mice were separately administered monocrotaline (MCT) or senecionine (SEN) via gavage to induce HSOS. Human hepatic sinusoidal endothelial cell (HHSEC) and primary human neutrophil treated with MCT, SEN, or corresponding metabolites were used as in vitro models. In separate experiments, animals received a neutrophil elastase (NE) inhibitor or neutrophil-depleting antibody to evaluate the contribution of NETs. We detected NETs within the sinusoids of necrotic liver lobes, as evidenced by robust immunostaining for NETs markers, accompanied with elevated level of circulating cell-free DNA, and hepatic citrullinated histone H3. Sinusoidal microthrombi containing VWF, CD41, and fibrin(ogen) were also stained positive for neutrophils in PA-HSOS models. Treatment with NE inhibitor or neutrophil depletion markedly attenuated PA-induced liver injury by concomitantly suppressing NETs formation and intrahepatic thrombogenesis. In vitro, conditioned medium from HHSEC exposed to MCT or SEN metabolites potently induced NETs formation. HHSEC injured by PA metabolites released CXCL8. Furthermore, hepatic mRNA and protein levels of CXCL1 and CXCL2 (murine homologous to human CXCL8) were up-regulated in PA-HSOS models, accompanied by a parallel elevation of their high-affinity receptors, CXCR1 and CXCR2. Our study identifies NETs-mediated immunothrombosis as a key driver in HSOS, which provides a promising target for PA-HSOS treatment.

pharmacology and toxicology↗

bioRxiv2025

Constrained Diffusion for Protein Design with Hard Structural Constraints

AO_SCPLOWBSTRACTC_SCPLOWDiffusion models offer a powerful means of capturing the manifold of realistic protein structures, enabling rapid design for protein engineering tasks. However, existing approaches observe critical failure modes when precise constraints are necessary for functional design. To this end, we present a constrained diffusion framework for structure-guided protein design, ensuring strict adherence to functional requirements while maintaining precise stereochemical and geometric feasibility. The approach integrates proximal feasibility updates with ADMM decomposition into the generative process, scaling effectively to the complex constraint sets of this domain. We evaluate on challenging protein design tasks, including motif scaffolding and vacancy-constrained pocket design, while introducing a novel curated benchmark dataset for motif scaffolding in the PDZ domain. Our approach achieves state-of-the-art, providing perfect satisfaction of bonding and geometric constraints with no degradation in structural diversity.

molecular biology↗

bioRxiv2025

Water body size shapes body size evolution via subgenomic selection in allopolyploid fish

Allopolyploidization is a major force in species evolution, and studying allopolyploids in diverse environments provides insights into their genetic mechanisms. Here, we investigated how water body size variation influences body size in allopolyploid fish. We used a nascent allopolyploid lineage derived from goldfish (Carassius auratus) and common carp (Cyprinus carpio) and established two populations in small and large water bodies for two generations. Genomic, transcriptomic, and epigenomic analyses revealed that the larger population had higher genetic diversity in subgenome R, while the smaller population had higher diversity in subgenome C. Potential selection regions were associated with growth regulatory genes, suggesting a role for gene regulation in body size adaptation. Dynamic gene expression patterns in various tissues indicated potential mechanisms underlying muscle development and adaptation to smaller water bodies. Furthermore, selection regions were linked to changes in chromatin accessibility, with subgenome R being more susceptible to these changes during genetic divergence. Our findings demonstrate that water body size variation drives body size in allopolyploid fish through subgenomic selection, impacting genetic variation, gene regulation, and epigenetic mechanisms.

genomics↗

bioRxiv2025

Abnormal shear stress induces ferroptosis in endothelial cells via KLF6 downregulation

Stable laminar flow maintains vascular tone regulation, while abnormal blood flow, such as disturbed flow or extreme shear stress, causes endothelial dysfunction, but the underlying mechanism is yet to be explored. We used a microfluidic device to deform flat microchannel into tunnel-like macrochannel. The cross-sectional area of this macrochannel varies with the flow rate and the thickness of the deformable layer, creating three levels of shear stress: low (0.99 dyn/cm2, LSS), medium (4.78 dyn/cm2, MSS), and high (24 dyn/cm2, HSS). Comparing different shear stress exposure to endothelial cells for 24 h, prominent ferroptosis features emerged under either LSS or HSS compared to MSS. These features included increased C11 BODIPY-labeled lipid peroxidation and 4-hydroxynonenal accumulation, CoQ10 depletion, reduced SLC7A11 protein expression, and diminished cell death with Ferrostatin-1 treatment. RNA-seq analysis (LSS/MSS) showed that LSS significantly downregulated transcription of cholesterol homeostasis and unfolded protein response (UPR). Compared to MSS, Western blot results showed that both LSS and HSS reduced the expression of two key enzymes (MVD and IDI1) in the mevalonate pathway, as well as the expression of two main UPR signaling regulators (PERK and BiP). Based on the binding prediction between transcription factors and gene promoters from differentially expressed genes identified through RNA-seq, we found KLF6 to be a key transcription factor. It regulates the PERK-mediated UPR and the mevalonate pathway, which are associated with SLC7A11 expression and CoQ10 synthesis, respectively. The overexpression of KLF6 restored SLC7A11 and CoQ10 levels under both LSS and HSS, significantly reducing foam cell formation, monocyte adhesion, and lipid peroxidation. Our findings reveal KLF6 as a crucial regulator of atherosclerosis induced by abnormal shear stress.

cell biology↗

bioRxiv2025

Rapid elicitation of a new class of neutralizing N332-glycan independent V3-glycan antibodies against HIV-1 in nonhuman primates.

Sequential immunization is a promising approach to elicit broadly neutralizing antibodies (bNAbs) against the HIV-1 Envelope (Env). However, available protocols are inefficient and involve multiple immunizations over long periods of time. Here, we present WIN332, a new engineered Env-immunogen that induces a new class of neutralizing N332-glycan-independent antibodies to the conserved V3-glycan epitope of Env after a single bolus immunization in nonhuman primates. WIN332 binds to precursors of canonical human N332-glycan-dependent (Type-I) V3-glycan bNAbs but also of a first-of-its-class N332-glycan-independent (Type-II) V3-glycan bNAb. A single immunization elicits neutralizing serum and monoclonal antibodies that are boosted and affinity matured with a heterologous immunogen. EMPEM analysis of serum antibodies, antibody cloning and cryo-EM analysis reveal that WIN332 elicits N332-glycan-independent antibodies with remarkable sequence and binding similarities with the most potent human type-I and type-II V3-glycan bNAbs. Thus, WIN332 is a promising vaccine candidate to streamline V3-glycan bNAb elicitation.

immunology↗

bioRxiv2025

Targeting RUNX1 in Macrophages Facilitates Cardiac Recovery

Despite advances in disease treatment, our understanding of how damaged organs recover and the mechanisms governing this process remain poorly defined. Here, we mapped the transcriptional and regulatory landscape of human cardiac recovery using single cell multiomics. Macrophages emerged as the most reprogrammed cell type. Deep learning identified the transcription factor RUNX1 as a key regulator of this process. Macrophage-specific Runx1 deletion recapitulated the human cardiac recovery phenotype in a chronic heart failure model. Runx1 deletion reprogrammed macrophages to a reparative phenotype, reduced fibrosis, and promoted cardiomyocyte adaptation. RUNX1 chromatin profiling revealed a conserved regulon that diminished during recovery. Mechanistically, the epigenetic reader BRD4 controlled Runx1 expression in macrophages. Chromatin activity mapping, combined with CRISPR perturbations, identified the precise regulatory element governing Runx1 expression. Therapeutically, small molecule Runx1 inhibition was sufficient to promote cardiac recovery. Our findings uncover a druggable RUNX1 epigenetic mechanism that orchestrates recovery of heart function.

immunology↗

bioRxiv2025

Hydraulic Activation of the AsLOV2 photoreceptor

How proteins transduce light into mechanical energy remains a central question in biology. This study tests the hypothesis that blue light activation of the LOV2 (light, oxygen, voltage sensitive) domain of Avena sativa phototropin 1 (AsLOV2), gives rise to concerted water movement that induces protein conformational extensions. Using electron and nuclear magnetic resonance spectroscopy, along with molecular dynamics simulations at high pressure, we find AsLOV2 activation can be initiated by blue light or high pressure, followed by selective and concerted expulsion of low-entropy, tetrahedrally coordinated "wrap" water from the protein hydration shell. These findings suggest that interfacial water serves as constituents to reshape the proteins free energy landscape during activation. Our study highlights hydration water as an active hydraulic fluid that can drive long-range conformational changes underlying protein mechanics upon light activation and offers a new concept for engineering externally controllable protein actuators. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=70 SRC="FIGDIR/small/660617v2_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1a35f9dorg.highwire.dtl.DTLVardef@da4991org.highwire.dtl.DTLVardef@113e9b2org.highwire.dtl.DTLVardef@53d9fc_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2025

Mechanisms of KCNQ1 gating modulation by KCNE1/3 for cell-specific function

KCNQ1 potassium channels are essential for physiological processes such as cardiac rhythm and intestinal chloride secretion. KCNE-family subunits (KCNE1-5) associate with KCNQ1, conferring distinct properties across various tissues. KCNQ1 activation requires membrane depolarization and phosphatidylinositol 4,5-bisphosphate (PIP2) whose cellular levels are controlled by Gq-coupled GPCR activation. While modulation of KCNQ1s voltage-dependent activation by KCNE1/3 is well-characterized, their effects on PIP2-dependent gating of KCNQ1 via GPCR signaling remain less understood. Here we resolved structures of KCNQ1-KCNE1 and reassessed reported KCNQ1-KCNE3 structures with and without PIP2. We revealed that KCNQ1-KCNE1/3 complexes feature two PIP2-binding sites, with KCNE1/3 contributing to a previously overlooked, uncharacterized site involving residues critical for voltage sensor and pore domain coupling. Via this site, KCNE1 and KCNE3 distinctly modulate the PIP2-dependent gating, in addition to the voltage sensitivity, of KCNQ1. Consequently, KCNE3 converts KCNQ1 into a voltage-insensitive PIP2-gated channel governed by GPCR signaling to maintain ion homeostasis in non-excitable cells. KCNE1, by significantly enhancing KCNQ1s PIP2 affinity and resistance to GPCR regulation, forms predominantly voltage-gated channels with KCNQ1 for conducting the slow-delayed rectifier current in excitable cardiac cells. Our study highlights how KCNE1/3 modulates KCNQ1 gating in different cellular contexts, providing insights for tissue-specifically targeting multi-functional channels.

biochemistry↗