Biology subjects

Lee, S.-Y.

Publications and source records attributed to Lee, S.-Y..

At least 19 recordsLinked to original sources

bioRxiv2026

Molecularly defined olfactory bulb output neurons exhibit selective yet overlapping projections

Early sensory processing decomposes rich signals into elemental features that can be conveyed through parallel neural pathways defined by cell types, with unique local circuit organization and long-range projection targets. Outputs from the olfactory bulb are typically categorized into two broad pathways corresponding to mitral and tufted cells. Previous high-throughput molecular profiling identified finer classes, but their morphology, projection patterns, and downstream convergence remain unclear. Here, we use an intersectional genetic targeting method with two Cre driver lines, retrograde projection mapping from six cortical targets, and RiboTag-based sequencing to study the subtypes of olfactory bulb output neurons. We found that retrograde labeling from the same cortical area recruited morphologically different OB neurons depending on the Cre driver used, suggesting a cortical area can be targeted by several subtypes. Comparing the marker expression, morphology, and downstream targets, we found that M1 neurons are densely labelled from the cortical amygdala, whereas M2 neurons project widely throughout the olfactory cortex but can be selectively recruited from the posterior piriform cortex. T1 cells differ from external tufted subtypes in their projections to the cap of the olfactory tubercle, while ET2 cells preferentially target the anterior olfactory nucleus pars externa. Electrophysiology of labelled neurons with different morphology and projection patterns differ in their intrinsic and functional properties. Further, our data suggest that extracellular recording methods may underrepresent mitral cells. Together, these findings reveal that molecularly defined olfactory bulb output neuron subtypes form anatomically and physiologically distinct channels that route information to overlapping yet selective downstream cortical targets.

neuroscience↗

bioRxiv2026

Non3nExonFinder: transcript-aware identification of shared non-triplet coding exons

Selection of an appropriate coding exon is an important upstream step in exon deletion-based loss-of-function strategies, particularly for genes with multiple transcript isoforms. However, manual comparison of exon structures across multiple transcripts can be time-consuming and may overlook transcript-specific differences in exon boundaries and coding sequence (CDS) composition. Non3nExonFinder was developed as an interactive R/Shiny-based tool for identifying shared non-triplet coding exons across user-defined human or mouse NCBI RefSeq protein-coding transcripts. Non3nExonFinder evaluates exon/CDS annotations and coding contributions for each transcript, identifies common exons with identical genomic boundaries, and prioritizes candidates based on CDS-only status and coding-length divisibility by three. Non-triplet CDS-only exons that are internal in all selected transcripts are classified as high-confidence candidates. Independent reconstruction from the original RefSeq GTF files across 80 genes (40 human and 40 mouse), comprising 6,848 transcript-specific exon instances, showed complete concordance with Non3nExonFinder outputs, with exact recovery of all candidate sets. Non3nExonFinder provides a reproducible upstream framework for transcript-aware prioritization of exon-deletion targets.

bioinformatics↗

bioRxiv2026

MitoAtlas: a Domain-Resolved Spatial Map of the HumanMitochondrial Proteome

Mitochondrial function depends on the precise sub-organellar localization and topological orientation of its proteome. However, mapping this architecture with domain-level precision across the double-membrane system has remained challenging. Here, we present MitoAtlas, a spatial systems biology resource built on SR-PL (super-resolution proximity labeling) that resolves the sub-mitochondrial architecture of 861 proteins at domain-level resolution. By deploying 42 spatially restricted APEX2 and BioID/BioID2 baits across mitochondrial compartments and cytosol in HEK293T-REx cells, we mapped 13,440 unique proximity-labeled sites. Integrating high-density spatial data with a machine-learning classification pipeline and AlphaFold structural modeling, we resolved membrane topologies for 196 transmembrane proteins and assigned sub-mitochondrial localizations to 160 previously unannotated mitochondrial proteins absent from MitoCarta3.0, including MINPP1 as an intermembrane space resident. Coupling bait-specific proximity labeling signatures with AlphaFold structure prediction and metabolite docking identified 47 protein-protein interactions, including the LETM1-Citrate Synthase complex, and 155 protein-metabolite interactions. MitoAtlas is available at mitoatlas.org as a domain-resolution spatial map of mitochondrial architecture and metabolism.

cell biology↗

bioRxiv2026

Virus-like particle delivery enables orthogonal genome editing in vitro and in vivo

The advent of CRISPR-Cas systems has revolutionized multiple fields, including basic science, biotechnology, and medicine. Central to this versatility is the use of programmable guide RNAs (gRNAs), which enable flexible and specific gene targeting. Building on this principle, various CRISPR-associated tools have been developed, including Cas9, base editors, prime editors, and gene-regulation platforms. However, because most CRISPR modalities share a common gRNA platform, the simultaneous use of multiple tools is constrained by interactions among different gRNAs, limiting orthogonal genome editing. This study presents a method for orthogonal multiplexed gene editing by packaging distinct CRISPR effectors and their corresponding gRNAs into separate virus-like particles (VLPs), thereby virtually eliminating the gRNA crosstalk. Furthermore, we demonstrate that this VLP-based strategy enables orthogonal, multiplexed gene editing in vivo in mouse eyes and ears. This platform expands the CRISPR toolkit by enabling simultaneous, non-interfering genetic manipulations in both cultured cells and living organisms.

genetics↗

bioRxiv2025

Early blood-spinal cord barrier stabilization preserves peri-lesional tissue and improves recovery after SCI

Vascular disruption is an early and critical event in spinal cord injury (SCI), but its contribution remains poorly understood. Using novel in vivo two-photon dual-dye imaging, we found simultaneous blood-spinal cord barrier (BSCB) leakage and venous dilation in both the injury epicenter and adjacent transitional segment after cervical SCI in rats. Notably, vascular permeability in the transitional zone preceded axonal and neuronal loss, revealing a therapeutic window. Systemic delivery of ferulic acid-glycol chitosan (FA-GC) nanoparticles, a membrane-sealant, rapidly stabilized the compromised vasculature, reduced neuronal loss in the transitional region, and improved forelimb muscle strength. These findings identified acute vascular leakage beyond the injury epicenter as a driver of secondary pathology and highlight early vascular stabilization as promising therapeutic strategy.

pathology↗

bioRxiv2025

Characterisation of LAMP1- and LAMP2A-positive organelles in neurons

LAMP1 and LAMP2A are abundant proteins of late endosomal/lysosomal compartments, which are often used interchangeably to label what is thought to be the same pool of organelles, potentially obscuring their unique physiological roles. Here, we characterised the transport dynamics of LAMP1- and LAMP2A-positive compartments in human iPSC-derived cortical neurons. We found that axonal LAMP1-positive organelles move more slowly in the retrograde direction, pause more frequently, and show a broader velocity distribution in the anterograde direction than LAMP2A-positive vesicles, suggesting they are distinct compartments with differential trafficking behaviour. To explore the molecular mechanism underlying these differences, we characterised with high spatiotemporal precision, the protein interactomes of LAMP1 and LAMP2A-positive compartments through proximity labelling, using full-length LAMP1 or LAMP2A fused to the light-activated biotin ligase LOV-Turbo. We identified and validated the endosomal protein, ZFYVE16, as a novel member of LAMP1 and LAMP2A interactomes. We suggest that LAMP2A-positive organelles represent a subset of LAMP1-positive compartments, which are surprisingly enriched in synaptic vesicle proteins. Summary statementLAMP1- and LAMP2A-positive organelles have different axonal transport dynamics and form distinct organelle pools characterised by specific protein compositions.

neuroscience↗

bioRxiv2025

Structural Basis of Cold and Menthol Sensing by TRPM8

The transient receptor potential melastatin member 8 (TRPM8) is a polymodal ion channel that senses cold and menthol in mammals. Despite prior structural studies, the mechanisms by which cold and menthol activate TRPM8 remain unresolved. Here, we present cryo-EM structures representing the cold and menthol-dependent activation trajectories, combined with extensive functional analyses. We captured snapshots of cooling-dependent pore opening, which involves dramatic pore rearrangement, suggesting a mechanism for cold sensing. Moreover, menthol binds dynamically to induce channel activation, which may underlie menthol specificity for TRPM8. Finally, we show how TRPM8 integrates multiple modalities (cold and menthol) through overlapping but non-identical pathways, revealing the temperature-specific "cold spot". These findings enhance our understanding of the molecular basis of physically and chemically induced cool sensation in mammals.

biophysics↗

bioRxiv2025

Biomarker Discovery via Integrative Multi-omics for Children exposed to Humidifier Disinfectant

RationaleExposure to humidifier disinfectants has been linked to an array of pulmonary disorders and diminished lung functionality particularly reduced Forced Vital Capacity (FVC). ObjectivesThis investigation sought to identify diagnostic biomarkers for early detection of children at elevated risk of developing chronic respiratory conditions following such exposure. MethodsOur research employed a comprehensive multi-omics strategy analyzing 70 pediatric patients alongside 10 controls, seamlessly integrating clinical assessments with transcriptomics, methylomics, proteomics, and metabolomics data. The analytical framework utilized a sophisticated combination of Non-negative Matrix Factorization (NMF), Multi-Omics Factor Analysis (MOFA), and advanced machine learning algorithms. Measurements and Main ResultsNMF clustering uncovered distinctive protein expression patterns associated with integrin-mediated signaling pathways and immune response mechanisms. Complementarily, MOFA identified latent factors correlating with lung function metrics, highlighting critical molecular pathways involved in integrin cell surface interactions and lipid metabolism regulation. Machine learning-based analysis facilitated the development of a multi-marker panel-comprising IGHV2-70, LysoPC (16:0), and hexadecyl ferulate-which achieved 81.46% accuracy in identifying pulmonary dysfunction cohort. ConclusionsThese findings suggest that alterations in integrin-related signaling networks and dysregulation of lipid metabolism play pivotal roles in mediating the long-term pulmonary consequences of humidifier disinfectant exposure. The proposed multi-marker panel offers significant potential for enhanced risk stratification and timely therapeutic intervention. At a Glance CommentaryO_ST_ABSScientific Knowledge on the SubjectC_ST_ABSExtensive epidemiological evidence has established the causal relationship between humidifier disinfectant exposure and pulmonary dysfunction; however, clinically validated biomarkers for predicting chronic lung disease progression remain limited. Pediatric populations demonstrate unique pathophysiological mechanisms distinct from adults, highlighting the critical necessity for biomarker identification grounded in comprehensive molecular understanding. Despite advances in omics technologies, recent investigations have encountered significant obstacles in achieving deeper mechanistic insights, predominantly attributable to methodological constraints in harmonizing clinical phenotypes with high-dimensional molecular datasets. What This Study Adds to the FieldThis investigation elucidates the fundamental contributions of integrin-mediated signaling cascades and lipid metabolic networks to persistent pulmonary dysfunction following humidifier disinfectant exposure. Our analyses revealed coordinated regulation of integrin signaling pathways and immune response networks through NMF clustering, indicating dynamic temporal evolution of inflammatory responses during chronic disease progression, with temporally distinct molecular signatures identified across discrete observation intervals. Multi-omics factor analysis (MOFA) corroborated integrin pathway dysregulation while additionally uncovering systematic suppression of lipid metabolic processes. Furthermore, machine learning algorithms enabled development of a robust three-component biomarker panel--encompassing IGHV2-70, LysoPC (16:0), and hexadecyl ferulate--demonstrating 81.46% classification accuracy for pulmonary dysfunction phenotypes. Collectively, these findings substantially advance mechanistic understanding of chronic lung injury in vulnerable pediatric cohorts and identify clinically relevant biomarkers with translational potential for risk stratification and therapeutic targeting in clinical practice.

systems biology↗

bioRxiv2025

Empirical optimization of dual-sgRNA design for in vivo CRISPR/Cas9-mediated exon deletion in mice

CRISPR/Cas9 has transformed gene editing, enabling precise genetic modifications across species. However, existing sgRNA design prediction models based on in vitro data are difficult to generalize to in vivo contexts. In particular, approaches based on single-sgRNA design require additional filtering of in-frame mutations, which is inefficient in terms of both time and cost. In this study, we developed the first mammalian in vivo-trained prediction model to evaluate the efficiency of a dual-sgRNA-based exon deletion strategy. Using 230 editing outcomes of postnatal viable individuals, eight prediction models were constructed and evaluated based on generalized linear models and random forests. The final selected model, a Combined GLM, integrated the DeepSpCas9 score with k-mer sequence features, achieving an AUC of 0.759 (95% Confidence Interval: 0.697-0.821). Motif analysis revealed that CC sequences were associated with high efficiency and TT sequences were associated with low editing efficiency. This study demonstrates that integrating sequence-based features with existing design scores can improve sgRNA efficiency prediction in vivo. The proposed framework can be applied to the development of next-generation sgRNA design tools, with direct implications for gene therapy, effective animal model generation, and precision genome engineering. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/675005v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@12e6170org.highwire.dtl.DTLVardef@1ff2599org.highwire.dtl.DTLVardef@1fd61dborg.highwire.dtl.DTLVardef@23a426_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2025

Distributed control circuits across a brain-and-cord connectome

Just as genomes revolutionized molecular genetics, connectomes (maps of neurons and synapses) are transforming neuroscience. To date, the only species with complete connectomes are worms1-3 and sea squirts4 (103-104 synapses). By contrast, the fruit fly is more complex (108 synaptic connections), with a brain that supports learning and spatial memory5,6 and an intricate ventral nerve cord analogous to the vertebrate spinal cord7-11. Here we report the first densely reconstructed adult fly connectome that unites the brain and ventral nerve cord, and we leverage this resource to investigate principles of neural control. We show that effector neurons (motor neurons, endocrine cells and efferent neurons targeting the viscera) are primarily influenced by sensory neurons in the same body part, forming local feedback loops. These local loops are linked by long-range circuits involving ascending and descending neurons organized into behavior-centric modules. Single ascending and descending neurons are often positioned to influence the voluntary movements of multiple body parts, together with the endocrine cells or visceral organs that support those movements. Brain regions involved in learning and navigation supervise these circuits. These results reveal an architecture that is distributed, parallelized and embodied, reminiscent of distributed control architectures in engineered systems12,13.

neuroscience↗

bioRxiv2024

PAM-flexible adenine base editing rescues hearing loss in a humanized MPZL2 mouse model harboring an East Asian founder mutation

Hearing loss is one of the most prevalent sensory disorders, but no commercial biological treatments are currently available. Here, we identified an East Asia-specific founder mutation, the homozygous c.220C>T mutation in MPZL2, that contributes to a significant proportion of hereditary deafness cases in our cohort study. We found that the disease-causing mutation could be targetable by adenine base editors (ABEs) that enable A{middle dot}T-to-G{middle dot}C base corrections without DNA double-strand breaks. To demonstrate this, we developed a humanized mouse model (hMPZL2Q74X/Q74X) that recapitulates human MPZL2 deafness and leads to progressive hearing loss. A PAM-flexible ABE variant with reduced bystander and off-target effects (ABE8eWQ-SpRY:sgRNA3) was packaged in dual adeno-associated viruses (AAVs) and injected into the inner ear of hMPZL2Q74X/Q74X mice and effectively corrected the mutation. This treatment significantly restored hearing function, improved inner ear structural integrity, and reversed altered gene expression. Base editing may hold therapeutic potential for hereditary deafness, including most cases of MPZL2 deafness.

genetics↗

bioRxiv2024

Directed evolution of the multicopper oxidase laccase for cell surface proximity labeling and electron microscopy

Enzymes that oxidize aromatic substrates have shown utility in a range of cell-based technologies including live cell proximity labeling (PL) and electron microscopy (EM), but are associated with drawbacks such as the need for toxic H2O2. Here, we explore laccases as a novel enzyme class for PL and EM in mammalian cells. LaccID, generated via 11 rounds of directed evolution from an ancestral fungal laccase, catalyzes the one-electron oxidation of diverse aromatic substrates using O2 instead of toxic H2O2, and exhibits activity selective to the surface plasma membrane of both living and fixed cells. We show that LaccID can be used with mass spectrometry-based proteomics to map the changing surface composition of T cells that engage with tumor cells via antigen-specific T cell receptors. In addition, we use LaccID as a genetically-encodable tag for EM visualization of cell surface features in mammalian cell culture and in the fly brain. Our study paves the way for future cell-based applications of LaccID.

biochemistry↗

bioRxiv2024

Molecular basis of the urate transporter URAT1 inhibition by gout drugs

Hyperuricemia is a condition when uric acid, a waste product of purine metabolism, accumulates in the blood1. Untreated hyperuricemia can lead to crystal formation of monosodium urate in the joints, causing a painful inflammatory disease known as gout. These conditions are associated with many other diseases and affect a significant and increasing proportion of the population2-4. The human urate transporter 1 (URAT1) is responsible for the reabsorption of [~]90% of uric acid in the kidneys back into the blood, making it a primary target for treating hyperuricemia and gout5. Despite decades of research and development, clinically available URAT1 inhibitors have limitations because the molecular basis of URAT1 inhibition by gout drugs remains unknown5. Here we present cryo-electron microscopy structures of URAT1 alone and in complex with three clinically relevant inhibitors: benzbromarone, lesinurad, and the novel compound TD-3. Together with functional experiments and molecular dynamics simulations, we reveal that these inhibitors bind selectively to URAT1 in inward-open states. Furthermore, we discover differences in the inhibitor dependent URAT1 conformations as well as interaction networks, which contribute to drug specificity. Our findings illuminate a general theme for URAT1 inhibition, paving the way for the design of next-generation URAT1 inhibitors in the treatment of gout and hyperuricemia.

biophysics↗

bioRxiv2024

Miniature linear and split-belt treadmills reveal mechanisms of adaptive motor control in walking Drosophila

To navigate complex environments, walking animals must detect and overcome unexpected perturbations. One technical challenge when investigating adaptive locomotion is measuring behavioral responses to precise perturbations during naturalistic walking; another is that manipulating neural activity in sensorimotor circuits often reduces spontaneous locomotion. To overcome these obstacles, we introduce miniature treadmill systems for coercing locomotion and tracking 3D kinematics of walking Drosophila. By systematically comparing walking in three experimental setups, we show that flies compelled to walk on the linear treadmill have similar stepping kinematics to freely walking flies, while kinematics of tethered walking flies are subtly different. Genetically silencing mechanosensory neurons alters step kinematics of flies walking on the linear treadmill across all speeds, while inter-leg coordination remains intact. We also found that flies can maintain a forward heading on a split-belt treadmill by adapting the step distance of their middle legs. Overall, these new insights demonstrate the utility of miniature treadmills for studying insect locomotion.

neuroscience↗

bioRxiv2024

Connexin43 in mesenchymal lineage cells regulates body adiposity and energy metabolism in mice

Connexin43 (Cx43) is the most abundant gap junction protein present in the mesenchymal lineage. In mature adipocytes, Cx43 mediates white adipose tissue (WAT) "beiging" in response to cold exposure and maintains the mitochondrial integrity of brown adipose tissue (BAT). We found that genetic deletion of Gja1 (Cx43 gene) in cells that give rise to chondro-osteogenic and adipogenic precursors driven by the Dermo1/Twist2 promoter leads to lower body adiposity and partial protection against the weight gain and metabolic syndrome induced by a high fat diet (HFD) in both sexes. These protective effects from obesogenic diet are related to increased locomotion, fuel utilization, energy expenditure, non-shivering thermogenesis, and better glucose tolerance in conditionally Gja1 ablated mice. Accordingly, Gja1 mutant mice exhibit reduced adipocyte hypertrophy, partially preserved insulin sensitivity, increased BAT lipolysis and decreased whitening under HFD. This metabolic phenotype is not reproduced with more restricted Gja1 ablation in differentiated adipocytes, suggesting that Cx43 has a hitherto unknown function in adipocyte progenitors or other targeted cells, resulting in restrained energy expenditures and fat accumulation. These results disclose an hitherto unknown action of Cx43 in adiposity, and offer a promising new pharmacologic target for improving metabolic balance in diabetes and obesity. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=157 HEIGHT=200 SRC="FIGDIR/small/574415v1_ufig1.gif" ALT="Figure 1"> View larger version (72K): org.highwire.dtl.DTLVardef@1ade7d3org.highwire.dtl.DTLVardef@4edbd7org.highwire.dtl.DTLVardef@1431c1borg.highwire.dtl.DTLVardef@ea6bbb_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2024

E protein control of NKgammadeltaT cell development through both generation and function of the stereotypic Vgamma1Vdelta6.3 TCR

T cell receptor (TCR) signals regulate important developmental transitions through induction of the E protein antagonist, Id3; however, Id3-deficiency produces paradoxical effects on {gamma}{delta} T cell subsets. Here, we show here that Id3-deficiency attenuates the development of V{gamma}3-expressing {gamma}{delta} T cells, while markedly enhancing the development of V{gamma}1V{delta}6.3-expressing NK{gamma}{delta}T cells. Id3-deficiency does so by regulating both the generation of the stereotypic V{gamma}1V{delta}6.3 TCR expressed by NK{gamma}{delta}T cells and its capacity to support development. Indeed, we determined that the Trav15 segment, which encodes the V{delta}6.3 TCR subunit, is directly bound by E proteins that control its expression. Moreover, once expressed, the resulting V{gamma}1V{delta}6.3 TCR in capable of specifying the innate-like NK{gamma}{delta}T cell fate in a cell-autonomous and developmentally-unrestricted manner that is restrained by the Id3/E axis. Together, these data indicate that the paradoxical behavior of NK{gamma}{delta}T cells in the Id3-deficient setting is entirely determined by its stereotypic V{gamma}1V{delta}6.3 TCR complex.

immunology↗

bioRxiv2023

Spatial dissection of cellular heterogeneity and functional crosstalk in the tumor microenvironments of gastric cancers

Spatial transcriptomics offers unprecedented challenges in characterizing tumor ecosystems, encompassing tumor cells and their surrounding tumor microenvironments (TME) along with cellular interactions therein. Here, we conducted Visium-based spatial transcriptomics on nine primary gastric cancers (GC) to translate the spatial organization of GC ecosystems into the functional landscape of cellular crosstalk across malignant, stromal and immune cells. We identified three distinct GC subtypes of 3 immunogenic, 3 epithelial and 3 stromal GCs with elevated infiltration of immune, malignant cells and fibroblasts reflecting the heterogeneity of the cell types and their interactions in GC TME. GC spatial architecture was further delineated into 6 regional compartments with varying degree of TME infiltration, among which the fibroblast-enriched TME was associated with the upregulation of epithelial-to-mesenchymal transformation (EMT) and immune response in malignant and immune cells, respectively. Cell type-specific transcriptional dynamics representing cellular crosstalk between TME cells were further identified, e.g., the infiltration of malignant and endothelial cells promote the cellular proliferations of TME cells whereas the fibroblasts and immune cells are associated with pro-cancer and anti-cancer immunity, respectively. Ligand-receptor analysis further revealed that CCL2-expressing fibroblasts promote the pro-cancer signaling of immune cells including JAK-STAT3 signaling and inflammatory response in tumor-infiltration macrophages. The CCL2+fibroblasts and macrophages with activated JAK-STAT3 signals are co-localized in spatial GC architecture and their co-abundance was associated with unfavorable clinical outcomes. Taken together, GC spatial transcriptomes can be regionally and functionally delineated into functional cellular crosstalk involving multiple cell types with potential clinical relevance, particularly the interaction between CCL2+ fibroblasts and JAK-STAT3+macrophages contributing to cancer-favoring immune contexture in GC TME.

cancer biology↗

bioRxiv2023

Structural insights into the FtsEX-EnvC complex regulation on septal peptidoglycan hydrolysis in Vibrio cholerae

During bacterial cell division, hydrolysis of septal peptidoglycan (sPG) is crucial for cell separation. This sPG hydrolysis is performed by the enzyme amidases whose activity is regulated by the integral membrane protein complex FtsEX-EnvC. FtsEX is an ATP-binding cassette transporter and EnvC is a long coiled-coiled protein that interacts with and activates the amidases. The molecular mechanism by which the FtsEX-EnvC complex activates amidases remains largely unclear. We present the cryo-electron microscopy structure of the FtsEX-EnvC complex from the pathogenic bacteria V. cholerae (FtsEX-EnvCVC). FtsEX-EnvCVC in the presence of ADP adopts a distinct conformation where EnvC is "horizontally extended" rather than "vertically extended". Subsequent structural studies suggest that EnvC can swing between these conformations in space in a nucleotide-dependent manner. Our structural analysis and functional studies suggest that FtsEX-EnvCvc may employ spatial control of EnvC for amidase activation, suggesting species-specific diversity in the FtsEX-EnvC regulation on septal peptidoglycan hydrolysis.

biochemistry↗