Biology subjects

Kolotuev, I.

Publications and source records attributed to Kolotuev, I..

13 recordsLinked to original sources

bioRxiv2026

Transient morphogenetic constraints organize self-assembling axon neighborhoods for robust yet flexible wiring

Nervous systems form wiring patterns that are reproducible across individuals. This reproducibility is thought to emerge from molecular encoding and developmental events, but their relative contributions remain unclear. We address this question in the C. elegans neuropil, where embryonic developmental dynamics and adult anatomy are resolved at single-cell resolution. We find that transient morphogenetic structures -- rosettes, corridor cells, pioneer axon scaffold -- restrict which axons make contact, shaping the neuropil into overlapping neighborhoods. This demonstrates how early events constrain wiring choices, but not whether they explain the resulting reproducibility. To explain, we use an agent-based model of stochastic innervation that recapitulates macro- and micro-level reproducibility, revealing a trade-off between physical constraint and molecular specificity that limits neighborhood size. Counterintuitively, less selective axons produce more reproducible wiring when constrained within neighborhoods. This trade-off lets nervous systems maximize reproducibility without having to molecularly encode every axon-contact, a strategy for robust yet flexible wiring.

neuroscience↗

bioRxiv2026

A stereotyped glial attachment determines the morphology and function of neuronal cilia

Primary cilia are signaling hubs that influence nearly every aspect of cell physiology. Neuronal primary cilia were recently discovered to interact with glia in the mammalian brain, but how cilia-glia attachments form and what roles they play remain unknown. Here, we find that two C. elegans sensory neurons (URX and BAG) use their primary cilia to attach to a specific glial partner (ILsoL). Through a genetic screen, we find that cilia-glia attachment requires BUG-1, a secreted protein with conserved cell adhesion domains that localizes to the cilia-glia attachment site. In the absence of BUG-1, neuronal cilia are present but fail to attach to the glial cell. We find that loss of cilia-glia attachment alters cilia morphology and disrupts stimulus-evoked calcium dynamics in cilia. We propose that primary cilia not only act as antennae for long-range cell communication, but also form close-range cell attachments that modulate cell signaling.

developmental biology↗

bioRxiv2025

Tanycyte-mediated synapse pruning shapes hypothalamic circuits controlling metabolism.

Synaptic pruning by glial cells is critical for refining neural circuits, yet which glial populations mediate this process in hypothalamic networks controlling energy balance remains poorly understood. Here, using 3D-ultrastructural analyses and molecular approaches, we show that tanycytes --specialized glial cells lining the third ventricle-- actively engulf excitatory synapses in the mediobasal hypothalamus during a defined postnatal window. This phagocytic activity relies on tanycytic MFG-E8 and phosphatidylserine exposure on synaptic elements. Disruption of this pathway reduces synapse uptake in vitro and alters body weight control around weaning and glucose homeostasis in adulthood. These findings identify tanycytes as previously unrecognized glial mediators of synaptic pruning in hypothalamic circuits, establishing their active role in shaping neural networks underlying energy balance.

neuroscience↗

bioRxiv2025

A bioengineered human urothelial organoid model reveals the urine-urothelium interplay in tissue resilience and UPEC recurrence in urinary tract infections

Urine is a dynamic and highly variable biofluid. Urine-urothelium interactions are a critical yet underexplored factor in bladder homeostasis and urinary tract infections (UTIs). Here, we report on a human mini-bladder model that exposes a stratified urothelium to urine of defined composition, and incorporates micturition. Prolonged exposure to high-solute concentration urine weakens tight junctions, dysregulates immune responses, and reduces bladder tissue resilience. This increases susceptibility to colonization of the bladder by uropathogenic E. coli (UPEC) which reduces efficacy of antibiotic therapy. In high-solute concentration urine, Fosfomycin monotherapy - prescribed for uncomplicated UTIs, induces the formation of cell wall-deficient (CWD) UPEC in the urine (as observed in patients with recurrent UTIs) but also within deeper urothelial layers. Tissue-associated CWD UPEC directly contributed to recurrence. Our findings expand the conceptual role for CWD UPEC in UTIs, and demonstrate the power of the mini-bladder platform to capture urine-urothelial microenvironment dynamics that actively shape UTI pathogenesis and antibiotic tolerance.

bioengineering↗

bioRxiv2025

Tanycytic annexinA1-containing extracellular vesicles control thermogenesis by orchestrating microglial and neuronal functions.

Obesity, a major global health issue, results from disrupted energy balance driven by chronic hypothalamic inflammation and altered intercellular communication. Among the diverse cells orchestrating this regulation, tanycytes--specialized ependymal cells at the brain-blood interface--have emerged as key modulators, yet the molecular mechanisms by which they influence surrounding cells remain poorly understood. Here, we identify Annexin A1 (ANXA1) as a tanycyte-derived anti-inflammatory signal whose expression, localization, and secretion are dynamically regulated by nutritional state and altered under high-fat diet. During positive energy balance, ANXA1 is secreted in CD9 extracellular vesicles (EV), remodeling hypothalamic networks by altering microglial morphology, synaptic density, and neuronal activation. These EV-mediated effects extend systemically to regulate brown adipose tissue thermogenesis, glucose homeostasis, and overall energy balance. Our findings reveal a previously unrecognized tanycyte-microglia-neuron signaling axis and highlight EV-mediated glial communication as a potential therapeutic target in obesity-associated neuroinflammation.

neuroscience↗

bioRxiv2025

Embryonic development of C. elegans sense organs

C. elegans sense organs provide a powerful model for understanding how different cell types interact to assemble a functional organ. Each sense organ is composed of two glial cells, called the sheath and socket, and one or more neurons. A major challenge in studying their development has been the lack of methods to directly observe these structures in the embryo. Here, we mine a recently published high-resolution ultrastructural dataset of a comma-stage embryo that provides an untapped resource for visualizing early developmental events. From this dataset we reconstructed all head sense organs (two amphid (AM), four cephalic (CEP), six inner labial (IL), four outer labial quadrant (OLQ), and two outer labial lateral (OLL)). Symmetric sense organs were at different stages of morphogenesis, allowing us to infer developmental steps by which they form. First, we found that the sheath glial cell begins wrapping its partner neurons at the distal tip of the dendrites where it self-fuses into a seamless tube and then "zippers" down the dendrite. In many cases, sheath glia wrap the progenitors of partner neurons prior to their terminal division. After sheath wrapping has begun, the socket glia wraps the sheath glia circumferentially before presumably elongating to form the mature sheath-socket channel. We also observed transient interactions not found in the mature animal, such as amphid sheath glia wrapping the AUA neuron, that may reflect ancestral relationships. This study demonstrates the value of large public EM datasets that can be mined for new insights, and sheds light on how neurons and glia undergo coordinated morphogenesis.

developmental biology↗

bioRxiv2025

Complementary Volume Electron Microscopy-based approaches reveal ultrastructural changes in germline intercellular bridges of D. melanogaster

Intercellular bridges are essential to connect developing germline cells. The Drosophila melanogaster egg chamber is a powerful model system to study germline intercellular bridges, or ring canals (RCs). RCs connect the developing oocyte to supporting nurse cells, and defects in their stability or growth lead to infertility. Despite their importance, it has been technically difficult to use electron microscopy-based approaches to monitor changes in RC structure during oogenesis. Here, we describe the application of a complementary set of volume EM-based approaches to visualize ultrastructural changes in the germline RCs. The combination of array tomography (AT) and focused ion beam (FIB) scanning electron microscopy (SEM) has allowed us to gain insight into previously unappreciated aspects of RC structure. We were able to quantify differences in RC size and thickness within and between germ cell clusters at different developmental stages. Within a cluster, RC size correlates with lineage; the largest RCs were formed during the first division, and the smallest RCs were formed during the fourth mitotic division. We observed the formation of membrane interdigitations in the vicinity of RCs much earlier than previously reported, and reconstruction of a RC from a mid-stage EC provided insight into the 3D orientation of these extensive cell-cell contacts. Our imaging also revealed a novel membrane structure that appeared to line the interior of the RC lumen. Although the focus was on ultrastructural changes in the germline RCs, our dataset contains valuable details of additional cell types and structures, including the fusome, the germline stem cells and their niche, and the migrating border cells. This imaging framework could be applied to other tissues or samples that face similar technical challenges, where the small structure of interest is located within a large sample volume.

developmental biology↗

bioRxiv2023

A sex-specific switch in a single glial cell patterns the apical extracellular matrix

Apical extracellular matrix (aECM) constitutes the interface between every tissue and the outside world. It is patterned into diverse tissue-specific structures through unknown mechanisms. Here, we show that a male-specific genetic switch in a single C. elegans glial cell patterns the aECM into a [~]200 nm pore, allowing a male sensory neuron to access the environment. We find that this glial sex difference is controlled by factors shared with neurons (mab-3, lep-2, lep-5) as well as previously unidentified regulators whose effects may be glia-specific (nfya-1, bed-3, jmjd-3.1). The switch results in male-specific expression of a Hedgehog-related protein, GRL-18, that we discover localizes to transient nanoscale rings at sites of aECM pore formation. Blocking male-specific gene expression in glia prevents pore formation, whereas forcing male-specific expression induces an ectopic pore. Thus, a switch in gene expression in a single cell is necessary and sufficient to pattern aECM into a specific structure.

developmental biology↗

bioRxiv2022

Autophagy degrades myelin proteins and is essential for maintaining CNS myelin homeostasis

(Macro)autophagy comprises a major lysosome-dependent degradation mechanism which engulfs, removes and recycles unwanted cytoplasmic material, including damaged organelles and toxic protein aggregates. Although a few studies implicate autophagy in CNS demyelinating pathologies, its role, particularly in mature oligodendrocytes and CNS myelin, remains poorly studied. Here, using both pharmacological and genetic inhibition of the autophagic machinery, we provide evidence that autophagy is an essential mechanism for oligodendrocyte maturation in vitro. Our study reveals that two core myelin proteins, namely proteolipid protein (PLP) and myelin basic protein (MBP) are incorporated into autophagosomes in oligodendrocytes, resulting in their degradation. Furthermore, we ablated atg5, a core gene of the autophagic machinery, specifically in myelinating glial cells in vivo by tamoxifen administration (plp-CreERT2; atg5F/F) and showed that myelin maintenance is perturbed, leading to PLP accumulation. Significant morphological defects in myelin membrane such as decompaction accompanied with increased axonal degeneration are observed. As a result, the mice exhibit behavioral deficits. In summary, our data highlight that the maintenance of adult myelin homeostasis in the CNS requires the involvement of a fully functional autophagic machinery.

neuroscience↗

bioRxiv2021

Progenitor cell integration into a barrier epithelium during adult organ turnover

Barrier epithelial organs face the constant challenge of sealing the interior body from the external environment while simultaneously replacing the cells that contact this environment. These replacement cells--the progeny of basal stem cells--are born without apical, barrier-forming structures such as a protective, lumen-facing membrane and occluding junctions. How stem cell progeny acquire these structures to become part of the barrier is unknown. Here we use Focused Ion Beam-Scanning Electron Microscopy (FIB-SEM), Correlative Light-Electron Microscopy (CLEM), and volumetric imaging of live and fixed organs to investigate progenitor integration in the intestinal epithelium of adult Drosophila. We find that stem cell daughters gestate their future lumenal-apical membrane beneath a transient, basal niche formed by an umbrella-shaped occluding junction that shelters the growing cell and adheres it to mature neighbor cells. The umbrella junction both targets formation of a deep, microvilli-lined, apical invagination and closes it off from the contents of the gut lumen. When the growing cell is sufficiently mature, the umbrella junction retracts to expose this Pre-Assembled Apical Compartment (PAAC) to the gut lumen, thus incorporating the new cell into the intestinal barrier. When we block umbrella junctions, stem cell daughters grow and attempt to differentiate but fail to integrate; when we block cell growth, no umbrella junctions form and daughters arrest in early differentiation. Thus, stem cell progeny build new barrier structures in the shelter of a transient niche, where they are protected from lumenal insults until they are prepared to withstand them. By coordinating this dynamic junctional niche with progenitor cell differentiation, a physiologically active epithelial organ incorporates new cells while upholding integrity of its barrier.

developmental biology↗

bioRxiv2020

Multicellular rosettes organize neuropil formation

Neuropils are compartments in the nervous system containing dense networks of neurites and synapses which function as information processing centers. Neuropil formation requires structural and functional organization at and across different scales, achieving single-axon precision for circuits that carry out the core functions while simultaneously accommodating variability among individuals [1; 2; 3; 4]. How these organizational features emerge over development is poorly understood. The nerve ring is the primary neuropil in C. elegans, and its structure is thoroughly mapped [5; 6]. We show that prior to axon outgrowth, nerve ring neurons form a ring of multicellular rosettes with surrounding cells to organize the stratified nerve ring structure [7; 8]. Axon bundles which correspond to future nerve ring strata grow from rosette centers, travel along the ring on "bridge" cells that are simultaneously engaged in adjacent rosettes, and assemble into a topographic scaffold of the nerve ring. SAX-3/Robo is required for proper rosette formation and outgrowth from the center. Furthermore, axon contact sites that form early in development are more conserved than the later ones, indicating a temporal component in neuropil structural variability. Our results reveal an unexpected and critical role of collective cell behaviors prior to innervation to pattern a complex neuropil and orchestrate its formation across scales.

neuroscience↗

bioRxiv2018

Morphogenesis of neurons and glia within an epithelium

To sense the outside world, some neurons protrude across epithelia, the cellular barriers that line every surface of our bodies. To study the morphogenesis of such neurons, we examined the C. elegans amphid, in which dendrites protrude through a glial channel at the nose. During development, amphid dendrites extend by attaching to the nose via DYF-7, a type of protein typically found in epithelial apical ECM. Here, we show that amphid neurons and glia exhibit epithelial properties, including tight junctions and apical-basal polarity, and develop in a manner resembling other epithelia. We find that DYF-7 is a fibril-forming apical ECM component that prevents rupture of the tube-shaped glial channel, reminiscent of roles for apical ECM in other narrow epithelial tubes. We also identify a role for FRM-2, a homolog of EPBL15/moe/Yurt which promote epithelial integrity in other systems. Finally, we show that other environmentally-exposed neurons share a requirement for DYF-7. Together, our results suggest that these neurons and glia can be viewed as part of an epithelium continuous with the skin, and are shaped by mechanisms shared with other epithelia.

developmental biology↗

bioRxiv2017

CYK-4 functions independently of its centralspindlin partner ZEN-4 to cellularize oocytes in germline syncytia

Throughout metazoans, germ cells undergo incomplete cytokinesis to form syncytia connected by intercellular bridges. Formation of gametes ultimately requires bridge closure. Here, we investigate the contribution of the conserved bridge component centralspindlin to oocyte production in C. elegans. Centralspindlin is composed of the Rho family GTPase-activating protein (GAP) CYK-4/MgcRacGAP and the microtubule motor ZEN-4/kinesin-6, which are both essential for cytokinesis. In contrast, we show that oocyte production by the syncytial germline requires CYK-4 but not ZEN-4. Longitudinal imaging after conditional CYK-4 inactivation revealed a role in oocyte cellularization, rather than in generation of syncytial compartments. CYK-4s lipid-binding C1 domain and the GTPase-binding interface of its GAP domain were individually important for oocyte cellularization and for targeting CYK-4 to bridges, where it contributes to enrichment of active RhoA. These results identify a C1-GAP module in CYK-4 that recruits it to bridges in the germline and directs their closure to produce oocytes.\n\nIMPACT STATEMENTThe CYK-4 subunit of centralspindlin, a broadly conserved component of intercellular bridges across metazoa, is required for the cytokinesis-like closure of intercellular bridges that cellularizes oocytes to separate them from germline syncytia.\n\nMAJOR SUBJECT AREASCell Biology, Developmental Biology & Stem Cells

cell biology↗