Biology subjects

Classen, A.-K.

Publications and source records attributed to Classen, A.-K..

14 recordsLinked to original sources

bioRxiv2026

Context and exposure history shape insecticide sensitivity in Drosophila melanogaster

Current insecticide toxicology still lacks an integrated understanding of how lethal and sublethal exposure shape organismal performance across biologically and environmentally relevant contexts. In particular, insecticide susceptibility may vary with physiological state, environmental conditions, and recent exposure history, factors that are only partially captured by standardized toxicity assessments. Here, using Drosophila melanogaster, we systematically compared responses to three insecticides with distinct modes of action - acetamiprid, chlorantraniliprole, and deltamethrin - across developmental stages, sexes, adult ages, and ambient temperatures. Insecticide sensitivity varied substantially across these conditions, with temperature modulating toxicity in a compound-dependent manner. Notably, deltamethrin caused substantial lethality at concentrations more than 10,000-fold below approved field application rates. Beyond these context-dependent differences in acute toxicity, sublethal pre-exposure to each insecticide rapidly reduced lethality during a subsequent challenge with the same compound, with increased tolerance already detectable after 24 h. Importantly, these exposure conditions that enhanced insecticide tolerance were associated with reduced resistance to environmental challenges, including heat and nutrient starvation, revealing a potential trade-off in organismal stress resilience. Acetamiprid exposure further altered reproductive performance in a concentration-dependent manner, with higher exposure reducing egg production while increasing developmental success among the resulting offspring, such that the number of offspring reaching adulthood remained largely unchanged. Together, these findings demonstrate that insecticide susceptibility is highly context dependent and can be rapidly modified by recent exposure history. We identify short-term, compound-specific, and potentially costly tolerance as an underappreciated consequence of insecticide exposure and highlight the importance of incorporating biological context, environmental conditions, and sublethal exposure history into pesticide risk assessment.

pharmacology and toxicology↗

bioRxiv2026

The dynamic inositol phosphate network during development in Drosophila melanogaster

Inositol phosphates (InsPs) and inositol pyrophosphates (PP-InsPs) are key regulators of cellular signaling and metabolism, yet developmental changes in the InsP/PP-InsP network in eukaryotic systems remain poorly understood. Here, we combine isomer-resolved capillary electrophoresis-mass spectrometry (CE-MS) with genetic perturbations to characterize InsP metabolism during the Drosophila melanogaster development. Our analyses reveal extensive developmental reprogramming of the InsP pathway, with early developmental stages exhibiting broad lower-order isomer diversity that progressively narrows into a more restricted metabolic state enriched in Ins(1,3,4,5,6)P5, InsP6, and PP-InsPs at later stages. Metamorphosis is characterized by a marked increase in 5-PP-InsP5, which emerges as the predominant pyrophosphate species in adults. Perturbation of dedicated kinases suggests that this metabolic remodeling is essential for developmental progression. RNAi-mediated IPK2 depletion disrupts higher-order InsP synthesis by reducing InsP5 levels, and results in failed metamorphosis. IPK1 depletion delays development and induces pronounced isomer-selective redistribution within the InsP5 pool, indicating network-level changes beyond simple precursor accumulation. IP6K depletion specifically abolishes 5-PP-InsP5 production, impairs tissue morphogenesis during pupal stages, and causes rapid post-eclosion lethality. Unexpectedly, depletion of either IPK2 or IP6K leads to the presence of 2-PP-InsP5. Collectively, these findings establish the Drosophila melanogaster InsP pathway as a dynamically regulated developmental network, particularly rich in previously undescribed InsP isomers, in which stage-specific metabolic remodeling parallels metamorphosis, tissue maturation, and adult viability.

biochemistry↗

bioRxiv2025

Cell cycle dynamics regulate H3K27 and H3K9 histone modifications

Cell cycle progression presents a fundamental challenge to genome integrity, particularly due to the need to reestablish post-translational histone modifications (PTMs) following DNA replication. Although proliferative and differentiating tissues exhibit markedly different cell cycle dynamics, how these differences shape the histone modification landscape in vivo remains largely unexplored. Here, we show that levels of H3K27ac, H3K27me3, and H3K9me3 are tightly linked to cell cycle dynamics in the Drosophila wing imaginal disc. We demonstrate that both physiological and pathological elongation of the cell cycle led to an accumulation of H3K9me3 and H3K27me3, whereas cell cycle acceleration reduces their levels. In contrast, H3K27ac exhibits the opposite pattern: levels decrease in arrested cells and increase with faster cycling. Genome-wide CUT&Tag analysis reveals that these changes predominantly affect genomic loci already modified in normally proliferating tissue. Importantly, the regulation of methylation levels at H3K9 and H3K27 is not solely mediated by the cell cycle machinery but reflects a metabolically guided process in which the rate of methylation is coupled to the rate of cell proliferation through metabolic activity, including signaling via the Insulin/PI3K/Akt pathway. Our study thus reveals key principles for understanding histone methylation in proliferating, senescent, and differentiating cells. In contrast, H3K27 acetylation is regulated through a distinct, cell cycle-coupled mechanism. We find that CBP/Nejire-mediated acetylation of H3K27 peaks during S-phase and is reversed by HDAC1, as cells exit replication. Disruption of this acetylation cycle leads to replication stress and a G2 cell cycle arrest via a DNA damage checkpoint. Notably, this genome-protective function of CBP/Nejire depends specifically on acetylation of the H3K27 residue itself, revealing a novel role for H3K27ac beyond its well-established function in transcriptional activation. Together, our findings establish a robust link between cell cycle progression and histone modification dynamics, highlighting the necessity of maintaining balanced PTM levels under varying proliferative states. These insights have broad implications for our understanding of development, aging, and tumor growth.

cell biology↗

bioRxiv2025

Deep learning-based image quantification of epithelial cell shapes and its application to polycystic kidney disease

Cell shape is a fundamental determinant of tissue architecture and organ function. In epithelial tissues, cytoskeletal organization and tight junctions regulate cell geometry, shaping functional tissue units. Disruption of these mechanisms may cause diseases such as autosomal dominant polycystic kidney disease (ADPKD), in which cyst formation is characterized by abnormal regulation of epithelial cell shape. The mechanisms of cystogenesis remain incompletely understood, highlighting the need for robust, high-throughput methods to quantify the morphology of epithelial cells. Here, we present a fully automated, deep learning-based image analysis pipeline to quantify epithelial cell shape and tight junction morphology from immunofluorescence images. Our approach employs a U-Net convolutional neural network for accurate segmentation of fluorescence labeled tight junctions. We introduce novel algorithms to quantify overall cell shape and tight junction morphology, as well as to estimate cytoskeletal traction at shared cell borders. Our analysis pipeline objectively identifies subtle morphogenetic changes associated with disease-related mutations, applied to a genetically modified Madin-Darby Canine Kidney cell model of ADPKD. The method enables high-throughput, standardized analysis, reduces observer bias, and facilitates comparison across experiments. We further demonstrate the pipelines generalizability by applying it to Drosophila egg chamber epithelia. Our results establish a robust and scalable framework for analyzing cell shape and mechanical interactions in epithelial tissues, with broad applications in phenotypic screening, disease modeling, and morphogenesis research. Author SummaryThe shape of epithelial cells is critical for organ function. In the kidney, properly shaped epithelial cells assemble to tubules ensuring efficient waste excretion as well as body electrolyte and water balance. Disruption of cell shape regulation can lead to diseases such as autosomal dominant polycystic kidney disease (ADPKD), characterized by cyst formation and displacement of normal kidney tissue. Traditionally, analysis of epithelial cell morphology has relied on manual, low-throughput methods, which are time-consuming and prone to error. To overcome these limitations, we developed a fully automated, artificial intelligence-based pipeline that rapidly and reliably quantifies cell shape and junctional organization from microscopic images. We validated our approach using a cellular model of ADPKD, demonstrating clear differences in cell shape and junctional structure between normal and mutant cells harboring mutations in PKD-related genes. Our method enables efficient, objective analysis of large datasets and provides a powerful tool for understanding the mechanisms underlying cell shape regulation in health and disease.

bioinformatics↗

bioRxiv2025

Toll receptors mediate tissue intrinsic surveillance against aberrant cells by detecting cell fate aberrations

Tissue-intrinsic surveillance systems play an important role in maintaining tissue health by detecting and eliminating aberrant cells. One such system, interface surveillance, is activated by differences in cell fate programs between neighboring cells. Activation of interface surveillance drives separation and elimination of aberrant cells by recruitment of actin regulators and Myosin II to neighboring cell contacts, as well as activation of JNK signaling and apoptosis in cells at this interface. Here, we uncover the role of long Toll receptors (Toll-2, Toll-6, Toll-7, Toll-8) as key mediators of interface surveillance in Drosophila wing imaginal discs. Using genetic mosaics combined with spatial mapping of Toll receptor expression patterns, we demonstrate that differences in long Toll receptor expression levels between neighboring cells trigger all hallmark responses of interface surveillance. This response is position-dependent and relies on the comparison of expression levels established by cell-fate-specifying programs. Specifically, we show that the expression of long Toll receptors is regulated by multiple fate-patterning pathways, giving rise to a combinatorial cell surface code that is deregulated in developmentally aberrant or oncogenic cells. Remarkably, Toll receptor-mediated surveillance operates independently of canonical NF-{kappa}B signaling, and rather mirrors a distinct role of long Toll receptors in modulating cell-cell affinity via actomyosin dynamics during early embryogenesis. Our findings identify long Toll receptors as integrators of developmental patterning and tissue homeostasis mechanisms and provide insights into how tissues detect and respond to potentially oncogenic mutations that misregulate cell fate specification pathways.

cell biology↗

bioRxiv2025

The emergence of E-Cadherin spot junctions between germline and somatic cells facilitates late stage oogenesis in Drosophila

Coordinated tissue morphogenesis relies on precise interactions between distinct cell lineages. During development of the Drosophila oocyte, interactions between di_erent somatic follicle cell types and the germline-derived nurse cells and the future oocyte are crucial for coordinating oogenesis. However, the molecular basis of their interaction throughout enormous morphological changes remains largely unexplored. During late oogenesis, nurse cells transfer their cytoplasmic content into the oocyte, while anterior follicle cells (AFCs) must closely envelop the nurse cells to induce phagoptosis and remove their remnants. We identify E-cadherin-based spot junctions as novel adhesive complexes at this essential soma-germline interface. AFCs reorganize their E-cadherin from intraepithelial adherens junctions to their apical surface to form interlineage spot junctions with nurse cells, representing a switch in adhesive and thus mechanical coupling. The distribution and density of these junctions is stably organized, even as the interface undergoes dramatic size changes. Additionally, we show that these junctions integrate into a microvilli-rich nurse cell surface, influencing microvilli architecture. Importantly, disruption of E-cadherin spot junctions impairs nurse cell enveloping. Our findings reveal a switch from intra-lineage to inter-lineage mechanical coupling that is driven by the formation of E-Cadherin spot junctions and highlights their essential role in ensuring robust soma-germline coupling and coordination during nurse cell dumping in late oogenesis.

developmental biology↗

bioRxiv2024

PDK-1/S6K and mTORC1 bypass systemic growth restrictions to promote regeneration

Inflammation triggers systemic signals that induce growth restrictions in distant organs, a process well characterized in tumor cachexia. While mechanisms allowing tumors to circumvent these systemic growth restrictions have been established, the physiological processes that overcome inflammation-induced growth restrictions during regeneration remain largely unexplored. In our study, we use a model of tissue inflammation and regeneration in developing Drosophila imaginal discs to dissect key metabolic and signaling adaptations that, in physiological settings, both induce and overcome systemic growth restrictions. We find that expression of eiger, the Drosophila TNF- homolog, induces systemic insulin restriction, as evidenced by reduced expression of dILP2 and dILP5, as well as elevated nuclear dFOXO, and reduced protein translation and proliferation in peripheral tissues. Proliferating cells overcome systemic insulin restriction by upregulating Pdk1, which is both necessary and sufficient to promote phosphorylation of ribosomal protein S6 and protein translation via an Insulin/Akt-independent mechanism. We further demonstrate that JAK/STAT signaling acts upstream to increase Pdk1 levels, delineating a novel JAK/STAT-Pdk1-S6K axis essential for regenerative proliferation. The upregulation of amino acid transporters in the proliferative domain further suggests that regenerating cells preferentially import amino acids, fueling mTORC1 activation. Similar metabolic signatures are observed in a Drosophila RasV12, scrib tumor model, suggesting that tumors may co-opt these metabolic pathways to sustain growth in an insulin-restricted environment. Our findings reveal a specialized metabolic program that integrates systemic nutrient mobilization with local metabolic reprogramming, with important implications for understanding of physiological tissue repair but also pathologies such as chronic wounds and cancer.

developmental biology↗

bioRxiv2023

Drosophila macrophages control systemic cytokine levels in response to oxidative stress via a non-canonical DNA damage repair signaling cascade

Environmental factors, infection, or injury can cause oxidative stress in diverse tissues and loss of tissue homeostasis. Effective stress response cascades, conserved from invertebrates to mammals, ensure reestablishment of homeostasis and tissue repair. Hemocytes, the Drosophila blood-like cells, rapidly respond to oxidative stress by immune activation. However, the precise signals how they sense oxidative stress and integrate these signals to modulate and balance the response to oxidative stress in the adult fly are ill-defined. Furthermore, hemocyte diversification was not explored yet on oxidative stress. Here, we employed high throughput single nuclei RNA-sequencing to explore hemocytes and other cell types, such as fat body, during oxidative stress in the adult fly. We identified distinct cellular responder states in plasmatocytes, the Drosophila macrophages, associated with immune response and metabolic activation upon oxidative stress. We further define oxidative stress-induced DNA damage signaling as a key sensor and a rate-limiting step in immune-activated plasmatocytes controlling JNK-mediated release of the pro-inflammatory cytokine unpaired-3. We subsequently tested the role of this specific immune activated cell stage during oxidative stress and found that inhibition of DNA damage signaling in plasmatocytes, as well as JNK or upd3 overactivation, result in a higher susceptibility to oxidative stress. Our findings uncover that a balanced composition and response of hemocyte subclusters is essential for the survival of adult Drosophila on oxidative stress by regulating systemic cytokine levels and cross-talk to other organs, such as the fat body, to control energy mobilization.

immunology↗

bioRxiv2023

A cell surface code mediates tissue-intrinsic defense against aberrant cells in epithelia

Tissue-intrinsic error-correction mechanisms allow epithelial cells to detect aberrant neighboring cells and cause their removal from the tissue. The molecular mechanisms which grant cells the ability to compare their internal states is unknown. Here we demonstrate that comparison of cell identity, created by cell-fate-specifying transcription factors and patterning pathways, is conveyed through a specific set of cell surface molecules. We demonstrate that Drosophila imaginal discs express a range of cell surface molecules previously implicated in neuronal axon guidance processes, such as members of the Robo, Teneurin, Ephrin, Toll-like or atypical Cadherin families. Expression of these molecules is regulated by intrinsic fate-patterning pathways of the disc but also by aberrant expression of oncogenic RasV12. Importantly, mosaic clones deregulating individual cell surface molecules are sufficient to induce all hallmarks of interface surveillance, a tissue-intrinsic error-correction mechanism previously shown to be induced by cells with aberrant activation of fate-patterning pathways. Specifically, cells with deregulated expression of Robo2 and Robo3 induce actomyosin enrichment, bilateral JNK signaling and apoptosis at mosaic clone interfaces in imaginal discs. Moreover, deregulation of Robo2 levels, which is normally expressed in a complex endogenous pattern, induces these interface surveillance hallmarks in a Robo2-pattern-specific manner. Taken together, our work indicates that these cell surface molecules mediate cell fate recognition in epithelial tissues and thereby contribute to the maintenance of epithelial health by initiating detection and removal of aberrant cells during development and adult tissue homeostasis.

cell biology↗

bioRxiv2022

Eya-controlled affinity between cell lineages drives tissue self-organization during Drosophila oogenesis

The cooperative morphogenesis of cell lineages underlies the development of functional units and organs. To study mechanisms driving the coordination of lineages, we investigated soma-germline interactions during oogenesis. From invertebrates to vertebrates, oocytes develop as part of a germline cyst that consists of the oocyte itself and so-called nurse cells, which feed the oocyte and are eventually removed. The enveloping somatic cells specialize to either facilitate oocyte maturation or nurse cell removal, which makes it essential to establish the right match between germline and somatic cells. We uncover that the transcriptional regulator Eya, expressed in the somatic lineage, controls bilateral cell-cell affinity between germline and somatic cells in Drosophila oogenesis. Employing functional studies and mathematical modelling, we show that differential affinity proportional to Eya expression and the resulting forces drive somatic cell redistribution over the germline surface and control oocyte growth to match oocyte and nurse cells with their respective somatic cells. Thus, our data demonstrate that differential affinity between cell lineages is sufficient to drive the complex assembly of inter-lineage functional units and underlies tissue self-organization during Drosophila oogenesis.

developmental biology↗

bioRxiv2022

Bilateral JNK activation is a hallmark of interface contractility and promotes elimination of aberrant cells

Tissue-intrinsic defence mechanisms eliminate aberrant cells from epithelia and thereby maintain the health of developing tissues or adult organisms. Interface contractility comprises one such distinct mechanism that specifically guards against aberrant cells, which undergo inappropriate cell fate and differentiation programs. The cellular mechanisms which facilitate detection and elimination of these aberrant cells are currently unknown. We find that in Drosophila imaginal discs, interface contractility is associated with bilateral JNK activation at the clonal interface of wild type and aberrant cells. Bilateral JNK activation is unique to interface contractility and is not observed in other tissue-intrinsic defence mechanisms, such as cell-cell competition. We find that JNK is activated cell-autonomously by either of the contacting cell types and drives apoptotic elimination of cells at clonal interfaces. Ultimately, JNK interface signalling provides a collective tissue-level mechanism, which ensures elimination of aberrant, misspecified cells that cannot be identified by cell fitness criteria, as in cell-cell competition. Importantly, oncogenic Ras activates interface contractility but evades apoptotic elimination by bilateral JNK activation. Combined, our work establishes bilateral JNK interface signalling and interface apoptosis as a new hallmark of interface contractility, and it highlights how oncogenic mutations evade tumour suppressor function encoded by this tissue-intrinsic surveillance system.

developmental biology↗

bioRxiv2022

Shared enhancer gene regulatory networks between wound and oncogenic programs

Wound response programs are often activated during neoplastic growth in tumors. In both wound repair and tumor growth, cells respond to acute stress and balance the activation of multiple programs including apoptosis, proliferation, and cell migration. Central to those responses are the activation of the JNK/MAPK and JAK/STAT signaling pathways. Yet, to what extent these signaling cascades interact at the cis-regulatory level, and how they orchestrate different regulatory and phenotypic responses is still unclear. Here, we aim to characterize the regulatory states that emerge and cooperate in the wound response, using the Drosophila melanogaster wing disc as a model system, and compare these with cancer cell states induced by rasV12scrib-/- in the eye disc. We used single-cell multiome profiling to derive enhancer Gene Regulatory Networks (eGRNs) by integrating chromatin accessibility and gene expression signals. We identify a "proliferative" eGRN, active in the majority of wounded cells and controlled by AP-1 and STAT. In a smaller, but distinct population of wound cells, a "senescent" eGRN is activated and driven by C/EBP-like transcription factors (Irbp18, Xrp1, Slow border, and Vrille) and Scalloped. In tumor cells on the other hand, both eGRN signatures are simultaneously active within the same cell. Our single-cell multiome and eGRNs resource offers an in-depth characterisation of the senescence markers, together with a new perspective on the shared gene regulatory programs acting during wound response and oncogenesis.

cell biology↗

bioRxiv2022

Mutual repression between JNK/AP-1 and JAK/STAT stratifies cell behaviors during tissue regeneration

Epithelial repair relies on the activation of stress signaling pathways to coordinate cellular repair behaviors. Their deregulation is implicated in chronic wound and cancer pathologies. Despite such translational importance, an understanding of how spatial patterns of signaling pathways and repair behaviors arise in damaged tissues remains elusive. Using TNF-/Eiger-mediated inflammatory damage to Drosophila imaginal discs, we uncover that JNK/AP-1 signaling cells act as paracrine organizers and initiate a mutual repression network that spatially segregates JNK/AP-1 and JAK/STAT signaling cells into distinct populations. While JNK/AP-1 signaling cells produce JAK/STAT-activating Upd ligands, these signal-sending cells suppress activation of JAK/STAT via Ptp61F. Conversely, responding cells with activated JAK/STAT suppress JNK activation via Zfh2. The resulting bistable segregation of signaling domains is associated with distinct cellular tasks and regenerative potential. While JNK/AP-1 signaling cells at the wound center act as paracrine organizers, their cell cycle is senescently arrested. Thus, compensatory proliferation occurs exclusively in JAK/STAT signaling cells at the wound periphery. This spatial stratification is essential for proper tissue repair, as co-activation of JNK/AP-1 and JAK/STAT in the same cells creates conflicting inputs on cell cycle progression, leading to excess apoptosis of senescently arrested organizer cells. Finally, we demonstrate that bistable spatial segregation of JNK/AP-1 and JAK/STAT drives senescent and proliferative behaviors in transient as well as chronic tissue damage models, and importantly, in RasV12, scrib tumors under the influence of JNK/AP-1 activity. Revealing this previously uncharacterized regulatory network between JNK/AP-1, JAK/STAT and associated cell behaviors have important implications for our conceptual understanding of tissue repair, chronic wound pathologies and tumor microenvironments, where both pathways are strongly implicated.

developmental biology↗

bioRxiv2018

Regulation of tensile stress in response to external forces coordinates epithelial cell shape transitions with organ growth and elongation

The role of actomyosin contractility at epithelial adherens junctions has been extensively studied. However, little is known about how external forces are integrated to establish epithelial cell and organ shape in vivo. We use the Drosophila follicle epithelium to investigate how tension at adherens junctions is regulated to integrate external forces arising from changes in germline size and shape. We find that overall tension in the epithelium decreases despite pronounced growth of enclosed germline cells, suggesting that the epithelium relaxes to accommodate growth. However, we find local differences in adherens junction tension correlate with apposition to germline nurse cells or the oocyte. We demonstrate that medial Myosin II coupled to corrugating adherens junctions resists nurse cell-derived forces and thus maintains apical surface areas and cuboidal cell shapes. Furthermore, medial reinforcement of the apical surface ensures cuboidal-to-columnar cell shape transitions and imposes circumferential constraints on nurse cells guiding organ elongation. Our study provides insight into how tension within an adherens junction network integrates growth of a neighbouring tissue, mediates cell shape transitions and channels growth into organ elongation.

developmental biology↗