Biology subjects

Deckert, M.

Publications and source records attributed to Deckert, M..

9 recordsLinked to original sources

bioRxiv2026

Spectral CytoFRET2 identifies lysine acetyltransferase inhibitors as modulators of vimentin assembly

Cytoskeletal plasticity is a defining feature of cancer progression, enabling tumor cells to adapt their morphology, mechanics, and migratory behavior during invasion and metastasis. Although actin filaments, microtubules, and intermediate filaments are known to cooperate in these processes, the molecular mechanisms coordinating their dynamics remain incompletely understood, particularly the role of post-translational modifications (PTMs). Here, we developed CytoFRET2, a multiparametric cytometry-based FRET platform that enables real-time and simultaneous monitoring of the dynamics of actin filaments, microtubules, and vimentin in living suspension cells. The system combines fluorescently tagged cytoskeletal reporters with spectral flow cytometry, allowing simultaneous high-content analysis of multiple cytoskeletal networks while overcoming autofluorescence and fluorescence interference from small molecules. Using spectral CytoFRET2, we screened a small library of epigenetic compounds targeting regulators of acetylation and methylation pathways. The screen revealed that inhibition of lysine deacetylases (KDACs) and sirtuins promoted stabilization of both microtubules and vimentin filaments, without impacting actin filament organization. In contrast, inhibition of lysine acetyltransferases (KATs), particularly with garcinol and anacardic acid, induced rapid vimentin disassembly. Mechanistically, the study reveals acetylation as a key post-translational modification regulating the dynamics of microtubules and vimentin filaments, with KAT inhibitors emerging as potent modulators of vimentin organization. Together, the findings establish spectral CytoFRET2 as a versatile platform for systematic investigation of cytoskeletal regulatory networks and therapeutic vulnerabilities in cancer.

cell biology↗

bioRxiv2026

PSMD14 drives melanoma cell survival and MAPK inhibitor resistance through histone H2A deubiquitination

Melanoma progression and resistance to targeted therapies remain major clinical challenges, driven in part by the remarkable phenotypic plasticity of melanoma cells. Identifying molecular mechanisms that couple tumor survival to adaptative drug responses is therefore essential. Here, we identify PSMD14, a proteasome-associated deubiquitinase, as an essential regulator of melanoma plasticity, growth, survival, and therapeutic resistance with strong prognostic significance in metastatic disease. An unbiased siRNA screen targeting the human deubiquitinase family revealed PSMD14 (proteasome 26S subunit, non-ATPase 14) as a top regulator of melanoma cell proliferation. Integrative analyses of DepMap, TCGA, and patient-derived datasets revealed that PSMD14 is frequently upregulated in melanoma, enriched in metastatic lesions, and significantly associated with poor patient outcome. Functional and pharmacological studies demonstrated that genetic depletion or inhibition of PSMD14 suppresses proliferation, clonogenic and long-term growth, and viability of melanoma cells across BRAF-, NRAS-, and NF1-driven genotypes, while inducing DNA damage and apoptosis. Consistently, PSMD14 inhibition markedly reduced tumor growth in Nras and Braf syngeneic mouse models. Mechanistically, we uncover a non-proteolytic role for PSMD14 as an epigenetic regulator of chromatin state. Proteomic and biochemical analyses identified histone H2A as a direct interactor and substrate of PSMD14. PSMD14 deubiquitinates H2A at lysine 119 independently of the proteasome, antagonizing the Polycomb E3 ligase RING1B. Loss of PSMD14 allows the increment of H2AK119 ubiquitination, transcriptional repression of pro-survival genes, including MCL1 and BCL2, and apoptotic cell death, effects rescued by RING1B depletion. Importantly, we demonstrate that the PSMD14-H2A axis governs melanoma adaptation to MAPK pathway inhibition. PSMD14 expression and H2AK119 ubiquitination dynamically correlate with therapeutic response, drug-tolerant persistence, and acquired resistance. Targeting PSMD14 genetically or pharmacologically enhances the efficacy of BRAF and MEK inhibitors, suppresses the emergence of drug-tolerant persister cells, and prevents tumor relapse in vivo. Together, this findings establish PSMD14 as a chromatin-rewiring enzyme that links proteostasis to epigenetic control of melanoma plasticity and therapy resistance, highlighting PSMD14 as a promising biomarker and therapeutic target in aggressive and drug-resistant melanoma.

cancer biology↗

bioRxiv2024

USP9X is a mechanosensitive deubiquitinase that controls tumor cell invasiveness and drug response through YAP stabilization

Post-translational modification by ubiquitin is crucial for protein turnover. Deubiquitinases (DUBs) remove ubiquitin chains from target proteins to prevent their degradation by the proteasome, thus acting as gatekeepers of protein homeostasis alongside the ubiquitin-proteasome system (UPS). Tumor cells exhibit remarkable plasticity, enabling them to adapt to anticancer treatments and the conditions of the tumor microenvironment, including mechanical cues from the extracellular matrix (ECM). However, the role of DUBs in mechanotransduction remains unexplored. To identify DUBs involved in cancer cell mechanosignaling, we used melanoma cells grown on collagen matrices with varying stiffnesses and an activity-based ubiquitin probe to profile DUB activities. Our approach, combined with quantitative proteomics, revealed that ubiquitin-specific protease 9X (USP9X) is sensitive to ECM stiffness through discoidin domain receptors (DDR)/actomyosin signaling pathway. In silico analysis further indicated that the mechanosensor YAP is part of the USP9X interactome, and USP9X expression correlates with the YAP transcriptional signature in melanoma. We hypothesized that mechanical signals regulate YAP levels through USP9X DUB activity. Consistently, low collagen stiffness reduced YAP expression, and siRNA-mediated depletion or pharmacological inhibition of USP9X decreased YAP protein expression in tumor cells. Conversely, knockdown of the ubiquitin E3 ligase {beta}TrCP increased YAP protein levels. Affinity purification of polyubiquitinated proteins using Tandem Ubiquitin Binding Entities (TUBEs) showed that combined USP9X and proteasome inhibition increased YAP poly-ubiquitination, revealing that USP9X deubiquitinates YAP to prevent its proteasomal degradation. Targeting USP9X impaired stiffness-mediated responses, including YAP nuclear translocation and transcriptional activity, cell migration and invasion, and drug resistance. An experimental metastasis assay showed that stable knockdown of USP9X impaired melanoma cell lung colonization. Finally, targeting USP9X in a syngeneic BRAF-mutant melanoma model counteracted targeted therapy-induced ECM remodeling, enhanced treatment efficacy, and delayed tumor relapse. Our findings reveal a novel role of USP9X in cancer cell mechanobiology and drug resistance through stiffness-dependent stabilization of the oncoprotein YAP, proposing USP9X as a targetable "mechano-DUB" in cancer.

cancer biology↗

bioRxiv2024

Extracellular matrix stiffness determines the phenotypic behavior of dedifferentiated melanoma cells through a DDR1/2-dependent YAP mechanotransduction pathway

Extracellular matrix (ECM) stiffening, resulting from increased collagen deposition and cross-linking, is a key biophysical factor of the tumor microenvironment. Cutaneous melanoma is a deadly metastatic cancer. Its aggressiveness stems from high intratumoral heterogeneity, resulting from the plasticity of melanoma cells, which transit from a melanocytic state to dedifferentiated therapy-resistant and invasive phenotypes, characterized by mesenchymal and/or neural crest stem cell-like features. Phenotypic plasticity is regulated by stroma-derived soluble factors, but the functional impact of ECM stiffening on melanoma cell phenotypes remains ill defined. Here, we found that melanoma cell subpopulations display difference in mechanical responsiveness. Compared to melanocytic cells, mesenchymal dedifferentiated cells showed increased proliferation, migration and resistance to MAP kinase-targeted therapy when seeded on stiff collagen. By contrast, a soft ECM impaired their proliferation and migration and sensitized them to targeted therapy. In addition, extracellular mechanical signals are required to sustain melanoma cell identity and dedifferentiation features. Further analyses indicated that the mechanosensitivity nature of dedifferentiated cells relies on the expression and activation of collagen receptors DDR1 and DDR2 that control actomyosin cytoskeleton reorganization and YAP mechanotransduction pathway. Inhibiting both DDR in dedifferentiated melanoma cells abrogated their mechano-induced behavior and drug-resistant phenotype, while forcing their expression in melanocytic cells induced mechanical responsiveness and a less differentiated phenotype. Our results reveal that phenotypic reprogramming endows dedifferentiated melanoma cells with increased sensitivity and addiction to ECM stiffness. We propose that mechano-addiction mediated by DDR collagen receptors may represent a novel vulnerability for aggressive dedifferentiated cancer cells that can be exploited for therapeutic benefits.

cancer biology↗

bioRxiv2024

Disrupting USP39 deubiquitinase Function Impairs the Survival and Migration of Multiple Myeloma Cells through ZEB1 Degradation

RationaleMultiple Myeloma (MM) stands as the second most common hematological malignancy characterized by the accumulation of monoclonal plasmocytes within the bone marrow. Despite the introduction of proteasome inhibitors, immunomodulatory agents and CD38-targeting antibodies which have extended survival rates, the disease remains incurable for most patients due to the emergence of resistant clones and frequent relapses. The efficacy of the proteasome inhibitor bortezomib (BTZ) in MM treatment underscores the critical role of the ubiquitin proteasome system (UPS) in this cancer. Deubiquitinases (DUBs), a class of enzymes governing the stability, interactions or localization of cellular proteins by removing ubiquitin modifications, have emerged as promising therapeutic targets across various cancers, including MM. MethodsThrough an exhaustive loss-of-function approach, we have identified for the first time USP39 DUB as a pivotal survival determinant for MM cells. ResultsOur analysis reveals a direct correlation between heightened USP39 mRNA levels and shorter survival in MM patients. Additionally, robust USP39 protein expression is observed in MM patient plasmocytes compared to healthy counterparts. Knockdown of Usp39 not only impedes clonogenic capabilities, but also induces apoptosis, triggers cell cycle arrest and overcomes BTZ resistance. Complementary gain-of-function assays, further elucidate how USP39, by stabilizing the transcription factor ZEB1, enhances the trans-migratory potential of MM cells. ConclusionsIn summary, our findings underscores the pivotal role of the deubiquitinase USP39, suggesting that targeting the USP39/ZEB1 axis hold promise as a prospective diagnostic marker and therapeutic target in MM.

cancer biology↗

bioRxiv2021

Blockade of pro-fibrotic response mediated by the miR-143/-145 cluster prevents targeted therapy-induced phenotypic plasticity and resistance in melanoma

Lineage dedifferentiation towards a mesenchymal-like state is a common mechanism of adaptive response and resistance to targeted therapy in melanoma. Yet, the transcriptional network driving this phenotypic plasticity remains elusive. Remarkably, this cellular state displays myofibroblast and fibrotic features and escapes MAPK inhibitors (MAPKi) through extracellular matrix (ECM) remodeling activities. Here we show that the anti-fibrotic drug Nintedanib/BIBF1120 is active to normalize the fibrous ECM network, enhance the efficacy of MAPK-targeted therapy and delay tumor relapse in a pre-clinical model of melanoma. We also uncovered the molecular networks that regulate the acquisition of this resistant phenotype and its reversion by Nintedanib, pointing the miR-143/-145 pro-fibrotic cluster as a driver of the therapy-resistant mesenchymal-like phenotype. Upregulation of the miR-143/-145 cluster under BRAFi/MAPKi therapy was observed in melanoma cells in vitro and in vivo and was associated with an invasive/undifferentiated profile of resistant cells. The 2 mature miRNAs generated from this cluster, miR-143-3p and miR-145-5p collaborated to mediate phenotypic transition towards a drug resistant undifferentiated mesenchymal-like state by targeting Fascin actin-bundling protein 1 (FSCN1), modulating the dynamic crosstalk between the actin cytoskeleton and the ECM through the regulation of focal adhesion dynamics as well as contributing to a fine-tuning of mechanotransduction pathways. Our study brings insights into a novel miRNA-mediated regulatory network that contributes to non-genetic adaptive drug resistance and provides proof-of-principle that preventing MAPKi-induced pro-fibrotic stromal response is a viable therapeutic opportunity for patients on targeted therapy.

cancer biology↗

bioRxiv2021

Invasive dedifferentiated melanoma cells inhibit JAK1-STAT3-driven actomyosin contractility of human fibroblastic reticular cells of the lymph node

Fibroblastic reticular cells (FRC) are immunologically specialized fibroblasts controlling the size and microarchitecture of the lymph node (LN), partly through their contractile properties. Swelling is a hallmark of tumor-draining LN in lymphophilic cancers such as cutaneous melanoma, a very aggressive and heterogeneous tumor with high risk of early metastasis. Melanoma cells can dynamically switch between melanocytic proliferative and dedifferentiated mesenchymal-like invasive phenotypes, which are characterized by distinct transcriptional signatures. Melanoma secreted cues, such as extracellular vesicles, growth factors or proinflammatory cytokines, promote LN stroma remodeling and metastatic spreading. But how FRC integrate these pro-metastatic signals and modulate their contractile functions remains poorly characterized. Here, we show that factors secreted by dedifferentiated melanoma cells, but not by melanocytic cells, strongly inhibit FRC actomyosin-dependent contractile forces by decreasing the activity of the RHOA-ROCK pathway and the mechano-responsive transcriptional co-activator YAP, leading to a decrease in F-actin stress fibers and cell elongation. Transcriptional profiling and biochemical analyses indicate that FRC actomyosin cytoskeleton relaxation is driven by inhibition of JAK1 and its downstream transcription factor STAT3, and is associated with increased FRC proliferation and activation. Interestingly, dedifferentiated melanoma cells reduce FRC contractility in vitro independently of extracellular vesicle secretion. These data show that FRC are specifically modulated by proteins secreted by invasive dedifferentiated melanoma cells and suggest that melanoma-derived cues could modulate the biomechanical properties of distant LN before metastatic invasion. They also highlight that JAK1-STAT3 and YAP signaling pathways contribute to the maintenance of the spontaneous contractility of resting human FRC.

cancer biology↗

bioRxiv2019

Targeting DDR1 and DDR2 overcomes matrix-mediated melanoma cell adaptation to BRAF-targeted therapy

Resistance to BRAF and MEK inhibitors in BRAFV600E mutant melanomas remains a major obstacle that limits patient benefit. Microenvironment components including the extracellular matrix (ECM) can support tumor cell adaptation and tolerance to targeted therapies, however the underlying mechanisms remain poorly understood. Here, we investigated the process of matrix-mediated drug resistance (MM-DR) in response to BRAF inhibition in melanoma. We demonstrate that physical and structural cues from fibroblast-derived ECM abrogate anti-proliferative responses to BRAF/MEK inhibition. MM-DR is mediated by the drug-induced clustering of DDR1 and DDR2, two tyrosine kinase collagen receptors. Genetic depletion and pharmacological inhibition of DDR1 and DDR2 overcome ECM-mediated resistance to BRAF inhibition. In melanoma xenografts, targeting DDRs by Imatinib enhances BRAF inhibitor efficacy, counteracts drug-induced collagen remodeling and delays tumor relapse. Mechanistically, DDR-mediated MM-DR fosters a targetable pro-survival NIK/IKK/NF-{kappa}B2 pathway. Our study reveals a novel role of collagen-rich matrix and DDRs in tumor cell adaptation and therapy resistance, thus providing important insights into environment-mediated drug resistance and a pre-clinical rationale for targeting DDR1/2 signaling in combination with BRAF-targeted therapy in melanoma.

cancer biology↗

bioRxiv2019

Electrochemical Roughening and Carbon Nanotube Coating of Tetrodes for Chronic Single-Unit Recording

Recording from single neurons in the brain for long periods of time has been a central goal in both basic neuroscience and translational neurology, in order to understand mechanisms underlying brain processes such as learning and to understand the pathogenesis of neurodynamic disease states 1. Recent advances in materials engineering, digital signal acquisition, and analysis algorithms have brought us closer to achieving this goal, and the possibility has gathered much public attention 2,3. However, it remains a challenge to record from the same units for weeks to months. Here, we record many high-quality tetrode neuronal signals reliably over long periods of time in both deep and superficial areas of the brain. We achieve this by combining electrochemical roughening and carbon nanotube coating of a flexible platinum/iridium substrate, with materials, packaging, and insertion optimized to minimize tip movement with brain pulsation. This \"Magdeburger\" probe enables recordings with long-term signal stability and high signal-to-noise ratio at a reasonable cost in both rodent brains and in substantially larger primate brains. Robust tetrode tracking of identified neurons over longer time periods, in multiple independently targeted areas of the brain, will allow fundamental advances in the study of cognitive learning, aging, and pathogenesis, and opens new possibilities for brain interfaces in humans.

bioengineering↗