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Abstract: High-content screening offers great potential for in vitro toxicology, yet its application in environmental test systems remains limited by the lack of open, standardized workflows and biologically interpretable analysis frameworks. Here, we established and optimized a reproducible pipeline for image-based phenotypic profiling of rainbow trout gill cells (RTgill-W1) exposed to environmental chemicals, using Tebuthiuron as a case study. The workflow integrates multiple organelle dyes and systematically evaluates critical data processing steps (quality control, standardization, outlier removal, concentration–response-guided feature selection, and dimensionality reduction) to identify robust, biologically meaningful end points. Our systematic comparison demonstrated remarkable robustness: even minimal processing detected concentration–response relationships, while optimized standardization and feature selection substantially improved signal clarity. The framework identified 94 morphological features capturing Tebuthiuron’s concentration- and time-dependent cellular responses, with lysosomal redistribution and mitochondrial fragmentation emerging as early stress indicators. Maximum mean discrepancy and Uniform Manifold Approximation and Projection (UMAP) visualizations provided biologically interpretable maps of phenotypic divergence. This proof-of-concept demonstrates that our quality-controlled, transparent HCS pipeline can resolve concentration- and time-dependent subcellular responses in fish cells, establishing a methodological basis for future multichemical phenotypic profiling.
Keywords: high-content screening, phenotyping, toxicology, fishcells, concentration−response, ecotoxicology
Published in DiRROS: 11.08.2026; Views: 218; Downloads: 263
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Abstract: Functional genomics offers a powerful, unbiased approach to elucidating the molecular mechanisms underlying the toxicity of environmental pollutants. In this study, we applied genome-wide screening in Chlamydomonas reinhardtii to investigate two classical herbicides: paraquat and diuron. Our screen successfully uncovered critical nuclear-encoded pathways that govern susceptibility. For both herbicides, we identified genes regulating the assembly and maintenance of the photosynthetic machinery, highlighting the central role of nuclear control over these chloroplast-localized targets. Beyond these target-related factors, we discovered novel nontarget-site resistance mechanisms. For paraquat, we identified intracellular trafficking as the central determinant of toxicity, experimentally characterizing a P5B ATPase transporter and a fatty acid elongation pathway whose disruption, we propose, converges on the same endomembrane delivery route through sphingolipid depletion. In contrast, our screening data suggest that diuron tolerance may be associated with a metabolic strategy focused on energy conservation, where the inactivation of specific NADPH-consuming enzymes could preserve reducing power for essential antioxidant defense. Collectively, these findings demonstrate that functional genomics can reveal novel, complex modes of action even for well-characterized chemicals, providing the mechanistic resolution required to advance modern ecotoxicological risk assessments.
Keywords: paraquat, diuron
Published in DiRROS: 09.06.2026; Views: 321; Downloads: 363
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Keywords: odprta znanost, občanska znanost, raziskovalni podatki, podatkovni svetovalec, repozitoriji, projekt SPOZNAJ
Published in DiRROS: 06.11.2025; Views: 1123; Downloads: 1139
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Abstract: We performed detailed spatiotemporal analysis of chloroplast redox response to potato virus Y (PVY) infection in resistant Ny‐1-gene-bearing potato and its transgenic counterpart with impaired SA accumulation and compromised resistance. We found that the chloroplasts are highly oxidized in the cells adjacent to the cell death zone at different stages after virus inoculation in both genotypes. This hypothesis is further supported by highly induced formation of stroma filled tubules that extend from chloroplasts (stromules) in the cells adjacent to signalling cells. This dataset s a deposit of all the raw microscopy images of the study, plus the relevant metadata in ISA-tab compliant folder structure. After receiving reviews, we have made an additional experiment using a ROS inhibitor. The raw and processed data for this is in a separate file: _S_chlROS_inhibitor.zip
Keywords: cell signalling, chloroplasts, microscopy, ISA-tab, potato virus Y, redox responses, cell death, spatiotemporal analysis
Published in DiRROS: 24.10.2025; Views: 961; Downloads: 634
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Keywords: podatki, ravnanje s podatki, ravnanje z raziskovalnimi podatki, raziskovalni podatki, odprta znanost, programska orodja, Python, R
Published in DiRROS: 26.05.2025; Views: 1291; Downloads: 1119
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