TY - CONF A1 - Schumacher, Julia T1 - The roles of DHN melanin and the stress-activated MAP kinase in the rock inhabitant Knufia petricola N2 - Black fungi/yeasts exhibit high stress tolerance, yeast‑like or meristematic growth, and constitutive 1,8-dihydroxynaphthalene (DHN) melanin formation. Due to their slow growth, robust cell walls and the lack of sexual cycles and genetic tools, the underlying mechanisms of their phenotypic traits have remained largely unexplored. Using recently developed genetic tools, it is now possible to manipulate the genome of the rock-inhabiting model fungus Knufia petricola. Thus, gene functions and the cell biology of black fungi can be studied using CRISPR/Cas9-based genome editing and live-cell imaging with genetically encoded fluorescent proteins. Here, we are addressing the question to which extent constitutive pigment formation (melanin and carotenoids) and responses mediated by the stress-activated mitogen-activated protein (MAP) kinase contribute to the observed extremotolerance of K. petricola. The mutations of pks1, phs1 and both genes result in melanin-free (pink), carotenoid-free (black) and pigment-free (white) strains, respectively. The other putative melanogenic genes were identified in the genome, deleted to confirm their involvement in DHN melanogenesis and co-expressed in Saccharomyces cerevisiae for reconstruction of the synthesis pathway. Sak1 encoding the stress-activated MAP kinase was deleted in the wild-type and different pigment-deficient backgrounds. Growth of the obtained single, double and triple deletion mutants was tested by droplet tests on media supplemented with different stress-inducing agents. The Δsak1 mutants show slightly reduced growth rates even without environmental pressure and are hypersensitive to different stresses: e.g. osmotic, oxidative, membrane, pH and heat stress. Melanin-free Δsak1 mutants are more sensitive than black Δsak1 mutants to some but not all stress conditions, suggesting that melanin and the SAK1 pathway have complementary roles in protecting K. petricola from stress. T2 - 16th European Conference on Fungal Genetics CY - Innsbruck, Austria DA - 05.03.2023 KW - Black fungus KW - Extremotolerance KW - Pigments PY - 2023 AN - OPUS4-57147 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schumacher, Julia T1 - Fungi in extreme habitats: Lessons from the microcolonial black fungus Knufia petricola N2 - Fungi that share light-flooded habitats with phototrophs may profit from their excess photosynthetic products. But to cope with sunlight-associated stresses [e.g. high temperatures, UV radiation with associated DNA damage, accumulation of reactive oxygen species (ROS), desiccation and osmotic stresses] it is important for fungi to accurately sense and respond to changes in light. The genomes of black [dihydroxynaphthalene (DHN) melanin-containing] fungi from phyllosphere and exposed solid surfaces contain multiple photoreceptors (PRs). The plant pathogen Botrytis cinerea (Leotiomycetes) has a highly sophisticated photosensory and signalling system that helps to avoid light and to locate susceptible hosts. Rock-inhabiting Dothideomycetes and Eurotiomycetes including Knufia petricola possess equal numbers of PRs along with the same set of protective pigments. This similarity between black fungi from plant and rock surfaces suggests that photoperception and -regulation are important for fungi that receive nutrients through cooperation with phototrophs. T2 - Gordon Research Conference "Cellular and Molecular Fungal Biology" CY - Holderness, NH, USA DA - 26.06.2022 KW - Light KW - Stress KW - Photoperception KW - Pigments PY - 2022 AN - OPUS4-55249 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schumacher, Julia T1 - Unlocking black fungi: from genomes to material conservation and biotechnological innovation N2 - Black fungi are a polyphyletic group of melanized ascomycetes adapted to extreme environments such as deserts, rocks, and human-made surfaces. Their slow compact growth, and thick melanized cell walls confer exceptional stress tolerance but have long hindered molecular studies and genetic engineering. Consequently, the biology and biotechnological potential of these organisms remained largely unexplored. To overcome these limitations, we selected the rock-inhabiting black fungus Knufia petricola as a model. Its genome sequence (12 contigs, ~10,000 genes) enabled the development of a versatile genetic toolbox based on CRISPR/Cas9-mediated editing, achieving up to 100% homologous recombination and supporting multiplex editing of at least six genomic regions. Color-based selection systems targeting pigment genes enable rapid identification of strains with correctly integrated expression cassettes, including simultaneous insertion of optimized genes coding for blue, green, and red fluorescent proteins for co-localization and protein interaction studies. These tools enable functional analyses of traits central to extremotolerance, such as the role of melanin in stress protection, biofilm formation, and material interactions. Furthermore, K. petricola has been established as a eukaryotic expression platform: heterologous genes from bacteria, fungi, and plants can be expressed under constitutive or inducible promoters, including the Tet-on system for tunable gene regulation. Pigment-free strains provide access to metabolic precursors, enabling high-level synthesis of alternative secondary metabolites. Importantly, the optimized transformation protocols were successfully transferred to the Antarctic black fungus Cryomyces antarcticus, a cryptoendolithic extremophile and astrobiology model, demonstrating that even the most stress-tolerant fungi can be genetically engineered. Together, the genome sequence and advanced genetic engineering strategies have transformed K. petricola from a challenging extremotolerant organism into a tractable model for studying stress adaptation, material interactions and a promising host for biotechnology and synthetic biology approaches. These achievements, combined with emerging black fungal genomes, inform sustainable material protection strategies to mitigate colonization and biodeterioration of exposed surfaces, linking fundamental research with applied solutions for material conservation and industrial biotechnology. T2 - Annual Conference of the Association for General and Applied Microbiology (VAAM) 2026 CY - Berlin, Germany DA - 22.03.2026 KW - Functional genetics KW - Material colonizer KW - Pigments PY - 2026 AN - OPUS4-65752 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER -