Department Materials and the Environment, Bundesanstalt für Materialforschung und -prüfung (BAM), Berlin, Germany.
Department of Ecological and Biological Sciences (DEB), Università degli Studi della Tuscia, Viterbo, Italy.
IUBMB Life. 2024 Dec;76(12):1072-1090. doi: 10.1002/iub.2895. Epub 2024 Jul 16.
Cryomyces antarcticus, a melanized cryptoendolithic fungus endemic to Antarctica, can tolerate environmental conditions as severe as those in space. Particularly, its ability to withstand ionizing radiation has been attributed to the presence of thick and highly melanized cell walls, which-according to a previous investigation-may contain both 1,8-dihydroxynaphthalene (DHN) and L-3,4 dihydroxyphenylalanine (L-DOPA) melanin. The genes putatively involved in the synthesis of DHN melanin were identified in the genome of C. antarcticus. Most important is capks1 encoding a non-reducing polyketide synthase (PKS) and being the ortholog of the functionally characterized kppks1 from the rock-inhabiting fungus Knufia petricola. The co-expression of CaPKS1 or KpPKS1 with a 4'-phosphopantetheinyl transferase in Saccharomyces cerevisiae resulted in the formation of a yellowish pigment, suggesting that CaPKS1 is the enzyme providing the precursor for DHN melanin. To dissect the composition and function of the melanin layer in the outer cell wall of C. antarcticus, non-melanized mutants were generated by CRISPR/Cas9-mediated genome editing. Notwithstanding its slow growth (up to months), three independent non-melanized Δcapks1 mutants were obtained. The mutants exhibited growth similar to the wild type and a light pinkish pigmentation, which is presumably due to carotenoids. Interestingly, visible light had an adverse effect on growth of both melanized wild-type and non-melanized Δcapks1 strains. Further evidence that light can pass the melanized cell walls derives from a mutant expressing a H2B-GFP fusion protein, which can be detected by fluorescence microscopy. In conclusion, the study reports on the first genetic manipulation of C. antarcticus, resulting in non-melanized mutants and demonstrating that the melanin is rather of the DHN type. These mutants will allow to elucidate the relevance of melanization for surviving extreme conditions found in the natural habitat as well as in space.
南极冰藻(Cryomyces antarcticus)是一种黑色素 cryptoendolithic 真菌，仅存在于南极洲，能耐受类似于太空的极端环境条件。特别是，其耐受电离辐射的能力归因于厚而高度黑色素化的细胞壁的存在，根据先前的研究，这种细胞壁可能含有 1,8-二羟基萘（DHN）和 L-3,4 二羟基苯丙氨酸（L-DOPA）黑色素。在南极冰藻的基因组中鉴定出了参与 DHN 黑色素合成的假定基因。最重要的是 capks1 基因，它编码一种非还原型聚酮合酶（PKS），是来自岩石栖息地真菌 Knufia petricola 的功能表征的 kppks1 的同源物。CaPKS1 或 KpPKS1 与 4'-磷酸泛酰巯基乙胺转移酶在酿酒酵母中的共表达导致形成黄色色素，表明 CaPKS1 是提供 DHN 黑色素前体的酶。为了剖析南极冰藻外壁黑色素层的组成和功能，通过 CRISPR/Cas9 介导的基因组编辑生成了非黑色素突变体。尽管其生长缓慢（长达数月），但获得了三个独立的非黑色素化 Δcapks1 突变体。突变体表现出与野生型相似的生长速度和浅粉红色着色，这可能归因于类胡萝卜素。有趣的是，可见光对黑色素化的野生型和非黑色素化 Δcapks1 菌株的生长都有不利影响。进一步的证据表明，光可以穿透黑色素化的细胞壁，这源于表达 H2B-GFP 融合蛋白的突变体，该融合蛋白可以通过荧光显微镜检测到。总之，该研究报告了对南极冰藻的首次遗传操作，产生了非黑色素突变体，并证明黑色素主要是 DHN 类型。这些突变体将允许阐明黑色素化对于在自然栖息地和太空中发现的极端条件下生存的相关性。
