School of Life Sciences, East China Normal University, Shanghai, China.
School of Life Sciences, East China Normal University, Shanghai, China
mBio. 2021 Mar 23;12(2):e00007-21. doi: 10.1128/mBio.00007-21.
In filamentous fungi, 1,8-dihydroxynaphthalene (DHN) melanin is a major component of the extracellular matrix, endowing fungi with environmental tolerance and some pathogenic species with pathogenicity. However, the subcellular location of the melanin biosynthesis pathway components remains obscure. Using the gray mold pathogen , the DHN melanin intermediate scytalone was characterized via phenotypic and chemical analysis of mutants, and the key enzymes participating in melanin synthesis were fused with fluorescent proteins to observe their subcellular localizations. The Δ mutant accumulated scytalone in the culture filtrate rather than in mycelium. Excessive scytalone appears to be self-inhibitory to the fungus, leading to repressed sclerotial germination and sporulation in the Δ mutant. The BcBRN1/2 enzymes responsible for synthesizing scytalone were localized in endosomes and found to be trafficked to the cell surface, accompanied by the accumulation of BcSCD1 proteins in the cell wall. In contrast, the early-stage melanin synthesis enzymes BcPKS12/13 and BcYGH1 were localized in peroxisomes. Taken together, the results of this study revealed the subcellular distribution of melanin biosynthetic enzymes in , indicating that the encapsulation and externalization of the melanin synthetic enzymes need to be delicately orchestrated to ensure enzymatic efficiency and protect itself from the adverse effect of the toxic intermediate metabolite. The devastating gray mold pathogen propagates via melanized conidia and sclerotia. This study reveals that the sclerotial germination of is differentially affected by different enzymes in the melanin synthesis pathway. Using gene knockout mutants and chemical analysis, we found that excessive accumulation of the melanin intermediate scytalone is inhibitory to Subcellular localization analysis of the melanin synthesis enzymes of suggested two-stage partitioning of the melanogenesis pathway: the intracellular stage involves the steps until the intermediate scytalone was translocated to the cell surface, whereas the extracellular stage comprises all the steps occurring in the wall from scytalone to final melanin formation. These strategies make the fungus avert self-poisoning during melanin production. This study opens avenues for better understanding the mechanisms of secondary metabolite production in filamentous fungi.
在丝状真菌中，1,8-二羟基萘（DHN）黑色素是细胞外基质的主要成分，使真菌具有环境耐受性，某些病原真菌具有致病性。然而，黑色素生物合成途径成分的亚细胞定位仍然不清楚。本研究通过灰霉菌突变体的表型和化学分析，对 DHN 黑色素中间产物 3-脱氧-D-甘露酮醛（scytalone）进行了特征描述，并将参与黑色素合成的关键酶与荧光蛋白融合，观察其亚细胞定位。Δ突变体在培养滤液中积累 scytalone，而不是在菌丝体中。过量的 scytalone 似乎对真菌具有自抑制作用，导致 Δ突变体中菌核萌发和产孢受到抑制。负责合成 scytalone 的 BcBRN1/2 酶定位于内体，并发现它们被转运到细胞膜，同时 BcSCD1 蛋白在细胞壁中积累。相比之下，早期黑色素合成酶 BcPKS12/13 和 BcYGH1 定位于过氧化物酶体。综上所述，本研究揭示了 中黑色素生物合成酶的亚细胞分布，表明黑色素合成酶的封装和外化需要精心调控，以确保酶的效率，并防止自身受到有毒中间代谢物的不利影响。破坏性灰霉菌 通过黑色素化的分生孢子和菌核传播。本研究表明，不同黑色素合成途径中的酶对 菌核萌发的影响不同。通过基因敲除突变体和化学分析，我们发现黑色素中间体 scytalone 的过度积累对 具有抑制作用。黑色素合成酶的亚细胞定位分析表明，黑色素生成途径分为两个阶段：细胞内阶段涉及到中间产物 scytalone 转运到细胞膜的步骤，而细胞外阶段包括从 scytalone 到最终黑色素形成过程中发生在细胞壁上的所有步骤。这些策略使真菌在黑色素生成过程中避免自我中毒。本研究为更好地理解丝状真菌次生代谢产物的产生机制开辟了道路。
