Department of Biotechnology, The University of Tokyo, Tokyo, Japan.
Collaborative Research Institute for Innovative Microbiology, The University of Tokyo, Bunkyo-ku, Tokyo, Japan.
Biosci Biotechnol Biochem. 2024 Oct 22;88(11):1279-1288. doi: 10.1093/bbb/zbae122.
Fungal biosynthetic gene clusters often include genes encoding transmembrane proteins, which have been mostly thought to be transporters exporting the products. However, there is little knowledge about subcellular compartmentalization of transmembrane proteins essential for biosynthesis. Fungal mycotoxin cyclochlorotine is synthesized by non-ribosomal peptide synthetase, which is followed by modifications with three transmembrane UstYa-family proteins. Heterologous expression in Aspergillus oryzae revealed that total biosynthesis of cyclochlorotine requires additional two transporter proteins. Here, we investigated subcellular localizations of the five transmembrane proteins under heterologous expression in A. oryzae. Enhanced green fluorescent protein (EGFP) fusions to the transmembrane proteins, which were confirmed to normally function in cyclochlorotine production, were expressed together with organellar markers. All the transmembrane proteins exhibited localizations commonly in line of the trans-Golgi, endosomes, and vacuoles. This study suggests that subcellular compartmentalization of UstYa family proteins and transporters allows corporative functions of delivering intermediates and subsequent modifications, completing cyclochlorotine biosynthesis.
真菌生物合成基因簇通常包含编码跨膜蛋白的基因，这些基因大多被认为是输出产物的转运蛋白。然而，对于生物合成必需的跨膜蛋白的亚细胞区室化知之甚少。真菌霉菌毒素环氯毒素是由非核糖体肽合成酶合成的，然后由三个跨膜 UstYa 家族蛋白进行修饰。在米曲霉中的异源表达表明，环氯毒素的总生物合成需要另外两种转运蛋白。在这里，我们研究了在米曲霉中异源表达的五跨膜蛋白的亚细胞定位。与跨膜蛋白融合的增强型绿色荧光蛋白（EGFP），经证实可正常参与环氯毒素的产生，与细胞器标记物一起表达。所有跨膜蛋白均表现出通常在高尔基体内、内体和液泡中的共定位。这项研究表明，UstYa 家族蛋白和转运蛋白的亚细胞区室化允许传递中间产物和后续修饰的协同作用，完成环氯毒素的生物合成。
