Tokyo University of Agriculture, 1-1-1 Sakuragaoka, Setagaya-ku, Tokyo 156-8502, Japan.
Noda Institute for Scientific Research, 399 Noda, Noda, Chiba 278-0037, Japan.
Fungal Genet Biol. 2019 Jun;127:35-44. doi: 10.1016/j.fgb.2019.02.004. Epub 2019 Feb 18.
Aspergillus oryzae is an important microorganism in the bio- and food industries; therefore, understanding the mechanism underlying its secondary metabolism regulation is important for ensuring its safe use. Here, we screened a novel Zn(II)-Cys-type protein-encoding gene, AO090003001186, designated as kpeA (kojic acid production enhancement A), from an A. oryzae disruption mutant library of transcriptional regulators. kpeA is highly conserved among filamentous fungi and encodes a protein with Zn(II)-Cys motif located in the middle of the sequence. Phylogenetic analysis revealed that KpeA was classified into a distal group compared to other fungal Zn(II)-Cys-type transcriptional regulators. A Cys to Ala substitution mutant of KpeA showed identical phenotype to the kpeA disruption strain, confirming that KpeA is novel type Zn(II)-Cys binding protein. Colonies of the kpeA disruption strain (ΔkpeA) had longer aerial hyphae and showed decreased conidia production. Microscopic analysis suggested that the reduced vesicle size and conidial head formation in ΔkpeA strain account for the decreased conidia production. Transcriptional levels of brlA and downstream abaA and wetA were decreased in ΔkpeA strain. Moreover, ΔkpeA strain produced 6-fold more kojic acid than the control strains, and the expression of kojR and kojA was increased in ΔkpeA strain. Therefore, KpeA is a novel Zn(II)-Cys-type protein likely involved in conidiation and kojic acid production at the transcriptional level.
米曲霉是生物和食品工业中的一种重要微生物；因此，了解其次级代谢调控的机制对于确保其安全使用非常重要。在这里，我们从米曲霉转录调控因子敲除突变体库中筛选到一个新型 Zn(II)-Cys 型蛋白编码基因，命名为 kpeA（曲酸产量增强 A）。kpeA 在丝状真菌中高度保守，编码的蛋白序列中间具有 Zn(II)-Cys 基序。系统发育分析表明，与其他真菌 Zn(II)-Cys 型转录调控因子相比，KpeA 被分类为远群。KpeA 的 Cys 到 Ala 取代突变体表现出与 kpeA 敲除菌株相同的表型，证实 KpeA 是一种新型的 Zn(II)-Cys 结合蛋白。kpeA 敲除菌株（ΔkpeA）的菌落具有更长的气生菌丝，产孢量减少。显微镜分析表明，ΔkpeA 菌株中囊泡体积减小和分生孢子头部形成减少是产孢量减少的原因。ΔkpeA 菌株中 brlA 及其下游 abaA 和 wetA 的转录水平降低。此外，与对照菌株相比，ΔkpeA 菌株产生了 6 倍的曲酸，并且 ΔkpeA 菌株中 kojR 和 kojA 的表达增加。因此，KpeA 是一种新型的 Zn(II)-Cys 型蛋白，可能参与了孢子形成和转录水平上的曲酸生产。
