香菇菌丝后熟转色形成的活性氧作用与自噬特征

为了解活性氧(reactive oxygen species,ROS)在香菇菌丝后熟转色形成中的作用及其自噬细胞学特征,以香菇工厂化菌株KS11为研究材料,分析其在菌丝后熟转色过程中4个时间点(30、45、60、75 d)的活性氧含量(ROS)、丙二醛(MDA)含量、NADPH氧化酶浓度、抗氧化酶活性以及外源活性氧和DPI对其影响的表型试验,利用透射电镜观察该过程菌丝细胞自噬特征变化,并运用实时荧光定量PCR对自噬基因Atg8的表达水平进行比较分析。结果表明：(1) H₂O₂作为主要的活性氧因子在菌丝后熟转色形成中呈现显著动态变化,后熟转色过程中不断升高,并在转色中第60天呈高峰值。(2) NADPH氧化酶浓度与H₂O₂含量变化呈紧密正相关。(3)外源施加一定浓度H₂O₂显著促进香菇菌丝后熟转色,且DPI作为NADPH氧化酶抑制剂显著抑制了香菇菌丝后熟转色的发生。(4)香菇菌丝后熟转色过程中,细胞自噬特征逐渐增强,并在转色中后期最显著。上述结果表明以H₂O₂为主的活性氧在香菇菌丝后熟转色过程中起重要作用,并且可能参与了香菇菌丝后熟转色过程中的自噬进程。

Effects of reactive oxygen species and autophagy characteristics of Lentinula edodes during postripeness of mycelia

In order to understand the role of reactive oxygen species (ROS) and autophagy cytological characteristics during brown film (BF) formation and postripeness of mycelium in Lentinula edodes, the content of reactive oxygen species (ROS), MDA content, and the concentration of NADPH oxidase at four time points (d30, d45, d60, d75) were analyzed. The activity of antioxidant enzymes, the phenotype of exogenous ractive oxygen species, and DPI were tested and the autophagy characteristics were analyzed by transmission electron microscopy. The expression level of autophagy gene Atg8 was compared and analyzed by real-time fluorescence quantitative PCR. The results showed that H₂O₂ as the main reactive oxygen species showed dynamic changes in BF formation and postripeness process, and increased continuously and reached high peak value in 60 days during postripeness and BF formation. The concentration of NADPH oxidase was closely related to the change of H₂O₂ content. The application of H₂O₂ could promote significantly the BF formation and ripeness of mycelium, and DPI as NADPH oxidase inhibitor could significantly inhibit the postripeness and BF formation. The autophagy characteristics was enhanced gradually during BF formation and postripeness process, and showed most significant in the middle and late stage. The results proved that H₂O₂ played an important role in the process of postripeness and BF formation, and it might be involved in the autophagy process during BF formation and postripeness in Lentinula edodes.