AbGATA6转录因子在ATP调控CCP延缓双孢蘑菇衰老中的作用机制

Owing to the rapid deterioration and susceptible to senescence of postharvest white mushroom (Agaricus bisporus), to study the senescent mechanism and how to regulate it have become one of the most urgent problems to be solved in mushroom industry. Insufficient ATP is the key factor inducing the senescence of white mushroom, and cytochrome pathway (CCP) is one of the main pathways for ATP generation. Therefore, it is quite critical to study the CCP metabolism and its regulation. We found that exogenous ATP delayed the senescent process of white mushroom and induced the expression of AbGATA6 transcription factor (TF) which is related to CCP, but the mechanism is still unclear. In this project, the ATP will be utilized to treat white mushrooms, and the regulation of AbGATA6 on CCP will be studied. The phylogenetic tree will be constructed through bioinformatics analysis. Whether the AbGATA6 protein is localized exclusively in the nucleus will be determined by transcriptional activity analysis. The interactive relationship between AbGATA6 TF to genes related to key enzymes of CCP regulated by ATP will be confirmed. The role of AbGATA6 in ATP regulating CCP to delay the senescent process of white mushrooms will be elucidated, which will provide new insights into transcriptional regulation of delaying senescence of white mushroom. It is significant to accelerate industrialization development and to delay the senescent process of white mushroom by regulating respiration pathway and energy metabolism.

采后双孢菇品质劣变快、易衰老，深入研究双孢菇衰老机制并对其进行调控是蘑菇产业亟待解决的问题之一。ATP不足是引起双孢菇衰老的关键因素，细胞色素途径（CCP）是ATP最主要的来源途径之一，因此，研究CCP及其调控至关重要。申请人前期研究表明外源ATP延缓衰老同时诱导了与CCP途径相关的转录因子AbGATA6的表达，但其作用机制尚未阐明。本项目拟采用外源ATP处理采后双孢菇，研究AbGATA6对CCP的调控；通过生物信息学分析构建AbGATA进化树，通过亚细胞定位分析明确AbGATA6是否位于细胞核；采用双荧光素酶瞬时表达系统、凝胶阻滞分析技术分析AbGATA6与ATP调控CCP关键酶相关基因的互作关系，阐明AbGATA6在ATP调控CCP延缓双孢菇衰老过程中的作用。为抑制双孢菇衰老的转录调控提供了新的视角，对通过呼吸途径、能量代谢的调控来抑制衰老，加快双孢菇产业化发具有重要意义。

采后双孢菇品质劣变快、极易衰老，深入研究双孢菇衰老机制并对其进行调控是蘑菇产业亟待解决的问题之一。ATP不足是引起双孢菇衰老的关键因素，细胞色素途径（CCP）是ATP最主要的来源途径之一，因此，研究CCP及其调控至关重要。本项目研究了转录因子AbGATA6在ATP调控CCP延缓双孢菇衰老过程中的作用机制。研究了外源ATP处理双孢菇子实体的CCP途径及其关键酶活性变化，结果表明外源ATP处理可显著抑制双孢菇衰老过程中CCP的下降；CCP途径中的关键酶细胞色素氧化酶（COX）和 ATP合成酶（ATPase）活性显著高于对照组，结合编码关键酶相关基因表达情况，筛选出ATP作用CCP的调控关键酶COX以及编码该酶的基因AbCOX5。利用转录组测序（RNA-seq）技术筛选ATP处理双孢菇中差异表达最为显著的转录因子GATA。亚细胞定位实验结果显示转录因子AbGATA6定位在细胞核，烟草体内转录活性分析实验表明AbGATA6具有转录激活活性。利用凝胶阻滞（EMSA）、双荧光素酶报告基因实验结果表明AbGATA6直接结合AbCOX5基因的启动子。以上结果表明转录因子AbGATA6通过转录激活AbCOX5表达调控CCP途径。该研究为抑制双孢菇衰老的转录调控提供新的视角，也为果蔬采后呼吸代谢的调控奠定理论基础。