Table_8_Transcriptome Profiling and Cytological Assessments for Identifying Regulatory Pathways Associated With Diorcinol N-Induced Autophagy in A3 Cells.xlsx

Fungal secondary metabolites serve as a rich resource for exploring lead compounds with medicinal importance. Diorcinol N (DN), a fungal secondary metabolite isolated from an endophytic fungus, Arthrinium arundinis, exhibits robust anticancer activity. However, the anticancer mechanism of DN remains unclear. In this study, we examined the growth-inhibitory effect of DN on different human cancer cell lines. We found that DN decreased the viability of A3 T-cell leukemia cells in a time- and concentration-dependent manner. Transcriptome analysis indicated that DN modulated the transcriptome of A3 cells. In total, 9,340 differentially expressed genes were found, among which 4,378 downregulated genes and 4,962 upregulated genes were mainly involved in autophagy, cell cycle, and DNA replication. Furthermore, we demonstrated that DN induced autophagy, cell cycle arrest in the G1/S phase, and downregulated the expression of autophagy- and cell cycle-related genes in A3 cells. By labeling A3 cells with acridine orange/ethidium bromide, Hoechst 33,258, and monodansylcadaverine and via transmission electron microscopy, we found that DN increased plasma membrane permeability, structural disorganization, vacuolation, and autophagosome formation. Our study provides evidence for the mechanism of anticancer activity of DN in T-cell leukemia (A3) cells and demonstrates the promise of DN as a lead or even candidate molecule for the treatment of acute lymphoblastic leukemia.

真菌次级代谢产物是挖掘药用先导化合物的宝贵资源。二氧丝菌素N（Diorcinol N，DN）是从内生真菌芦苇节菱孢菌（Arthrinium arundinis）中分离得到的一种真菌次级代谢产物，其具备强效的抗肿瘤活性。然而，DN的抗肿瘤分子机制迄今尚未阐明。本研究考察了DN对多株人体癌细胞系的生长抑制作用，发现DN可通过时间与浓度依赖的方式降低A3 T细胞白血病细胞的存活率。转录组学分析结果表明，DN可调控A3细胞的转录组表达谱，共计筛选得到9340个差异表达基因，其中4378个下调基因与4962个上调基因主要富集于自噬、细胞周期及DNA复制通路。进一步实验证实，DN可诱导A3细胞发生自噬、阻滞细胞周期于G1/S期，并下调自噬与细胞周期相关基因的表达水平。通过吖啶橙/溴化乙锭（acridine orange/ethidium bromide）、Hoechst 33258及单丹磺酰尸胺（monodansylcadaverine）对A3细胞进行染色，并结合透射电子显微镜（transmission electron microscopy）观察，本研究发现DN可升高细胞膜通透性、引发细胞膜结构紊乱、诱导细胞空泡化及自噬体形成。本研究阐明了DN在T细胞白血病（A3）细胞中的抗肿瘤作用机制，证实了DN作为急性淋巴细胞白血病治疗用先导化合物乃至候选药物的开发潜力。