Department of Pharmaceutical Sciences, College of Pharmacy, Oregon State University, Corvallis, OR 97331, USA.
MRC Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.
Mar Drugs. 2018 Mar 1;16(3):77. doi: 10.3390/md16030077.
Our understanding of autophagy and lysosomal function has been greatly enhanced by the discovery of natural product structures that can serve as chemical probes to reveal new patterns of signal transduction in cells. Coibamide A is a cytotoxic marine natural product that induces mTOR-independent autophagy as an adaptive stress response that precedes cell death. Autophagy-related (ATG) protein 5 (ATG5) is required for coibamide-induced autophagy but not required for coibamide-induced apoptosis. Using wild-type and autophagy-deficient mouse embryonic fibroblasts (MEFs) we demonstrate that coibamide-induced toxicity is delayed in ATG5 cells relative to ATG5 cells. Time-dependent changes in annexin V staining, membrane integrity, metabolic capacity and caspase activation indicated that MEFs with a functional autophagy pathway are more sensitive to coibamide A. This pattern could be distinguished from autophagy modulators that induce acute ER stress (thapsigargin, tunicamycin), ATP depletion (oligomycin A) or mTORC1 inhibition (rapamycin), but was shared with the Sec61 inhibitor apratoxin A. Coibamide- or apratoxin-induced cell stress was further distinguished from the action of thapsigargin by a pattern of early LC3-II accumulation in the absence of CHOP or BiP expression. Time-dependent changes in ATG5-ATG12, PARP1 and caspase-3 expression patterns were consistent with the conversion of ATG5 to a pro-death signal in response to both compounds.
天然产物结构的发现极大地增强了我们对自噬和溶酶体功能的理解，这些结构可以作为化学探针，揭示细胞中信号转导的新模式。Coibamide A 是一种细胞毒性海洋天然产物，可诱导 mTOR 非依赖性自噬作为细胞死亡前的适应性应激反应。自噬相关蛋白 5 (ATG5) 是 Coibamide 诱导自噬所必需的，但不是 Coibamide 诱导凋亡所必需的。利用野生型和自噬缺陷型小鼠胚胎成纤维细胞 (MEFs)，我们证明 Coibamide 诱导的毒性在 ATG5 细胞中比 ATG5 细胞延迟。 Annexin V 染色、膜完整性、代谢能力和半胱天冬酶激活的时程变化表明，具有功能性自噬途径的 MEFs 对 Coibamide A 更敏感。这种模式可以与诱导急性内质网应激（他普西龙、衣霉素）、ATP 耗竭（寡霉素 A）或 mTORC1 抑制（雷帕霉素）的自噬调节剂区分开来，但与 Sec61 抑制剂 apratoxin A 相同。Coibamide 或 apratoxin 诱导的细胞应激与 thapsigargin 的作用进一步区分开来，因为在没有 CHOP 或 BiP 表达的情况下，LC3-II 早期积累。ATG5-ATG12、PARP1 和 caspase-3 表达模式的时程变化与 ATG5 对这两种化合物的反应转化为促死亡信号一致。
