Department of Psychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
Department of Psychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
J Biol Chem. 2021 Oct;297(4):101120. doi: 10.1016/j.jbc.2021.101120. Epub 2021 Aug 25.
GGGGCC (GC) repeat expansion in the C9orf72 gene has been shown to cause frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Dipeptide repeat proteins produced through repeat-associated non-AUG (RAN) translation are recognized as potential drivers for neurodegeneration. Therefore, selective inhibition of RAN translation could be a therapeutic avenue to treat these neurodegenerative diseases. It was previously known that the porphyrin TMPyP4 binds to GC repeat RNA. However, the consequences of this interaction have not been well characterized. Here, we confirmed that TMPyP4 inhibits C9orf72 GC repeat translation in cellular and in in vitro translation systems. An artificial insertion of an AUG codon failed to cancel the translation inhibition, suggesting that TMPyP4 acts downstream of non-AUG translation initiation. Polysome profiling assays also revealed polysome retention on GC repeat RNA, along with inhibition of translation, indicating that elongating ribosomes stall on GC repeat RNA. Urea-resistant interaction between GC repeat RNA and TMPyP4 likely contributes to this ribosome stalling and thus to selective inhibition of RAN translation. Taken together, our data reveal a novel mode of action of TMPyP4 as an inhibitor of GC repeat translation elongation.
C9orf72 基因中的 GGGGCC (GC) 重复扩展已被证明会导致额颞叶变性和肌萎缩性侧索硬化症。通过重复相关的非 AUG (RAN) 翻译产生的二肽重复蛋白被认为是神经退行性变的潜在驱动因素。因此，选择性抑制 RAN 翻译可能是治疗这些神经退行性疾病的一种治疗途径。先前已知卟啉 TMPyP4 与 GC 重复 RNA 结合。然而，这种相互作用的后果尚未得到很好的描述。在这里，我们证实 TMPyP4 抑制细胞内和体外翻译系统中的 C9orf72 GC 重复翻译。插入一个 AUG 密码子的人工操作未能取消翻译抑制，表明 TMPyP4 作用于非 AUG 翻译起始的下游。多核糖体分析还显示多核糖体在 GC 重复 RNA 上的保留以及翻译的抑制，表明延伸的核糖体在 GC 重复 RNA 上停滞。尿素抗性 GC 重复 RNA 和 TMPyP4 之间的相互作用可能导致这种核糖体停滞，从而选择性抑制 RAN 翻译。总之，我们的数据揭示了 TMPyP4 作为 GC 重复翻译延伸抑制剂的一种新作用模式。
