Department of Neurology, MassGeneral Institute for Neurodegenerative Disease (MIND), Massachusetts General Hospital and Harvard Medical School, Boston, MA, 02114, USA.
Broad Institute of Harvard University and MIT, Cambridge, MA, 02142, USA.
Nat Commun. 2018 Jan 11;9(1):152. doi: 10.1038/s41467-017-02643-5.
Expansion of GC repeats in the C9ORF72 gene is the most prevalent inherited form of amyotrophic lateral sclerosis and frontotemporal dementia. Expanded transcripts undergo repeat-associated non-AUG (RAN) translation producing dipeptide repeat proteins from all reading frames. We determined cis-factors and trans-factors influencing translation of the human C9ORF72 transcripts. GC translation operates through a 5'-3' cap-dependent scanning mechanism, requiring a CUG codon located upstream of the repeats and an initiator Met-tRNA. Production of poly-GA, poly-GP, and poly-GR proteins from the three frames is influenced by mutation of the same CUG start codon supporting a frameshifting mechanism. RAN translation is also regulated by an upstream open reading frame (uORF) present in mis-spliced C9ORF72 transcripts. Inhibitors of the pre-initiation ribosomal complex and RNA antisense oligonucleotides selectively targeting the 5'-flanking GC sequence block ribosomal scanning and prevent translation. Finally, we identified an unexpected affinity of expanded transcripts for the ribosomal subunits independently from translation.
C9ORF72 基因中 GC 重复序列的扩增是最常见的肌萎缩性侧索硬化症和额颞叶痴呆的遗传性形式。扩增的转录本通过重复相关的非 AUG（RAN）翻译产生来自所有阅读框的二肽重复蛋白。我们确定了影响人类 C9ORF72 转录本翻译的顺式和反式因素。GC 翻译通过 5'-3' 帽依赖性扫描机制进行，需要位于重复序列上游的 CUG 密码子和起始 Met-tRNA。来自三个框架的 poly-GA、poly-GP 和 poly-GR 蛋白的产生受相同 CUG 起始密码子突变的影响，支持移码机制。RAN 翻译也受到存在于错误剪接的 C9ORF72 转录本中的上游开放阅读框 (uORF) 的调节。起始核糖体复合物抑制剂和针对 5'-侧翼 GC 序列的 RNA 反义寡核苷酸选择性地阻断核糖体扫描并阻止翻译。最后，我们发现扩增的转录本与核糖体亚基具有出乎意料的亲和力，而与翻译无关。
