Yahara Masao, Kitamura Akira, Kinjo Masataka
Laboratory of Molecular Cell Dynamics, Faculty of Advanced Life Science, Hokkaido University, Sapporo, Japan.
PLoS One. 2017 Nov 10;12(11):e0187813. doi: 10.1371/journal.pone.0187813. eCollection 2017.
Depletion of amyotrophic lateral sclerosis (ALS)-associated transactivation response (TAR) RNA/DNA-binding protein 43 kDa (TDP-43) alters splicing efficiency of multiple transcripts and results in neuronal cell death. TDP-43 depletion can also disturb expression levels of small nuclear RNAs (snRNAs) as spliceosomal components. Despite this knowledge, the relationship between cell death and alteration of snRNA expression during TDP-43 depletion remains unclear. Here, we knocked down TDP-43 in murine neuroblastoma Neuro2A cells and found a time lag between efficient TDP-43 depletion and appearance of cell death, suggesting that several mechanisms mediate between these two events. The amount of U6 snRNA was significantly decreased during TDP-43 depletion prior to increase of cell death, whereas that of U1, U2, and U4 snRNAs was not. Downregulation of U6 snRNA led to cell death, whereas transient exogenous expression of U6 snRNA counteracted the effect of TDP-43 knockdown on cell death, and slightly decreased the mis-splicing rate of Dnajc5 and Sortilin 1 transcripts, which are assisted by TDP-43. These results suggest that regulation of the U6 snRNA expression level by TDP-43 is a key factor in the increase in cell death upon TDP-43 loss-of-function.
肌萎缩侧索硬化症（ALS）相关的反式激活应答（TAR）RNA/DNA结合蛋白43 kDa（TDP-43）的缺失会改变多个转录本的剪接效率，并导致神经元细胞死亡。TDP-43的缺失还会扰乱作为剪接体成分的小核RNA（snRNA）的表达水平。尽管有这些认识，但在TDP-43缺失期间细胞死亡与snRNA表达改变之间的关系仍不清楚。在这里，我们在小鼠神经母细胞瘤Neuro2A细胞中敲低TDP-43，发现在有效的TDP-43缺失与细胞死亡出现之间存在时间滞后，这表明在这两个事件之间有几种机制起介导作用。在细胞死亡增加之前的TDP-43缺失期间，U6 snRNA的量显著减少，而U1、U2和U4 snRNA的量则没有减少。U6 snRNA的下调导致细胞死亡，而U6 snRNA的瞬时外源表达抵消了TDP-43敲低对细胞死亡的影响，并略微降低了由TDP-43辅助的Dnajc5和Sortilin 1转录本的错配剪接率。这些结果表明，TDP-43对U6 snRNA表达水平的调节是TDP-43功能丧失时细胞死亡增加的关键因素。
