The aggregation mechanisms of TDP-43 inclusion in neuronal cell

TAR DNA结合蛋白43 kDa（TDP-43）是肌萎缩侧索硬化症（ALS）和额颞叶变性伴TDP-43阳性包涵体（FTLD-TDP）特征性神经元和神经胶质包涵体聚集体的主要成分。然而，TDP-43聚集体是否有毒，以及TDP-43的异常，如聚集、泛素化、过度磷酸化、片段化和核定位丢失，如何导致神经元变性，仍有待澄清。我们在这里报告，增殖的人神经母细胞瘤细胞系SH-SY 5 Y与TDP-43夹杂物被强烈抑制，相比，没有夹杂物的细胞。TDP-43的C端片段对细胞生长的抑制作用尤为明显。有趣的是，TDP-43全长片段的生长抑制机制与阻滞于G2/M期有关，而TDP-43的C端片段的生长抑制机制与此无关。在TDP-43表达细胞中，RNA聚合酶II和几种转录因子被发现与TDP-43聚集体共定位。此外，在FTLD-TDP脑中检测到磷酸化TDP-43包涵体中RNA聚合酶II的积累。这些结果表明，异常TDP-43内含物本身是细胞毒性的，并通过募集正常转录因子导致细胞功能障碍，从而导致生长停滞。这种转录失调可能导致TDP-43蛋白病中的神经元变性。

TAR DNA binding protein of 43 kDa(TDP-43) is the major component of neuronal and glial inclusions aggregates characteristic of amyotrophic lateral sclerosis(ALS) and frontotemporal lobar degeneration with TDP-43-positive inclusions(FTLD-TDP). However, it remains to be clarified whether or not TDP-43 aggregates are toxic, and how abnormalities of TDP-43, such as aggregation, ubiquitination, hyperphosphorylation, fragmentation and loss of nuclear localization, lead to neuronal degeneration. We report here that proliferation of human neuroblastoma cell line SH-SY5Y with TDP-43 inclusions is strongly suppressed, compared to that of cells without the inclusions. Growth inhibition was especially strong in cells expressing a C-terminal fragment of TDP-43.Interestingly, the mechanism of cell growth inhibition by full-length TDP-43 involved arrest at the G2/M phase, whereas that by C-terminal fragment of TDP-43 did not. In TDP-43-expressing cells, RNA polymerase II and several transcription factors were found to be co-localized with TDP-43 aggregates. Furthermore, accumulation of RNA polymerase II in phosphorylated TDP-43 inclusions was detected in FTLD-TDP brains. These results suggest that abnormal TDP-43 inclusions itself is cytotoxic and lead to cellular dysfunction by recruiting normal transcription factors, resulting in growth arrest. Such transcriptional dysregulation may contribute to neuronal degeneration in TDP-43 proteinopathy.