While cytoplasmic aggregation of TDP-43 is a pathological hallmark of amyotrophic lateral sclerosis and frontotemporal dementia, how aggregates form and what drives its nuclear clearance have not been determined. Here we show that TDP-43 at its endogenous level undergoes liquid-liquid phase separation (LLPS) within nuclei in multiple cell types. Increased concentration of TDP-43 in the cytoplasm or transient exposure to sonicated amyloid-like fibrils is shown to provoke long-lived liquid droplets of cytosolic TDP-43 whose assembly and maintenance are independent of conventional stress granules. Cytosolic liquid droplets of TDP-43 accumulate phosphorylated TDP-43 and rapidly convert into gels/solids in response to transient, arsenite-mediated stress. Cytoplasmic TDP-43 droplets slowly recruit importin-α and Nup62 and induce mislocalization of RanGap1, Ran, and Nup107, thereby provoking inhibition of nucleocytoplasmic transport, clearance of nuclear TDP-43, and cell death. These findings identify a neuronal cell death mechanism that can be initiated by transient-stress-induced cytosolic de-mixing of TDP-43.

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are associated with loss of nuclear transactive response DNA-binding protein 43 (TDP-43). Here we identify that TDP-43 regulates expression of the neuronal growth-associated factor stathmin-2. Lowered TDP-43 levels, which reduce its binding to sites within the first intron of stathmin-2 pre-messenger RNA, uncover a cryptic polyadenylation site whose utilization produces a truncated, non-functional mRNA. Reduced stathmin-2 expression is found in neurons trans-differentiated from patient fibroblasts expressing an ALS-causing TDP-43 mutation, in motor cortex and spinal motor neurons from patients with sporadic ALS and familial ALS with GGGGCC repeat expansion in the C9orf72 gene, and in induced pluripotent stem cell (iPSC)-derived motor neurons depleted of TDP-43. Remarkably, while reduction in TDP-43 is shown to inhibit axonal regeneration of iPSC-derived motor neurons, rescue of stathmin-2 expression restores axonal regenerative capacity. Thus, premature polyadenylation-mediated reduction in stathmin-2 is a hallmark of ALS-FTD that functionally links reduced nuclear TDP-43 function to enhanced neuronal vulnerability.