. Active nuclear import and passive nuclear export are the primary determinants of TDP-43 localization. Sci Rep. 2018 May 4;8(1):7083. PubMed.


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  1. Overall, I was very impressed by this study of Ederle et al. This is one among a series of manuscripts, including the Pinarbarsi study and our own (all published in Scientific Reports), that show convincingly that nuclear export of TDP-43 and FUS are not as simple as they seem.

    The annotated nuclear export signal (NES) within TDP-43’s RRM2 was predicted to be a CRM1/XPO1 substrate in 2008, but subsequent data never really supported this hypothesis. There have been several proteomic studies attempting to identify CRM1 substrates, but TDP-43 was never among those listed. Our data indicated that leptomycin B and Karyopharm's selective inhibitor of nuclear export (SINE) compounds—potent inhibitors of CRM1—also failed to affect TDP-43 localization. These conclusions are supported by the Ederle and Pinarbarsi studies, both of which demonstrated that the annotated TDP-43 “NES” failed to function as such. 

    What about the implications for therapeutic design? Although the Karyopharm compounds demonstrated modest neuroprotective effects in ALS models, they did so at low concentrations that had no effect on CRM1-dependent nuclear export. When we used these compounds at higher doses, we noted pronounced inhibition of nuclear export, but also considerable toxicity in primary neuron preparations. Together with the data supporting CRM1-independent transport of TDP-43, these results indicate that the modest neuroprotection afforded by SINE compounds have little to do with TDP-43 localization.

    All three studies showed that CRM1 is not necessary for the nuclear egress of TDP-43. Even so, while no single exporter was necessary for TDP-43 nuclear export, we found several, including CRM1, XPO7, and NXF1, that were sufficient to drive nuclear export of TDP-43. This observation suggests that nucleocytoplasmic transport mechanisms for TDP-43 are partially redundant, as can be seen for some essential RNA binding proteins (i.e., hnRNPs). 

    Whether TDP-43 nuclear egress is passive, or actively mediated by several exporters, the therapeutic implications are similar—both possibilities significantly complicate the rational design of therapies aimed at preventing TDP-43 nuclear export. 

    View all comments by Sami Barmada
  2. Ever since 2006, when Virginia Lee and colleagues found cytoplasmic mislocalization and aggregation of phosphorylated nuclear proteins TDP-43 (and later FUS and hnRNPs) as a pathological hallmark of ALS and FTD (Neumann et al., 2006), researchers have striven to understand its molecular mechanism. In 2015, three contemporary studies (Zhang et al., 2015; Freibaum et al., 2015; Jovičić et al., 2015) brought nucleocytoplasmic transport to the focus of our attention, suggesting a tempting model that an imbalance of nuclear import and export of TDP-43, FUS, etc., causes the cytoplasmic mislocalization and subsequent phosphorylation and aggregation of these proteins. In accordance with this model, KPTs, a series of chemical compounds inhibiting nuclear export receptor Exportin-1, suppress neurodegeneration in animal and cell models of ALS (Zhang et al., 2015; Chou et al., 2018), likely through correcting this imbalance and thus the TDP-43 pathology.

    However, more recent studies suggested several caveats to this oversimplified model. Firstly, Mark Hipp, Ulrich Hartl, Wilfried Rossoll, and we reported that cytoplasmic protein aggregates, including TDP-43, disrupt nucleocytoplasmic transport through recruiting essential transport factors to these aggregates and/or stress granules induced by these aggregates (Woerner et al., 2016; Chou et al., 2018; Zhang et al., 2018). Furthermore, four recent studies by Jim Shorter, Dorothee Dormann, Yuh Min Chook, Peter St George-Hyslop, and colleagues showed that Transportin-1, the import receptor for FUS, functions as a chaperone preventing cytoplasmic FUS phase separation and aggregation (Guo et al., 2018; Hofweber et al., 2018; Yoshizawa et al., 2018; Qamar et al., 2018). Taken together, these studies suggest that nucleocytoplasmic transport and TDP-43/FUS cytoplasmic aggregation mutually regulate each other, with stress granule assembly/liquid-liquid phase separation as a key mediator. Importantly, this current paper by Dormann and colleagues shows convincing evidence that the nuclear export of TDP-43 does not need Exportin-1, arguing against our earlier explanation of how KPTs suppress neurodegeneration. Consistent with these findings, a prior study led by Sami Barmada reported that despite its protective effect against TDP-43, KPT-350 does not suppress TDP-43 cytoplasmic mislocalization! 

    These interesting findings have not only led us to better understand the ALS/FTD pathophysiology, but also raise many exciting questions. Firstly, what is the function of the nuclear export signals of TDP-43 and FUS if the proteins do not require exportins for their export? Secondly, how about phospho-TDP-43? Do KPTs affect its localization and aggregation? Importantly, and particularly interesting to people exploring the therapeutic potential of KPTs, what mediates the compounds’ protective effects in ALS/FTD models? Although other downstream targets of KPTs can be the answer, recent findings have suggested a possible pathway by which KPTs mitigate TDP-43 and FUS toxicity. Cellular stress disrupts RNA metabolism as well as nucleocytoplasmic transport (Zhang et al., 2018), causing TDP-43 and FUS to localize to cytoplasmic stress granules. Interestingly, as suggested by the current paper, RNA defects may also enhance the export of TDP-43 and FUS. Therefore, some cellular stress response pathways may be the answer. Indeed, several essential stress granule factors (e.g., TIA1 and G3BPs) can undergo nucleocytoplasmic shuttling. As inhibiting stress granule assembly suppresses TDP-43 toxicity and neurodegeneration in multiple ALS/FTD models (Elden et al., 2010; Kim et al., 2014; Becker et al., 2017; Zhang et al., 2018), KPTs may execute their protective roles via inhibiting stress granule assembly, which in turn prevents TDP-43/FUS phase separation and aggregation.


    . Ubiquitinated TDP-43 in frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Science. 2006 Oct 6;314(5796):130-3. PubMed.

    . The C9orf72 repeat expansion disrupts nucleocytoplasmic transport. Nature. 2015 Sep 3;525(7567):56-61. Epub 2015 Aug 26 PubMed.

    . GGGGCC repeat expansion in C9orf72 compromises nucleocytoplasmic transport. Nature. 2015 Sep 3;525(7567):129-33. Epub 2015 Aug 26 PubMed.

    . TDP-43 pathology disrupts nuclear pore complexes and nucleocytoplasmic transport in ALS/FTD. Nat Neurosci. 2018 Feb;21(2):228-239. Epub 2018 Jan 8 PubMed.

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    . Nuclear-Import Receptors Reverse Aberrant Phase Transitions of RNA-Binding Proteins with Prion-like Domains. Cell. 2018 Apr 19;173(3):677-692.e20. PubMed.

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    . Nuclear Import Receptor Inhibits Phase Separation of FUS through Binding to Multiple Sites. Cell. 2018 Apr 19;173(3):693-705.e22. PubMed.

    . FUS Phase Separation Is Modulated by a Molecular Chaperone and Methylation of Arginine Cation-π Interactions. Cell. 2018 Apr 19;173(3):720-734.e15. PubMed.

    . Stress Granule Assembly Disrupts Nucleocytoplasmic Transport. Cell. 2018 May 3;173(4):958-971.e17. Epub 2018 Apr 5 PubMed.

    . Ataxin-2 intermediate-length polyglutamine expansions are associated with increased risk for ALS. Nature. 2010 Aug 26;466(7310):1069-75. PubMed.

    . Therapeutic modulation of eIF2α phosphorylation rescues TDP-43 toxicity in amyotrophic lateral sclerosis disease models. Nat Genet. 2014 Feb;46(2):152-60. Epub 2013 Dec 15 PubMed.

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    View all comments by Ke Zhang
  3. This interesting and thought-provoking paper from the Dormann lab reports that TDP-43 and FUS appear to leave the nucleus from passive diffusion rather than active nucleocytoplasmic transport via the Exportin-1/CRM1 export receptor, at least in HeLa cells. This is a surprising finding, since it was widely believed that these RNA-binding proteins shuttle in and out of the nucleus via transport receptors binding to their nuclear localization and export sequences. Recent publications from several labs have demonstrated nucleocytoplasmic transport defects in C9-ALS models, and partial rescue of disease phenotypes via CRM1 inhibitors developed by Karyopharm (e.g., KPT-276 and KPT-350). We have shown similar defects and rescue in TDP-43 proteinopathy models of ALS.

    Does this new finding that TDP-43 and FUS are not actively exported from the nucleus call into question the use of KPT compounds as potential therapeutic interventions for ALS/FTD? I don’t think so.  In my opinion, it appears likely that these drugs address a general defect in nuclear protein import by inhibiting nuclear export, thus restoring a balance between these processes. I don’t think that correcting FUS and TDP-43 localization has been considered the most likely mechanism for the observed therapeutic effect in ALS disease models.

    View all comments by Wilfried Rossoll
  4. I am surprised to see data that TDP-43 export is occurring through passive diffusion rather than active export. I am not sure this will have huge implications on the effectiveness of KPT-350, as it seems to promote neuronal health independent of direct interactions with TDP-43 or FUS. It does appear that C9ORF72 dipeptides contribute to the mislocalization of TDP-43 but it is unclear whether this is due directly to impairment of import/export through the nuclear pore or indirectly through the accumulation of TDP-43 in stress granules, via the sequestration of nuclear import factors. The authors of this publication did not observe TDP-43 in stress granules, however, this could be due to an issue with the sensitivity of their antibodies.

    View all comments by Brian Freibaum

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  1. No, TDP-43 and FUS Are Not Actively Exported From the Nucleus