Dysregulation of the ALS-associated gene TDP-43 leads to neuronal death and degeneration in mice
Lionel M. Igaz, … , John Q. Trojanowski, Virginia M.-Y. Lee
Lionel M. Igaz, … , John Q. Trojanowski, Virginia M.-Y. Lee
Published January 4, 2011
Citation Information: J Clin Invest. 2011;121(2):726-738. https://doi.org/10.1172/JCI44867.
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Research Article Neuroscience

Dysregulation of the ALS-associated gene TDP-43 leads to neuronal death and degeneration in mice

Abstract

Amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) are characterized by cytoplasmic protein aggregates in the brain and spinal cord that include TAR-DNA binding protein 43 (TDP-43). TDP-43 is normally localized in the nucleus with roles in the regulation of gene expression, and pathological cytoplasmic aggregates are associated with depletion of nuclear protein. Here, we generated transgenic mice expressing human TDP-43 with a defective nuclear localization signal in the forebrain (hTDP-43-ΔNLS), and compared them with mice expressing WT hTDP-43 (hTDP-43-WT) to determine the effects of mislocalized cytoplasmic TDP-43 on neuronal viability. Expression of either hTDP-43-ΔNLS or hTDP-43-WT led to neuron loss in selectively vulnerable forebrain regions, corticospinal tract degeneration, and motor spasticity recapitulating key aspects of FTLD and primary lateral sclerosis. Only rare cytoplasmic phosphorylated and ubiquitinated TDP-43 inclusions were seen in hTDP-43-ΔNLS mice, suggesting that cytoplasmic inclusions were not required to induce neuronal death. Instead, neurodegeneration in hTDP-43 and hTDP-43-ΔNLS–expressing neurons was accompanied by a dramatic downregulation of the endogenous mouse TDP-43. Moreover, mice expressing hTDP-43-ΔNLS exhibited profound changes in gene expression in cortical neurons. Our data suggest that perturbation of endogenous nuclear TDP-43 results in loss of normal TDP-43 function(s) and gene regulatory pathways, culminating in degeneration of selectively vulnerable affected neurons.

Authors

Lionel M. Igaz, Linda K. Kwong, Edward B. Lee, Alice Chen-Plotkin, Eric Swanson, Travis Unger, Joe Malunda, Yan Xu, Matthew J. Winton, John Q. Trojanowski, Virginia M.-Y. Lee

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Figure 1

Generation and biochemical characterization of inducible TDP-43 Tg mice.

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Generation and biochemical characterization of inducible TDP-43 Tg mice....
(A) Generation of conditional Tg mice. Camk2a-tTA mice were crossed with tetO-TDP-WT or tetO-TDP-ΔNLS mice. In bigenic mice, Dox inhibits tTA binding to the tetracycline-responsive promoter element (tetO), repressing hTDP-43 expression. (B) Dox was removed at weaning (P28) followed by analysis at the indicated time points. (C) Expression of hTDP-43 in Tg mice. Immunoblot of hTDP-43 or total TDP-43 (h+mTDP-43) in cortical RIPA extracts (1 month off Dox) of nTg, tTA, tTA/TDP-WT (WT4, WT8, WT5 and WT12), and tTA/TDP-ΔNLS mice (ΔNLS4 and ΔNLS19) showed variable expression of hTDP-43. GAPDH is a loading control. Fold expression relative to nTg and tTA control mice is shown, as described in Methods. (D) Regional expression of hTDP-43. RIPA extracts from different brain regions of tTA/TDP-WT or tTA/TDP-ΔNLS mice (1 month off Dox) were immunoblotted for hTDP-43, h+m TDP-43, and GAPDH. Ctx, cerebral cortex; Hp, hippocampus; OB, olfactory bulb; Cb, cerebellum; Rest, brain stem plus subcortical gray regions; SC, spinal cord. (E) Low levels of insoluble hTDP-43. Sequential cortical extracts from tTA/TDP-WT and tTA/TDP-ΔNLS mice of increasing time off Dox were immunoblotted for hTDP-43 showing low levels of TDP-43 in urea fractions. Urea lysates were loaded at 4× relative to RIPA lysates so that the low level of insoluble TDP-43 could be seen. R, RIPA; U, urea. n = at least 3 animals per group analyzed in C–E.