Abstract
Background
TDP-43 is a nuclear protein involved in pivotal processes, extensively studied for its implication in neurodegenerative disorders. TDP-43 cytosolic inclusions are a common neuropathologic hallmark in amyotrophic lateral sclerosis (ALS) and related diseases, and it is now established that TDP-43 misfolding and aggregation play a key role in their etiopathology. TDP-43 neurotoxic mechanisms are not yet clarified, but the identification of proteins able to modulate TDP-43-mediated damage may provide crucial information to unveil the molecular basis of TDP-43 proteinopathies.
Methods
Here we generated and characterized novel models of TDP-43 toxicity in the yeast S. cerevisiae, which were used to investigate the effect of the nucleolar protein nucleolin (NCL) on TDP-43-damaged yeast cells, by employing multiple approaches (genetics, biochemistry, microscopy). We further characterized the NCL-TDP-43 relationship in human HEK293T cells, by the combination of biochemical and microscopy-based assays.
Results
We show for the first time that NCL acts as a potent suppressor of TDP-43 toxicity in yeast models, since NCL overexpression is able to rescue TDP-43-dependent damage on cell viability and morphology, by reducing the levels of TDP-43 aggregates, thus proteostatic stress. Interestingly, data in yeast cells point to the implication of the extra-nuclear fraction of NCL in the suppressive effect. We further provide evidence that NCL co-expression alleviates the TDP-43-induced toxicity also in HEK293T cells, as indicated by the restoration of cell viability, and the diminished apoptosis activation. Importantly, biochemical and microscopy data indicate that NCL protein in human cells reduces the amount of TDP-43 inclusions. Collectively, results in HEK293T cells further support the beneficial effects of NCL on TDP-43-dependent toxicity in a more consistent pathophysiological context.
Conclusions
Altogether, data in yeast and human cell models demonstrate that NCL potently supresses the cytotoxicity caused by the TDP-43 protein, and further suggest that NCL could act by promoting the TDP-43 nuclear retention, and thus reducing the formation of cytosolic TDP-43 toxic aggregates. Pinpointing NCL as a novel player in mediating TDP-43 toxicity, experimental evidence could support NCL as promising therapeutic target in ALS and ALS-related disorders.
Nucleolin Rescues TDP-43 Toxicity in Yeast and Human Cell Models