is an RNA-binding protein with pleiotropic functions, including pre-mRNA splicing, mRNA processing, and transport (1
). Notably, TDP-43 is the major component of ubiquitin-positive inclusions in diseased brains in a subtype of frontotemporal lobar degeneration (FTLD-U) as well as in many cases of amyotrophic lateral sclerosis (ALS) (1
). FTLD is the second most frequent form of presenile dementia after Alzheimer disease, whereas ALS constitutes the most common motor neuron disease (1
). In patients suffering from both familial and sporadic ALS, mutations in the TDP-43-encoding TARDBP
have been identified, supporting the notion that either the onset of toxicity due to TDP-43 aggregation or the loss of TDP-43 function triggers the neurological disorder (1
Diverse animal and cellular models, including mouse, rat, Drosophila
, Caenorhabditis elegans
, and mammalian cell culture, have been developed to dissect the molecular mechanisms governing TDP-43 proteinopathies (3
). Transgenic animal models expressing wild-type or ALS-associated variants of TDP-43 display pathological, ubiquitinated protein aggregates in motor neurons with subsequent neuronal cell loss and progressive neurodegenerative phenotypes (6
). Accordingly, expression of wild-type and disease-associated TDP-43 variants in mammalian cell cultures results in the formation of ubiquitin-positive cellular inclusions correlated with increased incidences of cell death (10
). However, the molecular pathways of TDP-43-triggered neuronal cell loss remain to be identified.
Although the yeast Saccharomyces cerevisiae
genome encodes no apparent ortholog of TDP-43 (13
), heterologous expression of human TDP-43 results in phenotypes noticeably resembling TDP-43 pathology in higher model organisms and in diseased humans (13
), including (i) the translocation of TDP-43 from the nucleus to the cytoplasm followed by the formation of cytoplasmic TDP-43 foci; (ii) the accumulation of TDP-43-specific inclusions, which are accelerated upon expression of disease-associated TDP-43 variants; (iii) the identification of the C terminus of TDP-43 to be essential for TDP-43 aggregation; (iv) the correlation of TDP-43 pathology with growth inhibition in yeast; and finally (v) the occurrence of plasma membrane permeabilization, which is highly suggestive of TDP-43-triggered cell death.
Yeast is a powerful, genetically tractable model organism for studying cell death (16
). Morphological markers of apoptosis and necrosis have been discovered in yeast (19
), and the molecular mechanisms of cellular demise resemble those from higher organisms. The yeast genome encodes conserved regulators of cell death, including proteases like the yeast caspase 1 (Yca1p), the serine proteases Nma111p and Kex1p, the calpain-like cysteine protease Cpl1p, and the vacuolar aspartyl protease Pep4p (22
), as well as mitochondrial proteins like apoptosis-inducing factor (Aif1p); Ndi1p, the yeast homolog of the AIF-homologous mitochondrion-associated inducer of death; and endonuclease G (Nuc1p) (23
). Consequently, yeast undergoes distinct cell death pathways, including cell death protease-dependent and -independent pathways (17
), or distinct modes of death involving the mitochondrion (16
). These pathways are mechanistically similar to those observed in neurodegenerative disorders, including critical contributions of oxidative stress, mitochondria, and aging (16
). The high degree of conservation of lethal signals and processes throughout evolution from yeast to humans has prompted the use of yeast for studies of neurotoxic cell death (33
In this study, we show that TDP-43 expression in yeast resulted in the formation of perinuclear and perimitochondrial aggregate-like foci and in the induction of oxidative stress and age-associated cytotoxicity culminating in apoptosis and necrosis. TDP-43-triggered cytotoxicity required mitochondrial functionality but depended neither on the release of the mitochondrial proteins Aif1p, Nuc1p, and cytochrome c nor on the activity of the cell death proteases Yca1p, Nma111p, Kex1p, Cpl1p, and Pep4p. Finally, we show that TDP-43-triggered cytotoxicity strictly correlated with respiratory capacity, mitochondrial DNA (mtDNA) stability, and respiratory chain activity, suggesting that oxidative stress, mitochondria, and respiration crucially influence neuronal death in TDP-43 pathologies.