Reduced stress granule formation and cell death in fibroblasts with the A382T mutation of TARDBP gene: evidence for loss of TDP-43 nuclear function.
TAR deoxyribonucleic acid-binding protein 43 (TDP-43) is a key protein in the pathogenesis of amyoptrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Recent studies suggest that mutations in the TDP-43 coding gene, TARDBP, as well as variations in TDP-43 protein expression levels may disrupt the dynamics of stress granules (SGs). However, it remains unclear whether the pathogenetic effect of the TDP-43 protein is exerted at the cytoplasmic level, through direct participation to SG composition, or at nuclear level, through control of proteins essential to SG assembly. To clarify this point, we investigated the dynamics of SG formation in primary skin fibroblast cultures from the patients with ALS together with the A382T mutation and the patients with ALS and healthy controls with wild-type TDP-43. Under stress conditions induced by sodium arsenite, we found that in human fibroblasts TDP-43 did not translocate to the SGs but instead contributed to the SG formation through a regulatory effect on the G3BP1 core protein. We found that the A382T mutation caused a significant reduction in the number of SGs per cell (P < 0.01) as well as the percentage of cells that form SGs (P < 0.00001). Following stress stimuli, a significant decrease of viability was observed for cells with the TDP-43 A382T mutation (P < 0.0005).

We can therefore conclude that the A382T mutation caused a reduction in the ability of cells to respond to stress through loss of TDP-43 function in SG nucleation. The pathogenetic action revealed in our study model does not seem to be mediated by changes in the localization of the TDP-43 protein.
Medical Genetics, Department of Medical Sciences, University of Cagliari, R. Binaghi Hospital, Cagliari, Italy.
Sandro
S
Orrù
Paola Coni, Department of Biomedical Sciences, University of Cagliari, Monserrato, Italy.
Paola
P
Coni
Medical Genetics, Department of Medical Sciences, University of Cagliari, R. Binaghi Hospital, Cagliari, Italy.
Andrea
A
Floris
Regional Transplant Center, R. Binaghi Hospital, ASL Cagliari, Cagliari, Italy.
Roberto
R
Littera
Medical Genetics, Department of Medical Sciences, University of Cagliari, R. Binaghi Hospital, Cagliari, Italy.
Carlo
C
Carcassi
Paola Coni, Department of Biomedical Sciences, University of Cagliari, Monserrato, Italy.
Valeria
V
Sogos
Paola Coni, Department of Biomedical Sciences, University of Cagliari, Monserrato, Italy.
Carla
C
Brancia
D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics / Q000378:metabolism; D018053:Arsenites / Q000633:toxicity; D016923:Cell Death; D002478:Cells, Cultured; D003594:Cytoplasmic Granules / Q000187:drug effects / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D057180:Frontotemporal Dementia / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020125:Mutation, Missense; D017670:Sodium Compounds / Q000633:toxicity; D013312:Stress, Physiological / Q000187:drug effects
D018053:Arsenites; D004268:DNA-Binding Proteins; D017670:Sodium Compounds; C000598624:TARDBP protein, human; C017947:sodium arsenite