Figure 2.

TARDBP is required for a homeostatic autophagy response. (a) TARDBP knockdown (KD) in HeLa cells leads to decreased ATG4B mRNA levels (left panel) linked to increased amounts of ATG4B mRNA with cryptic exons (right panel); this is associated with a failure to enhance ATG4B mRNA expression as a response to autophagy stimulation by nutrient deprivation. (b) TARDBP protein loss is associated with ATG4B decrease as shown by western blot (left panel) and densitometry (right panel). (c) Functional analyses reveal that loss of TARDBP in HeLa cells induces increased levels of SQSTM1 as shown by western blot (left panel) and immunofluorescence (right panel), suggesting a functional loss of autophagy flux. (d) The increase in SQSTM1 induced by TARDBP knockdown is rescued by ATG4B overexpression, as shown by densitometry analyses, either in HeLa cells (left panel) or in Human Neural Tissue primary cells (right panel). Blots are representative of different experiments (n = 3–5). Bars indicate mean values (± standard error). For (a), (b) and (c) **** p < 0.0001, ***p < 0.001 after 2-way ANOVA. For (d) *p < 0.05 and **p < 0.01 in Student’s t test.