Essential role of proteasomes in maintaining self-renewal in neural progenitor cells

Protein turnover and homeostasis are regulated by the proteasomal system, which is critical for cell function and viability. Pluripotency of stem cells also relies on normal proteasomal activity that mitigates senescent phenotypes induced by intensive cell replications, as previously demonstrated in human bone marrow stromal cells. In this study, we investigated the role of proteasomes in self-renewal of neural progenitor cells (NPCs). Through both in vivo and in vitro analyses, we found that the expression of proteasomes was progressively decreased during aging. Likewise, proliferation and self-renewal of NPCs were also impaired in aged mice, suggesting that the down-regulation of proteasomes might be responsible for this senescent phenotype. Lowering proteasomal activity by loss-of-function manipulations mimicked the senescence of NPCs both in vitro and in vivo; conversely, enhancing proteasomal activity restored and improved self-renewal in aged NPCs. These results collectively indicate that proteasomes work as a key regulator in promoting self-renewal of NPCs. This potentially provides a promising therapeutic target for age-dependent neurodegenerative diseases

Inhibition of protein ubiquitination by paraquat and 1-methyl-4-phenylpyridinium impairs ubiquitin-dependent protein degradation pathways

Intracytoplasmic inclusions of protein aggregates in dopaminergic cells (Lewy bodies) are the pathological hallmark of Parkinson’s disease (PD). Ubiquitin (Ub), alpha [α]-synuclein, p62/sequestosome 1 and oxidized proteins are major components of Lewy bodies. However, the mechanisms involved in the impairment of misfolded/oxidized protein degradation pathways in PD are still unclear. PD is linked to mitochondrial dysfunction and environmental pesticide exposure. In this work, we evaluated the effect of the pesticide paraquat (PQ) and the mitochondrial toxin 1-methyl-4-phenylpyridinium (MPP+) on Ub-dependent protein degradation pathways. No increase in the accumulation of Ub-bound proteins or aggregates was observed in dopaminergic cells (SK-N-SH) treated with PQ or MPP+, or in mice chronically exposed to PQ. PQ decreased Ub protein content, but not its mRNA transcription. Protein synthesis inhibition with cycloheximide depleted Ub levels and potentiated PQ–induced cell death. Inhibition of proteasomal activity by PQ was found to be a late event in cell death progression, and had no effect on either the toxicity of MPP+ or PQ, or the accumulation of oxidized sulfenylated, sulfonylated (DJ-1/PARK7 and peroxiredoxins) and carbonylated proteins induced by PQ. PQ- and MPP+-induced Ub protein depletion prompted the dimerization/inactivation of the Ub-binding protein p62 that regulates the clearance of ubiquitinated proteins by autophagic. We confirmed that PQ and MPP+ impaired autophagy flux, and that the blockage of autophagy by the overexpression of a dominant-negative form of the autophagy protein 5 (dnAtg5) stimulated their toxicity, but there was no additional effect upon inhibition of the proteasome. PQ induced an increase in the accumulation of α-synuclein in dopaminergic cells and membrane associated foci in yeast cells. Our results demonstrate that inhibition of protein ubiquitination by PQ and MPP+ is involved in the dysfunction of Ub-dependent protein degradation pathways