The role of GSH signaling in mitophagy induced by reactive oxygen species: potential target for neuroprotection in stroke?

• Autor(a) principal: Tavares, Diana Sofia Ferreira
• Data de Publicação: 2022
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10362/154800
• topic: Mitochondria; Oxidative Stress; Mitophagy; Glutathione; Protein S-glutathi- onylation; Stroke; Domínio/Área Científica::Engenharia e Tecnologia::Outras Engenharias e Tecnologias

Mitochondria are involved in numerous cellular processes, including energy production and cell death regulation. Simultaneously, these are the main cellular producers of reactive oxygen species (ROS), particularly when dysfunctional. ROS accumulation leads to oxidative stress which is associated with pathologies, such as stroke and neurodegenerative diseases. So, elimination of ROS-producing damaged mitochondria is key for maintaining cell homeostasis. Mitophagy, consists of the selective degradation of excessive or damaged mitochondria. The accumulation of ROS and mitochondrial depolarization stimulates PTEN-induced kinase 1 (PINK1)/PARKIN-mediated mitophagy. Concomitantly, ROS are counteracted by glutathione, a key antioxidant. ROS oxidize glutathione and stimulate protein S-glutathionylation, a post- translation modification capable of modulate protein function. Here, the main goal was to ex- plore the molecular mechanisms underlying mitophagy, in particular the role of ROS and/or glutathione signaling in mitophagy progression, with a special focus on protein S-glutathi- onylation. We hypothesized that mitophagy can be regulated by glutathione signaling, namely that the accumulation of mitochondrial ROS stimulates the production and oxidation of gluta- thione, which enhances/stimulates mitophagy by protein glutathionylation of mitophagy-re- lated proteins. Using human cell lines, SH-SY5Y and SH-SY5Y mitoQC, mitochondrial dysfunc- tion was stimulated with the classical uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP). ROS generation and PINK1/PARKIN-dependent mitophagy were observed after CCCP treatment. Likewise, the catalytic subunit of the rate limiting enzyme for glutathione synthesis, Glutamate-Cysteine Ligase, is upregulated in CCCP-treated cells. Notably, we show for the first time that CCCP treatment induces protein S-glutathionylation of PINK1. Furthermore, we also found fluorescence measure on a microplate reader to be a possible new technique to assess mitophagy. Understating mitophagy mechanisms and how these are regulated is key to stimulate cytoprotection by preventing cell death. Thus, this can eventually contribute to new strategies against diseases characterized by mitochondrial damage, like stroke.