Overexpression of α-synuclein inhibits mitochondrial Ca2+ trafficking between the endoplasmic reticulum and mitochondria through MAMs by altering the GRP75-IP3R interaction.

Mitochondria-associated ER membranes (MAMs) are formed by close and specific components in the contact sites between the endoplasmic reticulum (ER) and mitochondria, which participate in several cell functions, including lipid metabolism, autophagy, and Ca2+ signaling. Particularly, the presence of α-synuclein (α-syn) in MAMs was previously demonstrated, indicating a physical interaction among some proteins in this region and a potential involvement in cell dysfunctions. MAMs alterations are associated with neurodegenerative diseases such as Parkinson's disease (PD) and contribute to the pathogenesis features. Here, we investigated the effects of α-syn on MAMs and Ca2+ transfer from the ER to mitochondria in WT- and A30P α-syn-overexpressing SH-SY5Y or HEK293 cells. We observed that α-syn potentiates the mitochondrial membrane potential (Δψm ) loss induced by rotenone, increases mitophagy and mitochondrial Ca2+ overload. Additionally, in α-syn-overexpressing cells, we found a reduction in ER-mitochondria contact sites through the impairment of the GRP75-IP3R interaction, however, with no alteration in VDAC1-GRP75 interaction. Consequently, after Ca2+ release from the ER, α-syn-overexpressing cells demonstrated a reduction in Ca2+ buffering by mitochondria, suggesting a deregulation in MAM activity. Taken together, our data highlight the importance of the α-syn/MAMs/Ca2+ axis that potentially affects cell functions in PD.

Overexpression of α-synuclein in an astrocyte cell line promotes autophagy inhibition and apoptosis.

α-Synuclein is the major component of neuronal cytoplasmic aggregates called Lewy bodies, the main pathological hallmark of Parkinson disease. Although neurons are the predominant cells expressing α-synuclein in the brain, recent studies have demonstrated that primary astrocytes in culture also express α-synuclein and regulate α-synuclein trafficking. Astrocytes have a neuroprotective role in several detrimental brain conditions; we therefore analyzed the effects of the overexpression of wild-type α-synuclein and its A30P and A53T mutants on autophagy and apoptosis. We observed that in immortalized astrocyte cell lines, overexpression of α-synuclein proteins promotes the decrease of LC3-II and the increase of p62 protein levels, suggesting the inhibition of autophagy. When these cells were treated with rotenone, there was a loss of mitochondrial membrane potential, especially in cells expressing mutant α-synuclein. The level of this decrease was related to the toxicity of the mutants because they show a more intense and sustained effect. The decrease in autophagy and the mitochondrial changes in conjunction with parkin expression levels may sensitize astrocytes to apoptosis.

Exploring Caenorhabditis elegans as Parkinson's Disease Model: Neurotoxins and Genetic Implications.

Parkinson's disease (PD) is the second most common neurodegenerative disease in the world, the first being Alzheimer's disease. Patients with PD have a loss of dopaminergic neurons in the substantia nigra of the basal ganglia, which controls voluntary movements, causing a motor impairment as a result of dopaminergic signaling impairment. Studies have shown that mutations in several genes, such as SNCA, PARK2, PINK1, DJ-1, ATP13A2, and LRRK2, and the exposure to neurotoxic agents can potentially increase the chances of PD development. The nematode Caenorhabditis elegans (C. elegans) plays an important role in studying the risk factors, such as genetic factors, aging, exposure to chemicals, disease progression, and drug treatments for PD. C. elegans has a conserved neurotransmission system during evolution; it produces dopamine, through the eight dopaminergic neurons; it can be used to study the effect of neurotoxins and also has strains that express human α-synuclein. Furthermore, the human PD-related genes, LRK-1, PINK-1, PDR-1, DJR-1.1, and CATP-6, are present and functional in this model. Therefore, this review focuses on highlighting and discussing the use of C. elegans an in vivo model in PD-related studies. Here, we identified that nematodes exposed to the neurotoxins, such as 6-OHDA, MPTP, paraquat, and rotenone, had a progressive loss of dopaminergic neurons, dopamine deficits, and decreased survival rate. Several studies have reported that expression of human LRRK2 (G2019S) caused neurodegeneration and pink-1, pdr-1, and djr-1.1 deletion caused several effects PD-related in C. elegans, including mitochondrial dysfunctions. Of note, the deletion of catp-6 in nematodes caused behavioral dysfunction, mitochondrial damage, and reduced survival. In addition, nematodes expressing α-synuclein had neurodegeneration and dopamine-dependent deficits. Therefore, C. elegans can be considered an accurate animal model of PD that can be used to elucidate to assess the underlying mechanisms implicated in PD to find novel therapeutic targets.

Galectin-3 sensitized melanoma cell lines to vemurafenib (PLX4032) induced cell death through prevention of autophagy.

Melanoma is a current worldwide problem, as its incidence is increasing. In the last years, several studies have shown that melanoma cells display high levels of autophagy, a self-degradative process that can promote survival leading to drug resistance. Consequently, autophagy regulation represents a challenge for cancer therapy. Herein, we showed that galectin-3 (Gal-3), a ß-galactoside binding lectin which is often lost along melanoma progression, is a negative regulator of autophagy in melanoma cells. Our data demonstrated that Gal-3low/negative cells were more resistant to the inhibition of the activity of the cancer driver gene BRAFV600E by vemurafenib (PLX4032). Interestingly, in these cells, starvation caused further LC3-II accumulation in cells exposed to chloroquine, which inhibits the degradative step in autophagy. In addition, Gal-3 low/negative tumor cells accumulated more LC3-II than Gal-3 high tumor cells in vivo. Resistance of Gal-3low/negative cells was associated with increased production of superoxide and activation of the Endoplasmic Reticulum (ER) stress response, as evaluated by accumulation of GRP78. Pharmacological inhibition of autophagy with bafilomycin A reversed the relative resistance of Gal-3low/negative cells to vemurafenib treatment. Taken together, these results show that the autophagic flux is dependent on Gal-3 levels, which attenuate the prosurvival role of autophagy.