The polyphenol quercetin protects from glucotoxicity depending on the aggresome in Caenorhabditis elegans.

PURPOSE: Impaired proteostasis, i.e., protein homeostasis, is considered as a consequence of high-glucose exposure and is associated with reduced survival. The previous studies demonstrated that the polyphenol quercetin can protect from glucotoxicity. The aim of the present study was to unravel the contribution of the aggresome, sequestering potentially cytotoxic aggregates and also acting as a staging center for eventual autophagic clearance from the cell. METHODS: Knockdown of the aggresome-relevant genes dnc-1 and ubql-1 was achieved in stress-sensitive mev-1 mutants of the nematode Caenorhabditis elegans by RNA interference (RNAi). Survival assay was conducted under heat stress at 37 °C, protein aggregation using ProteoStat® and chymotrypsin-like proteasomal activity according to the cleavage of a fluorogenic peptide substrate. RESULTS: Survival was reduced by knockdown of ubql-1 and even more by knockdown of dnc-1 which both were not further reduced by addition of glucose. The rescue of survival due to quercetin in glucose-exposed nematodes was completely prevented under RNAi versus ubql-1 or dnc-1. Both knockdowns caused an increase of aggregated protein and prevented the reduction of aggregated protein caused by quercetin in glucose-exposed animals. Finally, the knockdown of ubql-1 and dnc-1 blocked the increase of proteasomal activity achieved by quercetin in glucose-treated nematodes. CONCLUSIONS: The study provides evidence that quercetin protects C. elegans from glucotoxicity through the activation of the aggresome, thereby, quercetin prevents the aggregation and functional loss of proteins, which is typically caused by enhanced glucose concentrations.

Betaine-rich sugar beet molasses protects from homocysteine-induced reduction of survival in Caenorhabditis elegans.

PURPOSE: Homocysteine (Hcy) in humans represents a blood-borne biomarker which predicts the risk of age-related diseases and mortality. Using the nematode Caenorhabditis elegans, we tested whether feeding betaine-rich sugar beet molasses affects the survival under heat stress in the presence of Hcy, in spite of a gene loss in betaine-homocysteine methyltransferase. METHODS: Knockdown of the genes relevant for remethylation or transsulfuration of Hcy was achieved by RNA interference (RNAi). Survival assay was conducted under heat stress at 37 °C and Hcy levels were determined by enzyme-linked immunosorbent assay. RESULTS: Addition of 500 mg/l betaine-rich sugar beet molasses (SBM) prevented the survival reduction that was caused by exposure to Hcy at 37 °C. Although SBM was no longer capable of reducing Hcy levels under RNAi versus homologues for 5, 10-methylenetetrahydrofolate reductase or cystathionine-ß-synthase, it still enabled the survival extension by SBM under exposure to Hcy. In contrast, RNAi for the small heat shock protein hsp-16.2 or the foxo transcription factor daf-16 both prevented the extension of survival by betaine-rich molasses in the presence of Hcy. CONCLUSIONS: Our studies demonstrate that betaine-rich SBM is able to prevent survival reduction caused by Hcy in C. elegans in dependence on hsp-16.2 and daf-16 but independent of the remethylation pathway.

Betaine reduces ß-amyloid-induced paralysis through activation of cystathionine-ß-synthase in an Alzheimer model of Caenorhabditis elegans.

BACKGROUND: The neurodegenerative disorder Alzheimer's disease is caused by the accumulation of toxic aggregates of ß-amyloid in the human brain. On the one hand, hyperhomocysteinemia has been shown to be a risk factor for cognitive decline in Alzheimer's disease. On the other hand, betaine has been demonstrated to attenuate Alzheimer-like pathological changes induced by homocysteine. It is reasonable to conclude that this is due to triggering the remethylation pathway mediated by betaine-homocysteine-methyltransferase. In the present study, we used the transgenic Caenorhabditis elegans strain CL2006, to test whether betaine is able to reduce ß-amyloid-induced paralysis in C. elegans. This model expresses human ß-amyloid 1-42 under control of a muscle-specific promoter that leads to progressive, age-dependent paralysis in the nematodes. RESULTS: Betaine at a concentration of 100 µM was able to reduce homocysteine levels in the presence and absence of 1 mM homocysteine. Simultaneously, betaine both reduced normal paralysis rates in the absence of homocysteine and increased paralysis rates triggered by addition of homocysteine. Knockdown of cystathionine-ß-synthase using RNA interference both increased homocysteine levels and paralysis. Additionally, it prevented the reducing effects of betaine on homocysteine levels and paralysis. CONCLUSION: Our studies show that betaine is able to reduce homocysteine levels and ß-amyloid-induced toxicity in a C. elegans model for Alzheimer's disease. This effect is independent of the remethylation pathway but requires the transsulfuration pathway mediated by cystathionine-ß-synthase.

Resveratrol reduces amyloid-beta (Aß1₋42)-induced paralysis through targeting proteostasis in an Alzheimer model of Caenorhabditis elegans.

PURPOSE: Resveratrol is a polyphenol present in red wine for which the capability of directly interfering with the hallmark of Alzheimer's disease (AD), i.e. toxic ß-amyloid protein (Aß) aggregation, has been shown recently. Since the stimulation of proteostasis could explain reduced Aß-aggregation, we searched for proteostasis targets of resveratrol. METHODS: The transgenic Caenorhabditis elegans strain CL2006, expressing Aß1-42 under control of a muscle-specific promoter and responding to Aß-toxicity with paralysis, was used as a model. Target identification was accomplished through specific knockdowns of proteostasis genes by RNA interference. Effects of resveratrol on protein aggregation were identified using ProteoStat(®) Detection Reagent, and activation of proteasomal degradation by resveratrol was finally proven using a specific fluorogenic peptide substrate. RESULTS: Resveratrol at a concentration of 100 µM caused a 40 % decrease in paralysis. UBL-5 involved in unfolded protein response (UPR) in mitochondria proved to be necessary for the prevention of Aß-toxicity by resveratrol. Also XBP-1, which represents an endoplasmic reticulum-resident factor involved in UPR, was identified to be necessary for the effects of resveratrol. Regarding protein degradation pathways, the inhibition of macroautophagy and chaperone-mediated autophagy prevented resveratrol from reducing paralysis as did the inhibition of proteasomal degradation. Finally, resveratrol reduced the amount of lysosomes, suggesting increased flux of proteins through the autophagy pathways and activated proteasomal degradation. CONCLUSIONS: Resveratrol reduces the Aß-induced toxicity in a C. elegans model of AD by targeting specific proteins involved in proteostasis and thereby reduces the amount of aggregated Aß.

A catechin-enriched green tea extract prevents glucose-induced survival reduction in Caenorhabditis elegans through sir-2.1 and uba-1 dependent hormesis.

Hyperglycemia is a hallmark of diabetes mellitus which leads to the onset of complications in the long term. Green tea through its high content of polyphenolic catechins, on the other hand, is suggested to prevent or at least delay such detrimental complications. In the present study we fed the nematode Caenorhabditis elegans on a liquid medium supplemented with 10mM glucose in the absence or presence of a catechin-enriched green tea extract (CEGTE). After exposure of young adults for 48h survival was subsequently measured under heat stress at 37°C. Whereas CEGTE at 0.01% did not affect the survival of wild type nematodes, it completely reversed the glucose-induced survival reduction. Those effects were not achieved through the monomeric catechins included in CEGTE. RNA interference (RNAi) for sir-2.1 not only prevented the survival extension by CEGTE under simultaneous glucose exposure but also caused a further reduction of survival. Likewise, the knockdown of uba-1, encoding the only E1-ubiquitin-activating enzyme in C. elegans, proved that UBA-1 is essential for the survival extension by CEGTE and that its loss of function changes CEGTE from a survival extending into a survival reducing extract. Stimulation of the proteasome by CEGTE was finally proven through measurements of the proteolytic cleavage of a fluorogenic peptide substrate. To conclude, our studies provide evidence that CEGTE reverses glucose-induced damage in C. elegans through activation of adaptive responses mediated by SIR-2.1 and proteasomal degradation. The hormetic mode of action is revealed by a reduction of survival once the adaptive processes were blocked.

The polyphenol quercetin protects the mev-1 mutant of Caenorhabditis elegans from glucose-induced reduction of survival under heat-stress depending on SIR-2.1, DAF-12, and proteasomal activity.

SCOPE: Hyperglycemia is a hallmark of diabetes mellitus but slighter increases of blood glucose levels are observed also during ageing. Using the Caenorhabditis elegans mev-1 mutant, we identified molecular mechanisms underlying the protection from glucose toxicity by the polyphenol quercetin. METHODS AND RESULTS: We fed C. elegans mev-1 mutants on a liquid medium supplemented with 10 mM glucose, which resulted in a reduced survival at 37°C. The polyphenol quercetin (1 µM) was able to prevent glucose-induced lifespan reduction completely. RNA interference revealed that the sirtuin SIR-2.1, the nuclear hormone receptor DAF-12, and its putative co-activator MDT-15 were critical for the quercetin effects. Moreover, RNA interference for key factors of proteostasis reduced survival, which was not further affected by glucose or quercetin, suggesting that those proteins are a target for both substances. Besides unfolded protein response, proper functionality of the proteasome was shown to be crucial for the survival enhancing effects of quercetin and the polyphenol was finally demonstrated to activate proteasomal degradation. CONCLUSION: Our studies demonstrate that lowest concentrations of quercetin prevent a glucose-induced reduction of survival. SIR-2.1, DAF-12, and MDT-15 were identified as targets that activate unfolded protein response and proteasomal degradation to limit the accumulation of functionally restricted proteins.