Gegg, M. E. | bioRxiv Science

bioRxiv2023

α-synuclein expression in response to bacterial ligands and metabolites in gut enteroendocrine cells

BackgroundCaudo-rostral migration of pathological forms of -synuclein from the gut to the brain is proposed as an early feature in Parkinson disease (PD) pathogenesis, but the underlying mechanisms remain unknown. Intestinal enteroendocrine cells sense and respond to numerous luminal signals, including bacterial factors, and transmit this information to the brain via the enteric nervous system and vagus nerve. There is evidence that gut bacteria composition and their metabolites change in PD patients and these alterations can trigger -synuclein pathology in animal models. ObjectiveHere we investigated the effect of toll-like receptor (TLR) and free fatty acid receptor (FFA2/3) agonists on -synuclein levels in mouse STC-1 enteroendocrine cells. MethodsSTC-1 cells were treated with TLR and FFA2/3 agonists alone and in combination with selective antagonists. The level of -synuclein protein was measured in cell lysates and cell culture media by western blot and ELISA. And the level of -synuclein and tumour necrosis factor (TNF) mRNA was measured by quantitative RT-PCR. ResultsTLR and FFA receptor agonists significantly increased intracellular and extracellular -synuclein levels and antagonists significantly reduced these effects. TLR and FFA receptor agonists also significantly increased TNF transcription and this was inhibited by corresponding antagonists. ConclusionsElevated intracellular -synuclein increases the likelihood of aggregation and conversion to toxic forms. Factors derived from bacteria induce -synuclein accumulation in STC-1 cells. Here we provide support for a mechanism by which exposure of enteroendocrine cells to specific bacterial factors found in PD gut dysbiosis might facilitate accumulation and transmission of -synuclein pathology from the gut to the brain.

neuroscience↗

bioRxiv2022

The GBA variant E326K is associated with alpha-synuclein aggregation and lipid droplet accumulation in human cell lines.

Sequence variants or mutations in the GBA gene are numerically the most important risk factor for Parkinson disease (PD). The GBA gene encodes for the lysosomal hydrolase enzyme, glucocerebrosidase (GCase). GBA mutations often reduce GCase activity and lead to impairment of the autophagy-lysosomal pathway, which is important in the turnover of alpha-synuclein, accumulation of which is a key pathological hallmark of PD. Although the E326K variant is one of the most common GBA variants associated with PD, there is limited understanding of its biochemical effects. We have characterised homozygous and heterozygous E326K variants in human fibroblasts. We found that E326K variants did not cause significant loss of GCase protein or activity, endoplasmic reticulum (ER) retention or ER stress, in contrast to the L444P GBA mutation. This was confirmed in human dopaminergic SH-SY5Y neuroblastoma cell lines over-expressing GCase with either E326K or L444P protein. Despite no loss of GCase activity, a significant increase of insoluble alpha-synuclein aggregates in E326K and L444P mutants was observed. Notably, SH-SY5Y over-expressing E326K demonstrated a significant increase in lipid droplet number under basal conditions, which was exacerbated following treatment with the fatty acid oleic acid. Similarly, a significant increase in lipid droplet formation following lipid loading was observed in heterozygous and homozygous E326K fibroblasts. In conclusion, the work presented here demonstrates that the E326K mutation behaves differently to common loss of function GBA mutations, however lipid dyshomeostasis and alpha-synuclein pathology is still evident.

genetics↗