Genetic Analysis of Neurite Outgrowth Inhibitor-Associated Genes in Parkinson's Disease: A Cross-Sectional Cohort Study

Abstract

Background: Parkinson's disease (PD) is a neurodegenerative disease caused by a combination of aging, environmental, and genetic factors. Previous research has implicated both causative and susceptibility genes in PD development. Nogo-A, a neurite outgrowth inhibitor, has been shown to impact axon growth through ligand-receptor interactions negatively, thereby involved in the deterioration of dopaminergic neurons. However, rare genetic studies have identified the relationship between neurite outgrowth inhibitor (Nogo)-associated genes and PD from a signaling pathway perspective.

Methods: We enrolled 3959 PD patients and 2931 healthy controls, categorized into two cohorts based on their family history and age at onset: sporadic early Parkinson's disease & familial Parkinson's disease (sEOPD & FPD) cohort and sporadic late Parkinson's disease (sLOPD) cohort. We selected 17 Nogo-associated genes and stratified them into three groups via their function, respectively, ligand, receptors, and signaling pathway groups. Additionally, we conducted the burden analysis in rare variants, the logistic regression analysis in common variants, and the genotype-phenotype association analysis. Last, bioinformatics analysis and functional experiments were conducted to identify the role of the MTOR gene in PD.

Results: Our findings demonstrated that the missense variants in the MTOR gene might increase PD risk, while the deleterious variants in the receptor subtype of Nogo-associated genes might mitigate PD risk. However, common variants of Nogo-associated genes showed no association with PD development in two cohorts. Furthermore, genotype-phenotype association analysis suggested that PD patients with MTOR gene variants exhibited relatively milder motor symptoms but were more susceptible developing dyskinesia. Additionally, bioinformatics analysis results showed MTOR gene was significantly decreased in PD, indicating a potential negative role of the mTOR in PD pathogenesis. Experimental data further demonstrated that MHY1485, a mTOR agonist, could rescue MPP⁺-induced axon inhibition, further implicating the involvement of mTOR protein in PD by regulating cell growth and axon growth.

Conclusions: Our preliminary investigation highlights the association of Nogo-associated genes with PD onset in the Chinese mainland population and hints at the potential role of the MTOR gene in PD. Further research is warranted to elucidate the mechanistic pathways underlying these associations and their therapeutic implications.

Keywords: Parkinson's disease; axon growth; genotype–phenotype association; neurite outgrowth inhibitor‐associated genes; rare variant.

FIGURE 1

An overview of the Nogo‐related proteins and the pathways they are involved in. Nogo‐A protein can regulate the activity of the RhoA‐ROCK1‐LIMK1 signaling pathway through a variety of ligand‐receptor interactions, including NgR‐LINGO1, NgR‐PlexinA2‐CRMP2, Nogo‐A‐S1PR2, and Nogo‐A‐Integrin αv, which causes actin depolymerization and microtubule decomposition, thereby exerting the function of inhibiting axon growth. Additionally, the Nogo‐A protein can inhibit the activity of TrkB and its downstream PI3K‐Akt–mTOR signal pathway by binding to PirB, thereby restraining protein transcription, translation, and cell proliferation and inducing cell apoptosis.

FIGURE 2

The schematic diagram of mTOR protein and the location of rare missense variants identified in the sEOPD & FPD cohort. The domain of mTOR protein contains HEAT repeats, FRAP‐ATM‐TTRAP (FAT) domain, FRB domain, kinase domain, and FATC domain. Mutations in this study of the MTOR gene are shown above the schematic. Mutation of the MTOR gene is only found in *Patients, ^#Healthy controls, and ^{^}Both patients and healthy controls.

FIGURE 3

Identification of the involvement of mTOR in Parkinson's disease via the GEO database. (A) Venn diagram of DEGs from the two PD datasets, with p‐adjusted < 0.05 and∣logFC∣ > 0.15 as the threshold. (B) KEGG pathway enrichment analysis (TOP 20) of the overlapping DEGs (5261 genes) in PD, with BH correction. (C) GO enrichment analysis (TOP 20) of the overlapping DEGs (5261 genes) in PD, with BH correction. (D) The relative mRNA expression of the MTOR gene in the two PD databases, with t‐tests. **p < 0.01, and ****p < 0.0001.

FIGURE 4

mTOR protein is involved in Parkinson's disease by regulating cell growth and axon growth. After differentiation via RA (10 μM, 7 days), SH‐SY5Y cells were treated with MPP⁺ (500 μM, 24 h), Rapamycin (2.5 μM, 18 h), or MHY1485 (10 μM, 18 h) to detect neurite length and cell death. (A) The procedure of cell plating and drug handling. (B) This bright field image of cells was taken, respectively, at 0 and 24 h with MPP⁺ treatment. Scale bar, 100 μm. (C) SH‐SY5Y cells were stained with Tuj1 (green), mitoTracker (red), and Hoechst (blue). (D and E) the quantification of axon length (Tuj1) and mitoTracker intensity (mitoTracker), analyzed by ImageJ and visualized by GraphPad Prism 9. Scale bar, 20 μm. Data were presented as the mean ± SD. ****p < 0.0001, **p < 0.01, ns, no different significance.