Alzheimer's disease-specific transcriptomic and epigenomic changes in the tryptophan catabolic pathway

Abstract

Background: Neurodegenerative disorders, including Alzheimer's disease (AD), have been linked to alterations in tryptophan (TRP) metabolism. However, no studies to date have systematically explored changes in the TRP pathway at both transcriptional and epigenetic levels. This study aimed to investigate transcriptomic, DNA methylomic (5mC) and hydroxymethylomic (5hmC) changes within genes involved in the TRP and nicotinamide adenine dinucleotide (NAD) pathways in AD, using three independent cohorts.

Methods: DNA derived from post-mortem middle temporal gyrus (MTG) tissue from AD patients (n = 45) and age-matched controls (n = 35) was analyzed, along with DNA derived from blood samples from two independent cohorts: the German Study on Ageing, Cognition, and Dementia in Primary Care Patients (AgeCoDe) cohort (n = 96) and the Dutch BioBank Alzheimer Center Limburg (BBACL) cohort (n = 262). Molecular profiling, including assessing mRNA expression and DNA (hydroxy)methylation levels, was conducted using HumanHT-12 v4 Expression BeadChip and HM 450 K BeadChip arrays, respectively. Functional interactions between genes and identification of common phenotype-specific positive and negative elementary circuits were conducted using computational modeling, i.e. gene regulatory network (GRN) and network perturbational analysis. DNA methylation of IDO2 (cg11251498) was analyzed using pyrosequencing.

Results: Twelve TRP- and twenty NAD-associated genes were found to be differentially expressed in the MTG of AD patients. Gene sets associated in the kynurenine pathway, the most common TRP pathway, and NAD pathway, showed enrichment at the mRNA expression level. Downstream analyses integrating data on gene expression, DNA (hydroxy)methylation, and AD pathology, as well as GRN and network perturbation analyses, identified IDO2, an immune regulatory gene, as a key candidate in AD. Notably, one CpG site in IDO2 (cg11251498) exhibited significant methylation differences between AD converters and non-converters in the AgeCoDe cohort.

Conclusion: These findings reveal substantial transcriptional and epigenetic alterations in TRP- and NAD-pathway-associated genes in AD, highlighting IDO2 as a key candidate gene for further investigation. These genes and their encoded proteins hold potential as novel biomarkers and therapeutic targets for AD.

Keywords: Alzheimer’s disease (AD); Blood; Brain; Epigenetics; Indoleamine 2,3-dioxygenase (IDO); Tryptophan (TRP).

Fig. 1

TRP and NAD pathway associated genes show significant differential gene expression. Shown are all the genes in the TRP and NAD pathway with up (red) or down (blue) regulation in the middle temporal gyrus of AD patients compared to age-matched controls. Differential expression is presented as log2 fold change. Differential regulation was assessed using false discovery rate (FDR) test and q < 0.05 was considered significant. * q < 0.05, ** q < 0.01 and *** q < 0.001. Error bars represent standard error of mean (SEM)

Fig. 2

TRP and NAD pathway associated genes show significant differential DNA (hydroxy)methylation levels. Shown are significant CpG sites with its corresponding gene name in the tryptophan metabolic pathway (A-C) and NAD pathway (D-F) with up (red) or down (blue) regulation in the middle temporal gyrus of AD patients compared to age-matched controls. A, D Differential methylation (5mC) expression. (B, E) Differential hydroxymethylation (5hmC) expression. C, F Differential unmodified (5uC) expression. Differential expression is presented as log2 fold change. Differential regulation was assessed using limma differential DNA (hydroxy)methylation level analysis and p < 0.05 was considered significant. * p < 0.05 and ** p < 0.01. Error bars represent standard error of mean (SEM)

Fig. 3

Overview of the differentially expressed genes and DNA (hydroxy)methylation levels within the TRP metabolic pathway and NAD pathway. Within the TRP metabolic pathway, a total of 12 genes exhibited significant differential gene expression, while 20 genes were differentially expressed within the NAD pathway. Genes exhibiting up-regulation in expression are highlighted with red box, whereas those with down-regulation are indicated by blue box. Genes with altered DNA (hydroxy)methylation levels are denoted by circular markers, with distinct colors representing specific modifications: 5mC (green), 5hmC (purple), and 5uC (orange)

Fig. 4

Spearman’s correlation analysis of TRP pathway. Spearman’s correlation analysis between methylation (■5hmC and ▲5uC) and mRNA expression levels. Spearman’s correlation analysis are presented with Spearman’s R-value, 95% confidence interval (CI), and p-value

Fig. 5

Spearman’s correlation analysis of NAD pathway. Spearman’s correlation analysis between methylation (●5mC, ■5hmC, and ▲5uC) and mRNA expression levels. Spearman’s correlation analysis are presented with spearman R-value, 95% confidence interval and, p-value

Fig. 6

Gene regulatory network (GRN) of TRP and NAD pathway. A TRP GRN representing the control phenotype and containing 22 nodes and 30 interactions, (B) TRP GRN representing Alzheimer’s disease (AD) phenotype and containing 22 nodes and 25 interactions, (C) NAD GRN representing the control phenotype and containing 13 nodes and 21 interactions, (D) NAD GRN representing Alzheimer’s disease (AD) phenotype and containing 14 nodes and 24 interactions. Green line indicates gene activation while red line indicates gene inhibition