The schizophrenia-associated variant in SLC39A8 alters N-glycosylation of critical proteins in the mouse brain

Robert G. Mealer MD, PhD

Massachusetts General Hospital
The schizophrenia-associated variant in SLC39A8 alters N-glycosylation of critical proteins in the mouse brain

Scientific Abstract

Background: Multiple glycosylation genes are associated with schizophrenia through GWAS, including a missense mutation (A391T) in the manganese transporter SLC39A8. Many glycosylation enzymes require manganese as a cofactor, and human carriers of A391T have reduced serum manganese, altered plasma glycosylation, and brain MRI changes consistent with altered metal transport. However, the molecular connection between the A391T mutation and schizophrenia risk in the brain remains unknown.

Methods: We investigated brain glycosylation changes in a knock-in mouse model homozygous for the A391T mutation. Different brain regions (cortex, hippocampus, striatum, and cerebellum) from male and female mice (N > 4 per group) were analyzed using MALDI-TOF MS glycomics, RNAseq, and glycoproteomics. Statistical analyses between groups was performed using t-tests while correcting for multiple comparisons on the transcriptome and proteome level.

Results: Mice homozygous for A391T display several changes in brain glycosylation, with N- linked glycosylation most significantly impaired. RNAseq analysis showed negligible variation, consistent with changes in the activity of glycosylation enzymes rather than gene expression. One third of all detected glycoproteins were differentially N-glycosylated in the cortex, including members of several pathways previously implicated in schizophrenia such as cell adhesion molecules and neurotransmitter receptors. Differentially glycosylated proteins originated from all cell types in the brain despite Slc39a8 expression being restricted to endothelial cells, suggesting it serves as a gate-keeper for brain manganese at the blood brain barrier.

Conclusions: These findings provide a mechanistic link between the A391T variant and biochemical changes in the brain, furthering our molecular understanding of how this validated risk-allele may contribute to the pathophysiology of schizophrenia. As many glycosylation deficiencies are treated through oral supplementation with enzymatic precursors and cofactors, brain glycosylation changes caused by the A391T mutation may be reversible with manganese supplementation.

SoundCloud Transcript

The A391T missense mutation in SLC39A8 is the most significantly associated coding variant identified to date by SCZ GWAS. 

 

SLC39A8 is a Manganese transporter, and many enzymes of glycosylation uniquely require Mn as a cofactor for activity.

 

Glycosylation is critical in the development and function of the nervous system, and involves the covalent attachment of carbohydrates to lipids and proteins.

 

Congenital Disorders of Glycosylation commonly present with neurologic symptoms, and common variants in glycosylation genes have been associated with several neuropsychiatric phenotypes including schizophrenia.

 

We recently reviewed the role of glycosylation in schizophrenia through post-mortem and genetic studies.

 

In this study, we utilize a mouse model expressing the schizophrenia-associated variant in SLC39A8 to investigate its effect on brain glycosylation, hypothesizing that A391T increases scz risk by altering Mn2+ dependent glycosylation.

 

To test this model, we employed a homozygous knock-in A391T mouse, and analyzed glycomics, transcriptomics, and glycoproteomics from several brain regions.

 

Figure 2 summarizes our MALDI-MS glycomics data, showing that A391T mice display significant glycosylation changes that differ based on region. A novel assay of fluorescent glycan derivatization confirmed that A391T mice have a reduced quantity of N-glycans in the CTX.

 

In Fig 3 we present transcriptomic analysis of A391T cortex and cerebellum, showing that gene expression differences cannot account for the observed glycosylation differences.

 

We next performed quantitative N-glycoproteomic analysis of mouse brain cortex as shown in Fig 4. A volcano plot comparing N-glycoproteins between A391T cortex and controls, shows that nearly one third of glycoprotein levels are altered in A391T cortex. Differentially N-glycosylated proteins in A391T cortex originate from nearly all cell types based on single cell expression atlases, and table 2 highlights several criticial glycoproteins with different levels in A391T cortex that have been previously implicated in schizophrenia pathogenesis.

 

In conclusion, our results show that SLC39A8, which is primarily expressed in brain endothelial cells, serves as a gate keeper for brain Mn, and the A391T mutation results broad changes of glycosylation activity across all cell types. These findings confirm A391T affects brain glycosylation and provide further evidence that altered glycosylation is involved in schizophrenia pathogenesis. It will be exciting to determine if these changes can be reversed through Mn supplementation, and an increased appreciation of dyglycosylation in schizophrenia may lead to new diagnostic tests and treatment opportunities.

 

Live Zoom Session – April 21st

research Areas

Authors

Robert G. Mealer MD PhD, Sarah E. Williams BS, Maxence Noel PhD, Bo Yang PhD, Alexandria D’Souza MS, Murat Cetinbas PhD, Ruslan Sadreyev PhD, Edward M. Scolnick MD, Ramnik J. Xavier MD PhD, Christina M. Woo PhD, Richard D. Cummings PhD, and Jordan W. Smoller MD ScD