The schizophrenia risk locus in SLC39A8 alters brain metal transport and plasma glycosylation

Abstract

A common missense variant in SLC39A8 is convincingly associated with schizophrenia and several additional phenotypes. Homozygous loss-of-function mutations in SLC39A8 result in undetectable serum manganese (Mn) and a Congenital Disorder of Glycosylation (CDG) due to the exquisite sensitivity of glycosyltransferases to Mn concentration. Here, we identified several Mn-related changes in human carriers of the common SLC39A8 missense allele. Analysis of structural brain MRI scans showed a dose-dependent change in the ratio of T2w to T1w signal in several regions. Comprehensive trace element analysis confirmed a specific reduction of only serum Mn, and plasma protein N-glycome profiling revealed reduced complexity and branching. N-glycome profiling from two individuals with SLC39A8-CDG showed similar but more severe alterations in branching that improved with Mn supplementation, suggesting that the common variant exists on a spectrum of hypofunction with potential for reversibility. Characterizing the functional impact of this variant will enhance our understanding of schizophrenia pathogenesis and identify novel therapeutic targets and biomarkers.

Figure 1

Dose-dependent effects of the A391T variant on regional T2w/T1w ratio. (a) Representative maps of the T2w/T1w ratio in CT and TT carriers overlaid on CC controls where ratio is either increased or decreased with a p value of < 0.05 on a pixel-by-pixel basis via student’s t-test. Heat map corresponds to the direction of change of the T2w/T1w ratio relative to the CC group, with yellow/orange representing an increase and blue representing a decrease. (b) Quantification of ROIs including globus pallidus (GPi), lateral putamen (LPut), and substantia nigra (SN) based on rs13107325 genotype, compared using post-hoc Dunnett’s test. Data points are shown for each individual, with the black horizontal line representing mean for each genotype. CC (white circles) n = 46, CT (gray circles) n = 44, TT (black circles) n = 43.

Figure 2

A391T results in a specific reduction of serum manganese in heterozygous and homozygous carriers. (a) Mn, (b) Co, (c) Cu, and (d) Zn are all trace elements previously shown to be transported by SLC39A8. Data points are shown for each individual, with the black horizontal line representing mean for each genotype. Method Detection Limit (MDL) for each trace element shown as grey dashed line on each graph. Genotypes compared using student’s t-test. CC (white circles) n = 46, CT (gray circles) n = 46, TT (black circles) n = 25. *p < 0.05, **p < 0.01, ***p < 0.001.

Figure 3

Analysis of plasma protein N-glycans based on rs13107325 genotype. (a) Summary data for the 20 most abundant plasma protein N-glycans sorted by rs13107325 genotype. Data presented as mean +/− standard error of the mean (SEM) for the percent (%) abundance of each N-glycan relative to the total N-glycan pool. Corresponding N-glycan structures are shown above each predicted m/z (mass/charge ratio) including a key for individual monosaccharide components of human N-glycans. (b) Heat map illustrating percent change of each individual glycan in CT and TT relative to CC; scaled from dark blue → white → bright red as − 50.0% → 0% →  + 50.0%. Genotypes compared using student’s t-test. Individual N-glycans that are significantly different in CT and TT compared to CC are marked with an asterisk; *p < 0.05. CC (white bars) n = 33, CT (gray bars) n = 31, TT (black bars) n = 25.

Figure 4

A391T carriers have reduced branching of plasma protein N-glycans. Data presented as mean +/− SEM for the % abundance of N-glycans with (a) one (mono-), (b) two (bi-), (c) three (tri-) or (d) four (tetra-) antennas, defined as the number of GlcNAc attachments to core Man residues (Supp. Table 3). Genotypes compared using student’s t-test. CC (white bars) n = 33, CT (gray bars) n = 31, TT (black bars) n = 25.

Figure 5

A391T has a larger effect on branching in male carriers. a, Males. b, Females. Data presented as mean +/− SEM for the percent abundance of N-glycans with one (mono-), two (bi-), three (tri-) or four (tetra-) antennas. Males: CC n = 17, CT n = 15, TT n = 10; Females: CC n = 16, CT n = 16, TT n = 12. *p value < 0.05, **p value < 0.01.

Figure 6

Mn supplementation increases large N-glycans in severe SLC39A8 mutation carriers with CDGs. Partial MALDI-TOF spectrum from plasma/serum of (a) subject A and (b) subject B pre- and post-Mn supplementation are shown. X-axis scaled for m/z of 3,550–3,800 kd and relative signal intensity on the Y-axis.

Figure 7

Plasma protein N-glycan changes in severe loss-of-function SLC39A8 mutation carriers after Mn treatment. Data presented as percent abundance of (a) tri-antennary, (b) tetra-antennary, (c) high-mannose, and (d) bisecting N-glycans before and after ~ 1 year of Mn supplementation in subjects A and B with congenital disorders of glycosylation due to severe SLC39A8 homozygous mutations. Samples from each individual were replicated twice with similar results.