The correlates of neonatal complement component 3 and 4 protein concentrations with a focus on psychiatric and autoimmune disorders

Abstract

Complement components have been linked to schizophrenia and autoimmune disorders. We examined the association between neonatal circulating C3 and C4 protein concentrations in 68,768 neonates and the risk of six mental disorders. We completed genome-wide association studies (GWASs) for C3 and C4 and applied the summary statistics in Mendelian randomization and phenome-wide association studies related to mental and autoimmune disorders. The GWASs for C3 and C4 protein concentrations identified 15 and 36 independent loci, respectively. We found no associations between neonatal C3 and C4 concentrations and mental disorders in the total sample (both sexes combined); however, post-hoc analyses found that a higher C3 concentration was associated with a reduced risk of schizophrenia in females. Mendelian randomization based on C4 summary statistics found an altered risk of five types of autoimmune disorders. Our study adds to our understanding of the associations between C3 and C4 concentrations and subsequent mental and autoimmune disorders.

Keywords: C3; C4; Mendelian randomization; autoimmune disorders; complement component; dried blood spots; genome-wide association study; phenome-wide association study; psychiatric disorders; schizophrenia.

Graphical abstract

Figure 1

Concise summary of methods (1) iPSYCH 2012 case-cohort study: N (protein concentration) = 68,768; N (genotype) = 75,764. (2) iPSYCH 2015 expanded sample provided additional genotyped samples: N ∼ 56,000 samples. (3) C4A and C4B isotypes with or without HERV insertions were imputed (long and short, respectively) results in four alleles (AL, AS, BL, BS). See Tables S2 and S3. (4) Based on the imputed C4A, C4B, and HERV allele count, copy numbers were estimated. See Figure 3. (5) Based on Sekar et al., brain C4A expression was estimated based on imputed C4 haplotypes. (6) We examined the association between variants associated with C4 and C3 protein concentration and mRNA expression in GTEx datasets, using summary-data-based Mendelian randomization (SMR). See Tables S12 and S16. (7) We examined the bidirectional association between variants associated with C4 and C3 protein concentration and summary statistics for a range of outcomes (see below) using generalized summary-data-based Mendelian randomization (GSMR). See Tables S22–S26. (8) We examined the association between variants associated with C4 and C3 protein concentration and a range of phenotypes identified in the UK Biobank. See Tables S27 and S28, and Figures S39 and S40. (9) The eight mental and neurological disorders were schizophrenia (SCZ), depression (DEP), bipolar disorder (BIP), autism spectrum disorder (ASD), attention deficit/hyperactivity disorder (ADHD), anorexia nervosa (AN), Alzheimer’s disease (ALZ), and amyotrophic lateral sclerosis (ALS). (10) The six autoimmune disorders were multiple sclerosis (MS), type 1 diabetes (T1D), Crohn’s disease (CD), ulcerative colitis (UC), rheumatoid arthritis (RhA), and systemic lupus erythematosus (SLE).

Figure 2

GWASs of neonatal C4 and C3 protein concentrations (A) Unadjusted C4 protein concentration, (B) C4 protein concentration adjusted for the cis-pQTLs from COJO (fitted as covariates in the regression model), (C) unadjusted C3 protein concentration, and (D) C3 protein concentration adjusted for the cis-pQTLs from COJO. The COJO SNPs fitted as covariates in GWAS of adjusted protein concentration (B and D) were identified from GCTA-COJO of unadjusted protein concentration. The COJO SNPs are highlighted with red triangles. The location of the C3 (on chromosome 19) and C4 (on chromosome 6) genes are highlighted in the relevant panels. The top-associated SNPs were annotated with their overlapped or nearest genes. The GWAS threshold was 5.0 × 10⁻⁸.

Figure 3

Plot of C4 copy number versus C4 protein concentration The center line shows the median value, and the lower and upper edges of the box shows the 25^th and 75^th quantiles respectively. The upper and lower whiskers extend 1.5 times the interquartile range from the top and bottom of the box. Individual outliers are shown as dots. There were three C4 alleles: (A) C4A, (B) C4B, and (C) C4L/HERV. The colors represent C4 allele counts.

Figure 4

Locus plots for GWASs of C4 protein concentration and three mental disorders The four panels show the differences in locations and distributions of the top-associated GWAS SNPs in the MHC region between C4 protein concentration (A) and three mental disorders, SCZ (B), BIP (C), and DEP (D). The y axis in the plots are squared Z scores from GWASs of (A) C4 protein concentration in iPSYCH2012, and −log₁₀(p values) from GWASs of (B) SCZ, (C) BIP, and (D) DEP. The GWAS summary statistics of these three mental disorders were from the latest meta-analysis. The p values from GWAS of C4 protein at the cis-SNPs were extremely small. Squared Z scores (i.e., Z = b/SE) were used to show the significance at SNPs. The GWAS threshold was 5 × 10⁻⁸, and the equivalent squared Z-score threshold was 29.8. The quasi-independent SNPs are highlighted with red triangles. The quasi-independent SNPs from GWAS of C4 were conducted from COJO (Table S7). The quasi-independent SNPs for the three types of mental disorders were based on those reported in the related publications.^,, The red dashed line represents the position of C4 gene.

Figure 5

Forward GSMR results of C4 protein concentration The GSMR analyses examined the relationships between circulating C4 protein concentration versus psychiatric disorders (A and B) and autoimmune disorders (C and D), using the unadjusted C4 GWAS. The pleiotropic SNPs identified by HEIDI-outlier were excluded in (A) and (C). All genome-wide significant SNPs were used in (B) and (D). The dot symbols show the estimates, and 95% confidence intervals are provided. The red dots represent the significant results with p value of GSMR <1.8 × 10⁻³, the Bonferroni-corrected threshold.

Figure 6

Summary of the results from GWASs of neonatal C3 and C4 protein concentrations displayed within the complement cascade For significant loci identified from COJO, proteins encoded by annotated genes are highlighted in red.