Assessment of complement C4 gene copy number using the paralog ratio test

Area of Science:

• Immunogenetics
• Human Genetics
• Molecular Biology

Background:

• The complement C4 locus, within the MHC class III region, exhibits copy number variation.
• Complement C4 null alleles are associated with diseases like systemic lupus erythematosus (SLE).
• Previous studies relied on protein analysis, lacking direct genomic interrogation for C4 null allele status, and accurate gene copy number (GCN) estimation was limited.

Purpose of the Study:

• To develop and validate accurate methods for determining total C4 GCN, C4A GCN, and C4B GCN.
• To assess the range of C4 GCN in human populations.
• To evaluate the utility of SNP genotypes as proxies for C4 GCN.

Main Methods:

• Development of a high-throughput paralog ratio test (PRT) combined with two restriction enzyme digest variant ratio tests (REDVRs).
• Validation against Southern blot analysis, achieving a 9% discrepancy rate.
• Analysis of C4 GCN in the CEU and 1958 British Birth Cohort populations.

Main Results:

• The PRT and REDVRs method accurately and reproducibly determines C4 GCN.
• A wide spectrum of C4 GCN was observed in the studied populations.
• SNP-C4 copy number variation (CNV) analyses showed only moderate correlation with actual C4 GCN.

Conclusions:

• The developed PRT and REDVRs methods provide accurate C4 GCN quantification.
• Significant variation in C4 GCN exists within human populations.
• SNP genotypes are not reliable proxies for complement C4 GCN, necessitating direct GCN measurement for accurate disease association studies.