Fine mapping and conditional analysis identify a new mutation in the autoimmunity susceptibility gene BLK that leads to reduced half-life of the BLK protein

• Published in: Annals of the rheumatic diseases Vol. 71; no. 7; pp. 1219 - 1226
• Main Authors: Delgado-Vega, Angélica M, Dozmorov, Mikhail G, Quirós, Manuel Bernal, Wu, Ying-Yu, Martínez-García, Belén, Kozyrev, Sergey V, Frostegård, Johan, Truedsson, Lennart, de Ramón, Enrique, González-Escribano, María F, Ortego-Centeno, Norberto, Pons-Estel, Bernardo A, D'Alfonso, Sandra, Sebastiani, Gian Domenico, Witte, Torsten, Lauwerys, Bernard R, Endreffy, Emoke, Kovács, László, Vasconcelos, Carlos, da Silva, Berta Martins, Wren, Jonathan D, Martin, Javier, Castillejo-López, Casimiro, Alarcón-Riquelme, Marta E
• Format: Journal Article
• Language: English
• Published: London BMJ Publishing Group Ltd and European League Against Rheumatism 01.07.2012; BMJ Publishing Group; BMJ Publishing Group LTD; BMJ Group
• Series: Extended report
• Subjects: Basic and Translational Research; Biological and medical sciences; Biomarkers - metabolism; Chromosome Mapping; Clinical Medicine; Diseases of the osteoarticular system; Gene expression; Gene Expression Regulation, Enzymologic; Genealogy; Genetic Markers - genetics; Genetic Predisposition to Disease; Genomes; Genotype; Half-Life; Haplotypes; HEK293 Cells; Humans; Hypotheses; Kinases; Klinisk medicin; Lupus; Lupus Erythematosus, Systemic - genetics; Medical and Health Sciences; Medical sciences; Medicin och hälsovetenskap; Mutation; NF-kappa B - metabolism; Polymorphism, Single Nucleotide; Protein Binding; Protein Stability; Proteins; Quality control; Reumatologi och inflammation; Rheumatology and Autoimmunity; src-Family Kinases - genetics; src-Family Kinases - metabolism; Stratigraphy; Transcription factors; Transfection; California; Autoimmunity; Cartography; Mutation; Protein; Rheumatology; Half life
• ISSN: 0003-4967; 1468-2060; 1468-2060
• DOI: 10.1136/annrheumdis-2011-200987

Objectives To perform fine mapping of the autoimmunity susceptibility gene BLK and identify functional variants involved in systemic lupus erythematosus (SLE). Methods Genotyping of 1163 European SLE patients and 1482 controls and imputation were performed covering the BLK gene with 158 single-nucleotide polymorphisms. Logistic regression analysis was done using PLINK and conditional analyses using GENABEL's test score. Transfections of BLK constructs on HEK293 cells containing the novel mutation or the wild type form were analysed for their effect on protein half-life using a protein stability assay, cycloheximide and western blot. CHiP-qPCR for detection of nuclear factor κ B (NFkB) binding. Results Fine mapping of BLK identified two independent genetic effects with functional consequences: one represented by two tightly linked associated haplotype blocks significantly enriched for NFκB-binding sites and numerous putative regulatory variants whose risk alleles correlated with low BLK mRNA levels. Binding of NFkBp50 and p65 to an associated 1.2 Kb haplotype segment was confirmed. A second independent genetic effect was represented by an Ala71Thr, low-frequency missense substitution with an OR=2.31 (95% CI 1.38 to 3.86). The 71Thr decreased BLK protein half-life. Conclusions These results show that rare and common regulatory variants in BLK are involved in disease susceptibility and both, albeit independently, lead to reduced levels of BLK protein.