Post-GWAS functional studies reveal an RA-associated CD40-induced NF-kB signal transduction and transcriptional regulation network targeted by class II HDAC inhibitors

Currently, it remains difficult to identify which single nucleotide polymorphisms (SNPs) identified by genome-wide association studies (GWAS) are functional and how various functional SNPs (fSNPs) interact and contribute to disease susceptibility. GWAS have identified a CD40 locus that is associated with rheumatoid arthritis (RA). We previously used two techniques developed in our laboratory, single nucleotide polymorphism-next-generation sequencing (SNP-seq) and flanking restriction enhanced DNA pulldown-mass spectrometry (FREP-MS), to determine that the RA risk gene RBPJ regulates CD40 expression via a fSNP at the RA-associated CD40 locus. In the present work, by applying the same approach, we report the identification of six proteins that regulate RBPJ expression via binding to two fSNPs on the RA-associated RBPJ locus. Using these findings, together with the published data, we constructed an RA-associated signal transduction and transcriptional regulation network (STTRN) that functionally connects multiple RA-associated risk genes via transcriptional regulation networks (TRNs) linked by CD40-induced nuclear factor kappa B (NF-kB) signaling. Remarkably, this STTRN provides insight into the potential mechanism of action for the histone deacetylase inhibitor givinostat, an approved therapy for systemic juvenile idiopathic arthritis. Thus, the generation of disease-associated STTRNs based on post-GWAS functional studies is demonstrated as a novel and effective approach to apply GWAS for mechanistic studies and target identification.