Mutational and epigenetic analysis of the promoter region of BMPR2 gene in patients with pulmonary arterial hypertension

Pulmonary arterial hypertension (PAH; OMIM 178600) is a progressive vascular disorder characterized by obstruction of precapillary pulmonary arteries. Approximately 80% of patients with familiar PAH have a mutation in bone morphogenetic protein receptor type II gene (BMPR2). This gene has a long promoter region with many binding sites of transcription factors.Given the complex and multifactorial nature of PAH, it seems likely that epigenetic events and mutations, in promoter region, could influence the penetrance, establishment or progression of PAH. We included 55 PAH patients and 52 controls. Using specifically designed primers we amplified by PCR and sequenced the 5´-UTR region.We searched for CpG islands in 5000 bp upstream the 5´-UTR region of BMPR2 and genomic DNA was modified with sodium bisulfite in order to establish the methylation pattern of this region. A total of 4 sequence changes were identified in 7.5% of patients with PAH, and none were detected in a panel of 100 chromosomes from controls. These mutations are c.1-301G>A, c.1-279C>A, c.1-212delC and c.1-92C>A, which have appeared alike in idiopathic and associated PAH patients. We found one CpG island of 1300 bp containing numerous CpG sites. To analyse the methylation status within this region we designed 3 pairs of methylation and unmethylation specific primers. No differences in the methylation status were observed between patients and controls. In conclusion, mutations in 5´-UTR region are not frequent, but can cause alterations in gene expression by disrupting the binding site of transcription factors. Methylation does not seem to be related to PAH, at least in the region of BMPR2 studied.