BMPR2 mutations have short lifetime expectancy in primary pulmonary hypertension.

In a nationwide study, we identified a total of 59 patients diagnosed with primary pulmonary hypertension (PPH) in Finland between the years 1987 and 1999. These data support a minimum estimate for a PPH population prevalence of 5.8 cases/million with an incidence of 0.2-1.3 cases/million/year. The male-to-female ratio among the patients was 1:4, while 7% (4/59) of the PPH probands had a known family history of the disorder. Familial or sporadic PPH showed no geographic clustering to any region of Finland. Sequencing of the coding regions and exon-intron boundaries of the bone morphogenetic protein receptor type 2 (BMPR2) identified heterozygous BMPR2 mutations in 12% (3/26) of the sporadic and 33% (1/3) of the familial patients. All four mutations were different, and two of those have been previously reported in other populations. Pathogenic defects in BMPR2 include a novel missense mutation (c.2696G>C encoding R899P), located within the receptor intracellular cytoplasmic domain whose function has been poorly characterized. Our analysis demonstrates that this mutant, while localizing to the cell surface, does not impact on SMAD-mediated (mothers against decapentaplegic homolog) intracellular signaling, but leads to constitutive activation of the p38(MAPK) pathway. The absence of a founder mutation in a genetically homogeneous population, such as the Finns, suggests that all identified BMPR2 mutations have to be rather young while the ancestral (if any) mutations have been lost either due to repetitive genetic bottlenecks or due to significant negative selection. Hum Mutat 26(2), 1-6, 2005. (c) 2005 Wiley-Liss, Inc.

Functional interaction between BMPR-II and Tctex-1, a light chain of Dynein, is isoform-specific and disrupted by mutations underlying primary pulmonary hypertension.

Diverse heterozygous mutations of bone morphogenetic receptor type II (BMPR-II) underlie the inherited form of the vascular disorder primary pulmonary hypertension (PPH). As yet, the molecular detail of how such defects contribute to the pathogenesis of PPH remains unclear. BMPR-II is a member of the transforming growth factor-beta cell signalling superfamily. Ligand binding induces cell surface receptor complex formation and activates a cascade of phosphorylation events of intracellular intermediaries termed Smads, which initiate transcriptional regulation. Some 30% of PPH-causing mutations localize to exon 12, which may be spliced out forming an isoform depleted of the unusually long BMPR-II cytoplasmic tail. To further elucidate the consequences of BMPR2 mutation, we sought to characterize aspects of the cytoplasmic domain function by seeking intracellular binding partners. We now report that Tctex-1, a light chain of the motor complex dynein, interacts with the cytoplasmic domain of BMPR-II and demonstrate that Tctex-1 is phosphorylated by BMPR-II, a function disrupted by PPH disease causing mutations within exon 12. Finally we show that BMPR-II and Tctex-1 co-localize to endothelium and smooth muscle within the media of pulmonary arterioles, key sites of vascular remodelling in PPH. Taken together, these data demonstrate a discrete function for the cytoplasmic domain of BMPR-II and justify further investigation of whether the interaction with and phosphorylation of Tctex-1 contributes to the pathogenesis of PPH.

Functional analysis of bone morphogenetic protein type II receptor mutations underlying primary pulmonary hypertension.

A wide range of mutations in the type II receptor for bone morphogenetic protein (BMPR-II) have been shown to underlie primary pulmonary hypertension. To determine the mechanism of altered BMPR-II function, we employed transient transfection studies in cell lines and primary cultures of pulmonary vascular smooth muscle cells using green fluorescent protein (GFP)-tagged wild-type and mutant BMPR2 constructs and confocal microscopy to localize receptors. Substitution of cysteine residues in the ligand binding or kinase domain prevented trafficking of BMPR-II to the cell surface, and reduced binding of (125)I-BMP4. In addition, transfection of cysteine-substituted BMPR-II markedly reduced basal and BMP4-stimulated transcriptional activity of a BMP/Smad responsive luciferase reporter gene (3GC2wt-Lux), compared with wild-type BMPR-II, suggesting a dominant-negative effect of these mutants on Smad signalling. In contrast, BMPR-II containing non-cysteine substitutions in the kinase domain were localized to the cell membrane, although these also suppressed the activity of 3GC2wt-Lux. Interestingly, BMPR-II mutations within the cytoplasmic tail trafficked to the cell surface, but retained the ability to activate 3GC2wt-Lux. Transfection of mutant, but not wild-type, constructs into a mouse epithelial cell line (NMuMG cells) led to activation of p38(MAPK) and increased serum-induced proliferation compared with the wild-type receptor, which was partly p38(MAPK)-dependent. We conclude that mutations in BMPR-II heterogeneously inhibit BMP/Smad-mediated signalling by diverse molecular mechanisms. However, all mutants studied demonstrate a gain of function involving upregulation of p38(MAPK)-dependent proproliferative pathways.