Whole-Exome Sequencing Reveals TopBP1 as a Novel Gene in Idiopathic Pulmonary Arterial Hypertension

Abstract

Rationale: Idiopathic pulmonary arterial hypertension (IPAH) is a life-threatening disorder characterized by progressive loss of pulmonary microvessels. Although mutations in the bone morphogenetic receptor 2 (BMPR2) are found in 80% of heritable and z15% of patients with IPAH, their low penetrance (z20%) suggests that other unidentified genetic modifiers are required for manifestation of the disease phenotype. Use of whole-exome sequencing (WES) has recently led to the discovery of novel susceptibility genes in heritable PAH, but whether WES can also accelerate gene discovery in IPAH remains unknown.Objectives: To determine whether WES can help identify novel gene modifiers in patients with IPAH.Methods: Exome capture and sequencing was performed on genomic DNA isolated from 12 unrelated patients with IPAH lacking BMPR2 mutations. Observed genetic variants were prioritized according to their pathogenic potential using ANNOVAR. Measurements and Main Results:A total of nine genes were identified as high-priority candidates. Our top hit was topoisomerase DNA binding II binding protein 1 (TopBP1), a gene involved in the response to DNA damage and replication stress. We found that TopBP1 expression was reduced in vascular lesions and pulmonary endothelial cells isolated from patients with IPAH. Although TopBP1 deficiency made endothelial cells susceptible to DNA damage and apoptosis in response to hydroxyurea, its restoration resulted in less DNA damage and improved cell survival.Conclusions: WES led to the discovery of TopBP1, a gene whose deficiency may increase susceptibility to small vessel loss in IPAH. We predict that use of WES will help identify gene modifiers that influence an individual's risk of developing IPAH.

PPARγ Interaction with UBR5/ATMIN Promotes DNA Repair to Maintain Endothelial Homeostasis

“…We used prolonged HU treatment to induce replication stress-dependent double-strand breaks and, hence, ATM signaling. HU-induced damage also closely resembles chronic replication-induced genotoxic insults associated with genomic instability in vascular ECs from PAH patients (Aldred et al, 2010; de Jesus Perez et al, 2014). In both 293T and ECs, increased ATMIN resulting from silencing PPARγ or UBR5 inhibited ATM signaling.…”

“…PPARγ/UBR5-dependent ATM signaling was also evident in response to DoxR treatment (Figure S3B). We further investigated the role of PPARγ in HU-induced DNA damage because replication stress damage is relevant to PAH (de Jesus Perez et al, 2014). …”

“…We showed that PPARγ promotes UBR5 ubiquitin ligase activity and regulates ATM interactor (ATMIN) levels, thereby permitting efficient ATM phosphorylation and the initiation of DNA repair upon DNA damage. Perturbation of this axis is observed in PAH and can account for unresolved DNA damage that is associated with impaired endothelial functions (de Jesus Perez et al, 2014; Diebold et al, 2015). …”

Identification of a novel truncating mutation in PALB2 gene by a multigene sequencing panel for mutational screening of breast cancer risk‐associated and related genes

“…One patient carried the c.1366G>A transition in TERF2 (p.Glu456Lys, rs150757154), reported as a variation causing potential impaired function (prediction score ≥0.50) by Wang and colleagues . On the other hand, the BARD1 rs1048108 identified in 4 of 34 cases studied and has been associated with medium‐high risk of sporadic neuroblastoma and low BC risk, and the TOpBP1 rs55633281 found in 8 of 34 cases, located in close proximity to the transactivation domain of the encoded protein, might be critical for cellular functions under stress conditions . We also found several variants with conflicting interpretation of pathogenicity and/or lacking sufficient information for classification, such as BLM rs183176301 (2 of 34 cases), CHEK2 rs200050883 (1 of 34 cases), FAM175A rs12642536 (11 of 34 cases), and RAD51C rs773998134 (1 of 34 cases).…”

Novel approaches to pulmonary arterial hypertension drug discovery

“…The contribution of these genes to PAH is less well understood but appears to be related to regulation of cell growth and survival in the pulmonary arteries. Recent studies using whole exome sequencing in families with PAH who are BMPR2 mutation negative have identified novel genes such as caveolin-1, TopBP1 and KCNK3 whose biological role in disease pathobiology is currently under study [22–24]. …”