Mutations of <scp><i>NOTCH3</i></scp> in childhood pulmonary arterial hypertension

<jats:title>Abstract</jats:title><jats:p>Mutations of <jats:styled-content style="fixed-case"><jats:italic>BMPR2</jats:italic></jats:styled-content> and other <jats:styled-content style="fixed-case">TGF</jats:styled-content>‐<jats:italic>β</jats:italic> superfamily genes have been reported in pulmonary arterial hypertension (<jats:styled-content style="fixed-case">PAH</jats:styled-content>). However, 60–90% of idiopathic <jats:styled-content style="fixed-case">PAH</jats:styled-content> cases have no mutations in these genes. Recently, the expression of NOTCH3 was shown to be increased in the pulmonary artery smooth muscle cells of <jats:styled-content style="fixed-case">PAH</jats:styled-content> patients. We sought to investigate <jats:styled-content style="fixed-case"><jats:italic>NOTCH3</jats:italic></jats:styled-content> and its target genes in <jats:styled-content style="fixed-case">PAH</jats:styled-content> patients and clarify the role of NOTCH3 signaling. We screened for mutations in <jats:styled-content style="fixed-case"><jats:italic>NOTCH3</jats:italic></jats:styled-content>,<jats:styled-content style="fixed-case"> <jats:italic>HES1</jats:italic></jats:styled-content>, and <jats:styled-content style="fixed-case"><jats:italic>HES5</jats:italic></jats:styled-content> in 41 <jats:styled-content style="fixed-case">PAH</jats:styled-content> patients who had no mutations in <jats:styled-content style="fixed-case"><jats:italic>BMPR2</jats:italic></jats:styled-content>,<jats:styled-content style="fixed-case"> <jats:italic>ALK1</jats:italic></jats:styled-content>,<jats:styled-content style="fixed-case"> <jats:italic>endoglin</jats:italic></jats:styled-content>,<jats:styled-content style="fixed-case"> <jats:italic>SMAD1/4/8</jats:italic></jats:styled-content>,<jats:styled-content style="fixed-case"> <jats:italic>BMPR1B</jats:italic></jats:styled-content>, or <jats:styled-content style="fixed-case"><jats:italic>Caveolin‐1</jats:italic></jats:styled-content>. Two novel missense mutations (c.2519 G>A p.G840E, c.2698 A>C p.T900P) in <jats:styled-content style="fixed-case"><jats:italic>NOTCH3</jats:italic></jats:styled-content> were identified in two <jats:styled-content style="fixed-case">PAH</jats:styled-content> patients. We performed functional analysis using stable cell lines expressing either wild‐type or mutant NOTCH3. The protein‐folding chaperone GRP78/BiP was colocalized with wild‐type NOTCH3 in the endoplasmic reticulum, whereas the majority of GRP78/BiP was translocated into the nuclei of cells expressing mutant NOTCH3. Cell proliferation and viability were higher for cells expressing mutant NOTCH3 than for those expressing wild‐type NOTCH3. We identified novel <jats:styled-content style="fixed-case"><jats:italic>NOTCH3</jats:italic></jats:styled-content> mutations in <jats:styled-content style="fixed-case">PAH</jats:styled-content> patients and revealed that these mutations were involved in cell proliferation and viability. NOTCH3 mutants induced an impairment in NOTCH3‐HES5 signaling. The results may contribute to the elucidation of <jats:styled-content style="fixed-case">PAH</jats:styled-content> pathogenesis.</jats:p>