UCSF

The mechanisms by which transcription factor haploinsufficiency alters the epigenetic and transcriptional landscape in human cells to cause disease are unknown. Here, I utilized human induced pluripotent stem cell (iPSC)-derived endothelial cells (ECs) to show that heterozygous nonsense mutations in NOTCH1 (N1) that cause calcific aortic valve disease (CAVD) disrupt the epigenetic architecture resulting in derepression of latent pro-osteogenic and -inflammatory gene networks. Hemodynamic shear stress, which protects valves from calcification in vivo, activated anti-osteogenic and anti-inflammatory networks in N1+/+, but not N1+/-, iPSC-derived ECs. N1 haploinsufficiency altered H3K27ac at N1-bound enhancers, dysregulating downstream transcription of over 1000 genes involved in osteogenesis, inflammation, and oxidative stress. Computational predictions of the disrupted N1-dependent gene network revealed regulatory nodes that when modulated restored the network toward the N1+/+ state. My results highlight how alterations in transcription factor dosage affect gene networks leading to human disease and reveal nodes for potential therapeutic intervention.

This iPSC model of CAVD provides a platform for screening for small molecules that target central regulatory nodes to alleviate the gene network dysregulation in N1 haploinsufficient cells. However, the lack of an in vivo animal model of CAVD caused by N1 haploinsufficiency severely limits safety and efficacy testing of potential therapeutics. Diseases caused by gene haploinsufficiency in humans commonly lack a phenotype in mice heterozygous for the orthologous factor. Since CAVD is an age-dependent disease, I generated N1+/- mice lacking telomerase activity and showed that telomere shortening reveals heart valve disease in N1+/- mice. Furthermore, N1 haploinsufficiency promoted proliferation that accelerated telomere shortening, potentially past the critical threshold at which valve disease ensues. Gene dysregulation in AVs of N1 haploinsufficient mice with shortened telomeres involved downregulation of osteoclast factors and upregulation of pro-calcific regulatory nodes paralleling gene network alterations in the human iPSC disease model. Concordance between the murine and human models supports that this model may represent an ideal in vivo setting in which to test therapeutics aimed at preventing or delaying the progression of CAVD.