A study using genome editing to delete large blocks of the genome in human induced pluripotent stem cells (iPSCs) has uncovered new functions of the 9p21.3 risk locus, one of the strongest common genetic risk factors for coronary artery disease (CAD), associated with 10–15% of CAD per year in the USA.

The 9p21.3 risk locus is an ∼60 kb block of single-nucleotide polymorphisms (SNPs) that comprises two haplotypes: risk and non-risk. This region is human-specific and lacks known coding genes, which has hampered the identification of its function. To assess the effects of the risk locus, Lo Sardo et al. used iPSCs derived from patients with CAD who were homozygous for the risk haplotype and from healthy individuals homozygous for the non-risk haplotype. Given that it is unclear which SNPs influence the risk of CAD, the investigators deleted the entire haplotype (known as haplotype editing) in the iPSCs using the TALENs genome editing tool. The iPSCs were then differentiated into vascular smooth muscle cells (VSMCs) to assess the effects in a CAD-relevant cell type.

The VSMCs with the risk haplotype had a phenotype consistent with CAD, including abnormal adhesion, contraction and proliferation, as well as a transcriptional profile with alterations in known CAD risk genes and pathways. Deleting the risk haplotype rescued VSMC proliferation, adhesion and contraction; by contrast, deletion of the non-risk region had a minimal effect on the cells. The 9p21.3 region encodes the terminal exons of the long non-coding RNA ANRIL and, interestingly, expressing ANRIL in non-risk VSMCs induced the phenotypes observed in VSMCs with the risk haplotype. Importantly, Lo Sardo et al. were able to identify new risk-dependent gene networks, with ∼3,000 differentially regulated genes implicated in the VSMC phenotypes observed with the risk haplotype and including more than one-third of known CAD risk genes. “This study … establishes haplotype-edited iPSCs as powerful tools for functionally annotating the human genome,” conclude the investigators.