Contribution of Nonsense Mediated mRNA Decay to the Complex Genetics of Congenital Heart Disease

Premature termination codon (PTC)-inducing genetic variants represent a large fraction of clinically relevant pathogenic sequence variation. While often assumed to undergo nonsense-mediated mRNA decay (NMD) and to cause loss-of-function (LOF), PTC-containing transcripts may escape NMD (NMDesc) and exert dominant effects via dominant-negative or gain-of-function (DNGOF). However, NMD-aware genomic analysis of PTCs and their potential contribution to CHD has not been examined. Herein, we conducted a systematic analysis of predicted NMDesc PTCs for their latent role in congenital heart disease (CHD). Using whole exome sequencing data from ~3,000 CHD patients and ~5,000 controls, we analyzed predicted NMD triggering (NMDtrig) and NMDesc PTC identified by AENMD, software we developed for annotating genomic variants with predicted transcript-specific sensitivity to NMD. Case-control burden association testing of gene-sets from developmental pathways showed NMDesc PTCs disproportionately fell in genes mediating heart development. Further, we observed a burden of NMDtrig PTCs in previously associated gene-sets, such as NOTCH/WNT signaling and chromatin modifiers, while NMDesc PTCs were associated with cell adhesion and migration, hedgehog signaling, and MAPK signaling, the latter of which is associated with dominant variants causing Noonan syndrome. Gene-/transcript-level association analysis of NMDtrig/NMDesc PTCs identified 11 genes enriched in CHD cases, 5 associated with LOF, including FLT4, a gene known to cause Tetralogy of Fallot (TOF) via haploinsufficiency, and 6 genes with NMDesc with DNGOF prediction. Interrogating the protein-protein interactions of our 11 genes revealed an enrichment for known CHD genes, supporting the theory of an CHD-interactome network providing the context for CHD pathogenesis. Further, CHD patients showed enrichment for digenic pairs within our 11 genes and, PTCs in heart developmental processes recovered from CHD patients displayed a genetic network of not only di, and high order of within-patient PTC combinations. Altogether indicating the previously unrecognized contribution of oligogenic contributions to the etiology of CHD, which only revealed itself upon comprehensive analysis of both NMDtrig and NMDesc PTCs.