Alagille syndrome is an autosomal dominant disease with a known molecular etiology of dysfunctional Notch signaling caused primarily by pathogenic variants in JAGGED1 (JAG1), but also by variants in NOTCH2. The majority of JAG1 variants result in loss of function, however disease has also been attributed to lesser understood missense variants. Conversely, the majority of NOTCH2 variants are missense, though fewer of these variants have been described. In addition, there is a small group of patients with a clear clinical phenotype in the absence of a pathogenic variant. Here, we catalog our single‐center study, which includes 401 probands and 111 affected family members amassed over a 27‐year period, to provide updated mutation frequencies in JAG1 and NOTCH2 as well as functional validation of nine missense variants. Combining our cohort of 86 novel JAG1 and three novel NOTCH2 variants with previously published data (totaling 713 variants), we present the most comprehensive pathogenic variant overview for Alagille syndrome. Using this data set, we developed new guidance to help with the classification of JAG1 missense variants. Finally, we report clinically consistent cases for which a molecular etiology has not been identified and discuss the potential for next generation sequencing methodologies in novel variant discovery.

Biliary atresia (BA) is a rare pediatric cholangiopathy characterized by fibrosclerosing obliteration of the extrahepatic bile ducts, leading to cholestasis, fibrosis, cirrhosis, and eventual liver failure. The etiology of BA remains unknown, although environmental, inflammatory, infectious, and genetic risk factors have been proposed. We performed a genome-wide association study (GWAS) in a European-American cohort of 343 isolated BA patients and 1716 controls to identify genetic loci associated with BA. A second GWAS was performed in an independent European-American cohort of 156 patients with BA and other extrahepatic anomalies and 212 controls to confirm the identified candidate BA-associated SNPs. Meta-analysis revealed three genome-wide significant BA-associated SNPs on 2p16.1 (rs10865291, rs6761893, and rs727878; P < 5 ×10−8), located within the fifth intron of the EFEMP1 gene, which encodes a secreted extracellular protein implicated in extracellular matrix remodeling, cell proliferation, and organogenesis. RNA expression analysis showed an increase in EFEMP1 transcripts from human liver specimens isolated from patients with either BA or other cholestatic diseases when compared to normal control liver samples. Immunohistochemistry demonstrated that EFEMP1 is expressed in cholangiocytes and vascular smooth muscle cells in liver specimens from patients with BA and other cholestatic diseases, but it is absent from cholangiocytes in normal control liver samples. Efemp1 transcripts had higher expression in cholangiocytes and portal fibroblasts as compared with other cell types in normal rat liver. The identification of a novel BA-associated locus, and implication of EFEMP1 as a new BA candidate susceptibility gene, could provide new insights to understanding the mechanisms underlying this severe pediatric disorder.

Cornelia de Lange syndrome (CdLS) is a complex disorder with multiple structural and developmental defects caused by mutations in structural and regulatory proteins involved in the cohesin complex. NIPBL, a cohesin regulatory protein, has been identified as a critical protein responsible for the orchestration of transcriptomic regulatory networks necessary for embryonic development. Mutations in NIPBL are responsible for the majority of cases of CdLS. Through RNA-sequencing of human induced pluripotent stem cells and in vitro-derived cardiomyocytes, we identified hundreds of mRNAs, pseudogenes, and non-coding RNAs with altered expression in NIPBL+/− patient-derived cells. We demonstrate that NIPBL haploinsufficiency leads to upregulation of gene sets identified in functions related to nucleosome, chromatin assembly, RNA modification and downregulation of Wnt signaling, cholesterol biosynthesis and vesicular transport in iPSC and cardiomyocytes. Mutations in NIPBL result in the dysregulation of many genes responsible for normal heart development likely resulting in the variety of structural cardiac defects observed in the CdLS population.