Integration of Protein Interactome Networks with Congenital Heart Disease Variants Reveals Candidate Disease Genes

Abstract

SUMMARYCongenital heart disease (CHD) is present in 1% of live births, yet identification of causal mutations remains a challenge despite large-scale genomic sequencing efforts. We hypothesized that genetic determinants for CHDs may lie in protein interactomes of GATA4 and TBX5, two transcription factors that cause CHDs. Defining their interactomes in human cardiac progenitors via affinity purification-mass spectrometry and integrating results with genetic data from the Pediatric Cardiac Genomic Consortium revealed an enrichment of de novo variants among proteins that interact with GATA4 or TBX5. A consolidative score that prioritized interactome members based on variant, gene, and proband features identified likely CHD-causing genes, including the epigenetic reader GLYR1. GLYR1 and GATA4 widely co-occupied cardiac developmental genes, resulting in co-activation, and the GLYR1 missense variant associated with CHD disrupted interaction with GATA4. This integrative proteomic and genetic approach provides a framework for prioritizing and interrogating the contribution of genetic variants in disease.

Transcription Factor GATA4 Regulates Cell Type–Specific Splicing Through Direct Interaction With RNA in Human Induced Pluripotent Stem Cell–Derived Cardiac Progenitors

“…More than 250 proteins were identified as potential GATA4 interacting partners, including several previously reported GATA4 interactors and novel interactors. 54 Surprisingly, there were many significant interactions between GATA4 and RNA-binding proteins belonging to the spliceosome complex (eg, SNRPF [small nuclear ribonucleoprotein polypeptide F], SF3A1 [splicing factor 3a subunit 1], SF3A3 [splicing factor 3a subunit 3]), RNA splicing regulators (eg, HNRNPF [heterogeneous nuclear ribonucleoprotein F], HNRNPR [heterogeneous nuclear ribonucleoprotein R]), and mRNA processing regulators (eg, DHX15 [DEAH-box helicase 15], SRRT [serrate RNA effector molecule homolog]; Figure 1B). Coimmunoprecipitation analysis of the GATA4 interaction with RNA-binding proteins, for which there are high quality antibodies, confirmed that interaction with SF3A3, HNRNPH1 (heterogeneous nuclear ribonucleoprotein H1), and SF3A120 (splicing factor 3 subunit 1) can indeed occur, even in the presence of RNAse (Figure 1C; Figure S1A).…”

“…As most transcription factors function in complexes to regulate gene expression, our recent work investigated GATA4's protein interactome by using an antibody to the endogenous GATA4 for affinity purification followed by mass spectrometry in human iPS-CPs treated with RNase and DNase in triplicates (Figure 1A). 54 Human GATA4-null iPS-CPs were used as the negative control for nonspecific interactions. More than 250 proteins were identified as potential GATA4 interacting partners, including several previously reported GATA4 interactors and novel interactors.…”