In vivo identification and validation of novel potential predictors for human cardiovascular diseases

Abstract

Genetics crucially contributes to cardiovascular diseases (CVDs), the global leading cause of death. Since the majority of CVDs can be prevented by early intervention there is a high demand for the identification of predictive causative genes. While genome wide association studies (GWAS) correlate genes and CVDs after diagnosis and provide a valuable resource for such causative candidate genes, often preferentially those with previously known or suspected function are addressed further. To tackle the unaddressed blind spot of understudied genes, we particularly focused on the validation of human heart phenotype-associated GWAS candidates with little or no apparent connection to cardiac function. Building on the conservation of basic heart function and underlying genetics from fish to human we combined CRISPR/Cas9 genome editing of the orthologs of human GWAS candidates in isogenic medaka with automated high-throughput heart rate analysis. Our functional analyses of understudied human candidates uncovered a prominent fraction of heart rate associated genes from adult human patients impacting on the heart rate in embryonic medaka already in the injected generation. Following this pipeline, we identified 16 GWAS candidates with potential diagnostic and predictive power for human CVDs.

Natural genetic variation quantitatively regulates heart rate and dimension

“…Embryos were mounted in EMR into injection molds (1.5 % (w/v) agarose in ERM). Heart rate quantification of CRISPR-Cas9 and evoBE4max-mediated knockout embryos and control embryos were obtained using an ACQUIFER Imaging Machine and the HeartBeat software as described previously 15,45 . To acquire the heart rate, fluorescent image sequences have been captured for 10 sec with 24 fps at 21°C, 28°C, and 35°C.…”

Biochemical Structure and Function of TRAPP Complexes in the Cardiac System

“… 49 Medaka knockout model showed increase in heart rate and abnormal heart looping. 18 TRAPPC12 expression is decreased in acute myocardial infarction. 19 Variants have been associated with hydrocephalus, intellectual disability, microcephaly, and hearing loss.…”

“…Furthermore, TRAPPC12 crispants resulted in morphological heart phenotypes including heart looping defects that caused improper atrium placement compared with the ventricle. 18 TRAPPC12 has also been linked to coronary artery disease, with acute myocardial infarction patients having markedly decreased TRAPPC12 gene expression compared with patients with normal coronary artery function and intermediate coronary lesions. 19 …”

“…TRAPPIII, in conjunction with other proteins, act as a membrane trafficking factor to recruit COP II to the ER by promoting outer layer assembly of the COPII coat to conduct its normal processes. 18 COPI mediates a recycling pathway that retrieves proteins from both the Golgi and ER-Golgi intermediate compartment and returns them to the ER. TRAPPI, in conjunction with TRAPPII, helps this intra-Golgi trafficking process function by facilitating COPI.…”

“… 12 , 13 However, TRAPPC subunit variants and/or changes in expression have also been recently associated with structural cardiac diseases such as small-vessel stroke, cardiomyocyte apoptosis, heart looping/atrium placement, and coronary artery disease, and electrical cardiac dysfunction such as atrial fibrillation and Ca 2+ handling defects. 4 , 14 , 15 , 16 , 17 , 18 , 19 , 20 …”