Project description:Next-generation sequencing (NGS) has supported precision therapeutic approaches that have improved the lives of children with rare diseases. Patients with congenital diarrhea and enteropathies (CODE) have a particularly difficult disease with high morbidity and mortality. However, there are now several targeted therapies including specific diets, pharmacological treatments, and surgical interventions that are based on an individual’s genetic diagnosis. We performed NGS on a large cohort of 139 infants with suspected monogenic congenital diarrheal disorders and identified known causal variants in 50% of cases, including a new founder NEUROG3 variant. We also uncovered and functionally characterized three novel CODE genes, GRWD1, MYO1A, and MON1A, using cell and zebrafish models.
Project description:Congenital Heart Disease (CHD) accounts for 1% of birth defects, and while large-scale genetic studies have uncovered genes associated with CHDs, identifying causal mutations remains a challenge. We hypothesized that genetic determinants for CHDs could be found in the protein interactomes of GATA4 and TBX5, two cardiac transcription factors (TFs) associated with CHDs. Defining their interactomes in human cardiac progenitors via affinity purification-mass spectrometry and integrating the results with genetic data from the Pediatric Cardiac Genomic Consortium (PCGC) revealed an enrichment of de novo variants among proteins that interact with GATA4 or TBX5. A consolidative score designed to prioritize TF interactome members based on distinctive variant, gene and proband features identified numerous 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 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 CHD and can be extended to other genetic diseases.
Project description:The glaucomas are a group of diseases characterized by optic nerve damage that together represent a leading cause of blindness in the human population and in domestic animals. Here we report a mutation in LTBP2 that causes primary congenital glaucoma (PCG) in domestic cats. We identified a spontaneous form of PCG in cats and established a breeding colony segregating for PCG consistent with fully penetrant, autosomal recessive inheritance of the trait. Elevated intraocular pressure, globe enlargement and elongated ciliary processes were consistently observed in all affected cats by 8 weeks of age. Varying degrees of optic nerve damage resulted by 6 months of age. Although subtle lens zonular instability was a common feature in this cohort, pronounced ectopia lentis was identified in less than 10% of cats examined. Thus, glaucoma in this pedigree is attributed to histologically confirmed arrest in the early post-natal development of the aqueous humor outflow pathways in the anterior segment of the eyes of affected animals. Using a candidate gene approach, significant linage was established on cat chromosome B3 (LOD 18.38, q = 0.00) using tightly linked short tandem repeat (STR) loci to the candidate gene, LTBP2. A 4 base-pair insertion was identified in exon 8 of LTBP2 in affected individuals that generates a frame shift that completely alters the downstream open reading frame and eliminates functional domains. Thus, we describe the first spontaneous and highly penetrant non-rodent model of PCG, identifying a valuable animal model for primary glaucoma that closely resembles the human disease providing valuable insights into mechanisms underlying the disease and a valuable animal model for testing therapies.
Project description:Hypoplastic left heart syndrome (HLHS) is a heterogeneous, lethal combination of congenital malformations that result in a heart unable to sustain systemic circulation. The genetic determinants of this disorder are largely unknown. Evidence of copy number variants (CNVs) contributing to the genetic etiology of HLHS and other congenital heart defects (CHDs) has been mounting. However, the functional effects of such CNVs have not been examined, particularly in cases where the variant of interest is found in only a single patient. Initially whole-genome SNP microarrays were employed to detect CNVs in two patient cohorts (N = 70 total) predominantly diagnosed with some form of nonsyndromic HLHS. We discovered 16 rare variants adjacent to or overlapping 20 genes associated with cardiovascular or premature lethality phenotypes in mouse knockout models. Fifteen of the 16 variants were identified in separate patients, suggestive of a private mutation model of disease. We evaluated the impact of selected variants on the expression of nine of these genes through quantitative PCR on cDNA derived from patient heart tissue. Four genes displayed significantly altered expression in patients with an overlapping or proximal CNV verses patients without such CNVs. Thus, rare and private genomic imbalances perturb transcription of genes affecting cardiogenesis in a subset of nonsyndromic HLHS patients. Some of these genes influence extracellular matrix structure, cardiac neural crest development, and coronary vascularization. A total of 70 samples from CHD patients, mostly with nonsyndromic HLHS, yielded SNP genotypes and probe intensity ratios of sufficient quality for CNV identification. Two classes of CNV detection algorithms (HMM and CBS) were employed. After identification, concordant entries were subjected to selection criteria based on rarity and gene content, which produced putative candidate genes for follow-up experiments.
Project description:MicroRNAs negatively regulate gene expression and may serve as biomarkers for human cardiomyopathy. In the domestic cat, hypertrophic cardiomyopathy (HCM) represents the most common primary cardiomyopathy. In humans, the etiology of HCM is linked to mutations in genes of contractile muscle proteins, while in cats a clear proof for causal mutations is missing. The etiology of feline HCM is uncertain. Diagnosis is made by heart ultrasound examination and measuring the serum level of N-terminal pro B-type natriuretic peptide. The purpose of this study was to investigate whether microRNA profiles in the serum of cats with HCM are different from the profiles of healthy cats and whether specific miRNAs can be detected to serve as potential biomarkers for feline HCM or may help in understanding the etiology of this disease
Project description:Pathogenic variants in CHRNE encoding the epsilon subunit of acetyl choline receptor (AChR) result in impaired neuromuscular transmission and congenital myasthenic syndromes (CMS). Clinical manifestations include facial, ocular and limb fatigability and weakness, whereby severity of symptoms may vary in patients harboring the same pathogenic variant. Although the underlying pathophysiology is well-known, blood biomarkers enabling a patient-stratification are lacking. Previous studies revealed CHRNE protein in white blood cells (WBC) rendering these cells suitable for the study of cellular and minimal-invasive marker proteins.
Project description:MicroRNAs negatively regulate gene expression and may serve as biomarkers for human cardiomyopathy. In the domestic cat, hypertrophic cardiomyopathy (HCM) represents the most common primary cardiomyopathy. In humans, the etiology of HCM is linked to mutations in genes of contractile muscle proteins, while in cats a clear proof for causal mutations is missing. The etiology of feline HCM is uncertain. Diagnosis is made by heart ultrasound examination and measuring the serum level of N-terminal pro B-type natriuretic peptide. The purpose of this study was to investigate whether microRNA profiles in the serum of cats with HCM are different from the profiles of healthy cats and whether specific miRNAs can be detected to serve as potential biomarkers for feline HCM or may help in understanding the etiology of this disease Blood was drawn from two groups of cats: 12 healthy cats and 11 cats suffering from hypertrophic cardiomyopathy. After clotting, samples were centrifuged and total mRNA was extracted from serum. These 23 serum samples were analyzed and the groups were compared
Project description:The relationship between maternal cholesterol deficiency and the risk of congenital heart defects (CHD) in offspring is not fully understood. In a birth cohort study of 5,041 family trios, we found that low maternal cholesterol levels were significantly associated with an increased risk of CHD, with relative risk (RR) of 1.52 in the second trimester and 1.73 in the third trimester. To further investigate this link, we treated pregnant mice with cholesterol-lowering agents, namely ezetimibe or atorvastatin. Both treatments led to a significant increase in the incidence of CHD in their offspring. To identify a pathogenic variant that could provide genetic evidence linking cholesterol synthesis to CHD occurrence and serve as a target for constructing genetic mouse models, we performed whole-genome sequencing (WGS) on 103 CHD cases from the birth cohort. We identified a recurrent functional variant in the CYP51A1 gene (c.1147A>G, p.Ile383Val). We then developed a CYP51A1I383V knock-in mouse model. This variant disrupted cholesterol synthesis, resulting in CHD through impaired hedgehog (Hh) signaling. Most intriguingly, maternal dietary intervention to increase cholesterol intake effectively reduced the risk of CHD in CYP51A1I383V mutant offspring. Our study suggests that low maternal cholesterol during pregnancy increases the risk of CHD in offspring by inhibiting Hh signaling, and that maternal cholesterol supplementation during pregnancy may reduce the occurrence of CHD.