A zebrafish model of nicotinamide adenine dinucleotide (NAD+) deficiency-derived congenital disorders

Congenital NAD deficiency disorder (CNDD) is a multisystem condition in which cardiac, renal, vertebral, and limb anomalies are most common, but anomalies in all organ systems have been identified. Patients with this condition have biallelic pathogenic variants involving genes in the nicotinamide adenine dinucleotide (NAD+) synthesis pathway leading to decreased systemic NAD+ levels. CNDD anomalies mimic the clinical features described in vertebral-anal-cardiac-tracheoesophageal fistula-renal-limb (VACTERL) association raising the possibility that CNDD and VACTERL association possess similar underlying causes. However, the mechanism by which NAD+ deficiency causes CNDD developmental anomalies has not been determined, nor has NAD+ deficiency been definitively linked to VACTERL association. Therefore, additional animal models amenable to detailed observation of embryonic development are needed to address the causes and progression of congenital anomalies in both CNDD and VACTERL association. Here, we describe a zebrafish model of NAD+ disruption to begin to model CNDD and VACTERL association phenotypes, assessing developmental anomalies in real-time. Treatment of zebrafish embryos with 2-amino-1,3,4-thiadiazole (ATDA), a teratogen known to disrupt NAD+ metabolism, resulted in neural tube, craniofacial, cardiac, and tail defects. These defects were rescued by the administration of nicotinamide (NAM) in a dose-dependent manner. Our work establishes zebrafish as a useful model for investigating the mechanistic causes and developmental dynamics of CNDD and VACTERL association. Further, as VACTERL association has been linked to teratogens, our zebrafish model provides a platform to assess these agents. One sentence summaryNAD(H) deficiency causes multiple congenital anomalies in zebrafish.