Abstract
Purpose :
Chromosome 19q13.11 microdeletion syndrome is characterized by microcephaly, dysmorphic facies, growth retardation, limb and skin anomalies. Genetic studies have defined a minimal deletion interval of 324 kb, which includes the UBA2 (SUMO-activating enzyme subunit 2) gene. Here, we report a de novo truncation of UBA2 in a patient with hypertelorism, growth retardation, syndactyly and ectrodactyly. To explore the requirements of sumoylation in craniofacial, eye, and brain development, we developed a zebrafish loss-of-function model of uba2.
Methods :
Whole Exome Sequencing (WES) was used to identify the underlying genetic deficit in the proband. uba2 expression in zebrafish was appraised by whole mount in situ hybridization. CRISPR/Cas9 technique was used to generate zebrafish knockout line. Homozygous mutant (uba2-/-) larvae at 5-8 days post fertilization were evaluated for optical and skeletal abnormalities through gross morphology analysis, immunohistochemistry, alcian blue staining, and transmission electron microscopy (TEM). Wild type (WT) and mutation-harboring human mRNAs were injected at 1-2 cell stage embryos to determine their ability to rescue the phenotypical abnormalities in uba2-/-.
Results :
WES revealed a heterozygous nonsense variant [NM_005499.2: c.800T>A: p.(Leu267Ter)] in UBA2, encoding a highly conserved sumoylation pathway protein. In situ hybridization studies revealed strong expression of uba2 in developing eye, brain and limb buds of zebrafish. Loss of uba2 in zebrafish resulted in multiple gross morphological defects, including microphthalmia, mild to severe hydrocephaly, uninflated swim bladder, severe edema, and small-upright pectoral fins, leading to complete lethality by day 12. Furthermore, uba2-/- mutants have several neural retina and lens defects, including disrupted outer plexiform and photoreceptor layers, and absence of synaptic connections with ganglion cell layer. Imaging studies revealed disrupted craniofacial and pectoral fin development with scattered pattern of actinotrichia and abnormal epidermal cells. Microinjections of WT but not mutant human UBA2 mRNA significantly rescued gross phenotypes in uba2-/- mutants.
Conclusions :
Our clinical and genetics studies define a novel human disorder and zebrafish model of UBA2 loss-of-function. Our studies highlight the critical function of sumoylation pathway proteins in the development of eye and skeletal structures in humans and zebrafish.
This is a 2020 ARVO Annual Meeting abstract.