The vertebrate eye development begins with the symmetric, bilateral evagination of the diencephalon to form optic vesicle (OV). As development continues the OV invaginates to form bilayer optic cup, the invagination of OV is asymmetric and creates a gap in the ventral region of the developing eye, known as the optic fissure. To continue normal development, the two edges of this fissure must fuse. Any abnormality in the optic fissure closure results in a potentially blinding congenital ocular malformation known as uveal coloboma, account for 10% of childhood blindness. Several modes of inheritance have been documented, yet, most cases are sporadic, impeding effective genetic counseling for families affected by the disease. Although the embryology has long been understood, the genetic and developmental mechanisms behind optic fissure closure defects are still largely unknown.
CRISPR/Cas9 mediated genome engineering technology made it possible to perform targeted mutagenesis in a fast and economical way in zebrafish that facilitate large-scale phenotyping efforts in mutant embryos. We have used CRISPRs to generate knock out (KO) lines for previously described nlz1 and nlz2 gene that are necessary for normal optic fissure closure (1) and nlz1 and nlz2 CRISPRs KO embryos exhibited ocular coloboma (2). Genes that are dynamically regulated at the edges of the optic fissure during the process of closure are likely to be important for proper execution of this developmental process. Our lab identified 164 annotated genes dynamically regulated during optic fissure closure in the mouse using laser capture microdissection (LCM) and microarray (1), and the function of 90% of the genes during eye development are unknown. To understand the function of the genes expressed during optic fissure closure, we are generating CRISPRs KO zebrafish lines and screen for ocular coloboma, and other eye abnormalities in the KO embryos. In future our research will shed light into genes and genetic network that are crucial for optic fissure closure and eye abnormalities as well as to move forward in the genetic research of uveal coloboma.
1. Brown JD et al. (2009) Expression profiling during ocular development identifies 2 Nlz genes with a critical role in optic fissure closure. PNAS 106(5):1462-7.
2. Dutta S et al. (2015) nlz1 is required for cilia formation in zebrafish embryogenesis.Dev Biol. 2015 Oct 15; 406(2):203-11.