Molecular Biology and Genetics Abstracts

Analyzing heme and pathways in Drosophila melanogaster

Pendleton Cox, 499

How heme biosynthesis is regulated is largely unclear. To study heme biosynthetic pathways, genes in the production pathway will be knocked down via RNAi with different Gal4 drivers in different tissues. As well, heme will be measured with a stain in wild type and mutant flies to further understand the level of heme required at the third instar larval stage. Finally, RNAi knockouts of genes used for iron synthesis will be done to view the effects on the fly and its relation to heme.


Defects in ciliary development in UNC119b knockdown zebrafish

Francesca Jean, M.Sc.

Many aspects of animal development and function require cilia, including organizing the embryonic axis, sensing the environment, and ensuring proper pathfinding of neurons. Though the structure of cilia is well established, many of the factors involved in cilia development are poorly understood. Our lab identified unc-119, a gene that has high sequence conservation across a wide range of organisms. It has been hypothesized that the functionally conserved protein product, UNC-119, may have a role in cilia development in both vertebrates and invertebrates. MO (morpholino oligonucleotide)-injected zebrafish display a small eye phenotype, along with exencephaly (a condition where the brain develops outside of the cranial cavity) and heart edema (swelling of the heart) while previous work has revealed axon pathfinding errors and misplaced neuronal cell bodies. The goal of my project is to identify how these phenotypes may be associated with defects in ciliary development. Using immunohistochemistry, I have found that incomplete development of the photoreceptors, which are a type of sensory cilia, occurs in MO zebrafish whereas development of the photoreceptor layer looks complete in equivalently aged WT zebrafish. In addition, when using in situ hybridization, MO zebrafish demonstrate a coloboma during early eye development, which is failure of the optic fissure to close. One possibility for the small eye phenotype is that UNC119b (one of the UNC-119 homologues in zebrafish) is involved in photoreceptor development; a lack of the gene likely results in truncation of these photoreceptors. These preliminary results suggests that knockdown of unc119b has a negative effect on cilia, particularly photoreceptor, development. However, analyzing MO zebrafish is often riddled with limitations, therefore, work is also in progress to create a stable mutation in UNC119b in zebrafish using TALENs (Transcription Activator-Like Effector Nuclease). By analyzing a stable mutation in UNC119b in zebrafish, we can verify the phenotype identified using MOs and further analyze both sensory and motile cilia during development.