Nikale Pettie and Matt Cring to present at Genetics Student Seminar 5/12/2016
The Conserved and Unique Determinants of Crossover Events
Meiotic recombination promotes genetic diversity and allows for the continuous adaptation of natural populations to ever changing environments. Mechanistically, recombination relies heavily on protein-protein and protein-DNA interactions. The goal of my project is to find out how genetic incompatibilities through failed interactions disrupt the tight control of recombination localization across the genome in interspecific hybrids. To this end, I am currently generating high-resolution recombination maps in two closely related species of fruit flies, Drosophila yakuba and D. santomea, and their hybrids. Our findings will have a positive impact in our understanding of the role of protein interactions in recombination control, will illuminate the pervasiveness of sexual reproduction and recombination in nature, and will enable the development of new theoretical models on the maintenance of genetic variation.
Genetic therapeutic strategies for the Bardet-Biedl Syndrome (BBS) M390R mutation
Bardet-Beidl Syndrome (BBS) is a pleiotropic ciliopathy that causes a variety of phenotypes in humans and animal models, including blindness and male infertility. BBS is considered a model disease for ciliopathies in general, which affect approximately 1 in 1000 people worldwide. There are no current efficient non-surgical treatments for ciliopathies, illustrating a critical need for new therapeutic strategies. The most common cause of BBS is the M390R mutation in Bardet-Biedl Syndrome 1 (BBS1) gene, an essential component of the BBsome that is required for basal body function and primary ciliogenesis. This mutation leads to photoreceptor degeneration and dysfunctional flagella in sperm. My objective is to use gene replacement and editing techniques to restore male fertility and prevent retinal degeneration in a Bbs1M390R/M390R mouse model. I hypothesize that these phenotypes are amenable to correction by gene therapy and gene correction. Here, I will test this hypothesis by using BBS1 gene therapy and correction as a strategy to preserve photoreceptors in a mouse model. First, I will elucidate the mechanism by which BBS1 overexpression in the retina causes toxicity. I hypothesize that overexpression of BBS1 leads to homodimerization and subsequent inability to incorporate into the BBsome, thereby affecting ciliary function. I will deliver adeno-associated viruses (AAVs) expressing BBS1 driven by the BBS1 promoter to the Bbs1M390R/M390R mice postnatally, and subsequently test photoreceptor preservation by electroretinogram (ERG) and immunohistology. I will also use CRISPR/Cas9 mediated homologous recombination to correct the M390R mutation in vitro and in vivo. In addition to Cas9, I will employ Cpf1, a Cas9-like class 2 CRISPR endonuclease, to simultaneously knock out dysfunctional BBS1 and introduce functional BBS1 cDNA driven by the BBS1 promoter. By correcting the germline in vivo, all downstream progeny should effectively be corrected. While most studies using gene editing techniques are completed ex vivo, my approach will show the level of correction that can occur in vivo using various novel methods. These techniques may be applied to other degenerative genetic diseases.