Affiliations: 1) RNA BioScience Initiative, University of Colorado School of Medicine, Aurora, CO ; 2) Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO; 3) Metropolitan State University of Denver, Denver, CO
Keywords: r. other (Maternal-to-Zygotic transition); b. oogenesis
The maternal-to-zygotic transition (MZT) is an essential developmental process whereby maternal gene products deposited into the oocyte are degraded, and zygotic gene products take their place. The degradation of maternal RNA has been well studied and is known to be an important part of the MZT. However, the removal of maternally deposited proteins is far less studied. The main mechanism for protein degradation pathway works through the ubiquitin-proteosome system, which uses E1s, E2s, and E3s and culminates in the ubiquitylation of target proteins, thus marking them for degradation. The Drosophila melanogaster gene encodes dozens of E2s and hundreds of E3s, but we know very little about their involvement in the MZT. As part of a multi-year undergraduate research program, we are performing an RNAi screen to identify E2s, E3s, and related proteins that are essential for oogenesis and embryogenesis. Here, we will describe the results of ten randomly selected E2 and E3 genes: CG11658, Cul-3, Cul-5, EloB, faf, slmb, l(3)73Ah, CSN6, Nedd4, and Prosalpha5. We will present results on the effectiveness of RNAi knockdown by RT-qPCR and western blotting. We will describe the extent to which each gene affects: egg laying; embryonic viability; other developmental effects. In the future, we will be able to use unbiased techniques, like mass spectrometry, to identify putative targets of maternally required E2s and E3s. Together, this screen will help reveal the factors required for proper protein degradation in the MZT.