Date of Award
Master of Science (MS)
Environmental Science and Biology
The Notch gene encodes a transmembrane protein that functions as a receptor for intracellular signals in Drosophila melanogaster. The Notch gene is required for cell proliferation as well as differentiation. A Notch mutation, Nnd-p, is caused by the insertion of a P element into the first exon upstream of the start of translation of the Notch gene. The insertion causes a weak Notch mutation. This Notch mutation is lethal in the presence of an enhancer of rudimentary mutation. This lethality of the X chromosome that contains Nnd-p and e(r)27-1 was hypothesized to be caused by an interaction between these two mutations. In order to support this hypothesis, two aims were done. The first aim was to revert Nnd-p and then examine the viability in the presence of e(r)27-1. The second aim was to revert the lethality of the Nnd-p e(r)27-1 chromosome and then examine the Nnd-p mutation. It was determined that the Nnd-p mutation is the cause of the lethality. In the first case where Nnd-p was reverted, the lethality was also reverted. In the second case where the lethality was reverted, Nnd-p was concurrently reverted. The two Nnd-p revertants, R(Nnd-p)4 and R(Nnd-p)9, were determined to be imprecise excisions of the P element where the P element left some of its DNA behind. New Notch mutations caused by the mobilization of the P element Nnd-p were also isolated. Each of them (N2, N3, and N5) resulted in a further decrease in Notch activity. Each of the mutations were analyzed by PCR and DNA sequencing in order to determine the precise structure of the mutation. The mutant phenotypes were studied by examining viability of both the larvae and the adults. By examining the viability, the point in the Drosophila life cycle where the lethal interaction occurs can be determined. Crosses were set up between males with the X chromosome tagged with GFP and female containing a Notch mutation on one of the X chromosomes and GFP on the other. The males and females were scored based on their visible phenotypes. The larvae were observed through the 1st, 2nd, and 3rd Instar stages. The N2 and N3 mutants were found to be embryonic lethal. Observing the viability of the larvae, shows that the e(r) gene can affect N expression in the embryo stage. The Notch mutant larvae did not give rise to adult flies, except for one N3 male that survived to adulthood. This revertant of N3 was found to have a wild type Notch gene probably due to recombination with the GFP-containing X chromosome. The lethal interaction between Nnd-p e(r)27-1 was previously shown to be during the pupal stage, however, N5 e(r)27-1 was found to be lethal during embryogenesis. These results indicate that e(r) is necessary for Notch activity both during embryogenesis and metamorphosis.
Kostrzebski, Melissa A., "Structure and Phenotypic Analysis of Notch Mutations" (2010). Environmental Science and Ecology Theses. 47.