Identification and characterization ofbemused, a Drosophila gene involved in neuronal excitability, gametogenesis and embryonic development
Walters, Karina Joanne
Doctor of Philosophy
During a screen for novel genes involved in the process of neurotransmission, a gene, bemused, was identified as the result of a transposon insertion at 85D on the third chromosome. Mutation of bem produces flies that are extremely uncoordinated and unable to fly. Subsequent electrophysiological recordings performed at the neuromuscular junction revealed neuronal hyperexcitability of bem mutants. Mutation of bemused also results in female sterility and reduced male fertility, both of which were determined to be the result of defects in oogenesis and spermatogenesis, respectively. Oogenesis defects include actin membrane degeneration within the egg chamber, resulting in nurse cell fusions and mispositioned nurse cell nuclei, abnormal dorsal appendage formation and in severe cases, a misplaced or absent oocyte. These phenotypes are often observed in mutants disrupting the cytoskeleton, suggesting bem may have a cytoskeletal function. Analysis of spermatogenesis revealed the lack of production and storage of mature sperm in the seminal vesicles of bem testes. Homozygous bem females deposit a small number of superficially normal eggs that fail to hatch, thus defects in embryogenesis were hypothesized. Indeed, the embryos of bem mothers have a number of defects in early embryogenesis, including the formation of polyploid nuclei presumably as a consequence of aberrant mitotic spindles, incomplete axial migration of the nuclei to the anterior pole, disruption of proper formation of the pole cells and abnormal syncytial blastoderm characteristics, such as nuclear withdrawal from the periphery and nuclear aggregates. These abnormalities resemble defects observed in embryos treated with cytoskeleton-disrupting drugs, further supporting a role for bemused in the structure or regulation of the cytoskeleton. Approximately 15 kb of genomic DNA from the bem region was isolated but identified only questionable partial transcription units when used to screen cDNA libraries. Subsequent sequencing of the 15 kb of genomic DNA surrounding the bem P element insertion failed to identify any transcribed regions; therefore, long term cloning strategies have been initiated to identify the bem transcription unit, including the isolation of additional genomic DNA and the creation of additional bem alleles.
Molecular biology; Neurosciences; Entomology