Date of Graduation
8-2010
Document Type
Dissertation (PhD)
Program Affiliation
Genes and Development
Degree Name
Doctor of Philosophy (PhD)
Advisor/Committee Chair
William Mattox, Ph.D.
Committee Member
Richard Behringer, Ph.D.
Committee Member
Andreas Bergmann, Ph.D.
Committee Member
Gilbert Cote, Ph.D.
Committee Member
Pierre McCrea, Ph.D.
Abstract
DMRT (Doublesex and Mab-3 related transcription factor) proteins generally associated with sexual differentiation in many organisms share a common DNA binding domain and are often expressed in reproductive tissues. Aside from doublesex, which is a central factor in the regulation of sex determination, Drosophila possesses three different dmrt genes that are of unknown function. Because the association with sexual differentiation and reproduction is not universal and some DMRT proteins have been found to play other developmental roles we chose to further characterize one of these Drosophila genes. We carried out genetic analysis of dmrt93B, which was previously found to be expressed sex-specifically in the developing somatic gonad and to affect testis morphogenesis in RNAi knockdowns. In order to disrupt this gene, the GAL4 yeast transcriptional activator followed by a polyadenylation signal was inserted after the dmrt93B start codon and introduced into the genome by homologous recombination. Analysis of the knock-in mutation as well as a small deletion removing all dmrt93B sequence demonstrate that loss of function causes partial lethality at the late pupal stage. Surprisingly, these mutations have no significant effect on gonad formation or male fertility. Analysis of GAL4-driven GFP reporter expression indicates that the dmrt93B promoter activity is highly specific to neurons in the suboesophageal and proventricular ganglion in larva and adult of both sexes suggesting a possible role in digestive tract function. Using the Capillary Feeder (CAFÉ) assay to measure daily food intake we find that reduction in this gene’s function leads to an increase in food consumption. These results suggest dmrt93 plays an important role in the formation or maintenance of neurons that affect feeding and support the idea that dmrt genes may not be restricted to roles in sexual differentiation.
Keywords
dmrt93B, Doublesex and Mab-3 related transcription factor, Drosophila, GAL4 knock-in, Capillary Feeder (CAFÉ) assay, ends-out gene replacement, homologous recombination in vivo