Tag: Chris Doe

Utilizing Drosophila to investigate novel regulatory pathways for the Hox gene Sex Combs Reduced

Presenter: Alanna Sowles – Neuroscience

Faculty Mentor(s): Kristen Lee, Chris Doe

Session: (In-Person) Poster Presentation

Across species, Hox genes are necessary for an organism’s anatomical development, including the nervous system. Interestingly, these proteins continue functioning within neurons of mature organisms. This research seeks to determine the functional purpose of Hox genes post-development, as these mechanisms could provide novel etiological insight into neurodevelopmental disorders. Drosophila melanogaster is an effective tool for this investigation because fly neurons are similar to mammals, and gene expression of individual neurons is easily manipulated. Within this model, I will utilize the well-characterized Pair1 pre-motor neuron, which expresses the Hox gene Sex combs reduced (Scr). I hypothesize that Scr is functioning in a conserved molecular pathway to preserve the morphology and function of Pair1 neurons. Past research provides intuitive candidates for exploring these mechanisms, like Pair1 proteins Hb and Bcd. RNAi-facilitated knockdown had no significant impact on Scr expression, prompting exploration of alternative genes. Using published resources, several genes with expression patterns similar to Scr were selected and visualized with GFP-tagged proteins. Colocalization of these genes with Scr was assessed via immunohistochemistry, revealing 8 promising candidates for further analysis. Scr expression will be measured after knockdown experiments are repeated for each gene. The results will hopefully illuminate novel regulatory pathways of Scr beyond development.

Decreasing Hunchback and Bicoid Levels in Pair1 Neurons Alters the Pair1 Circuit in Drosophila

Presenter: Amanda Linskens – Biochemistry

Faculty Mentor(s): Kristen Lee, Chris Doe

Session: (In-Person) Poster Presentation

Transcription factors (TFs) are essential for cell specification across multiple species, including humans. During Drosophila melanogaster development, neuroblasts produce neuronal progeny that acquires identity based on the temporal TF (tTF) present during birth. tTFs activate specific Homeodomain TFs (HDTFs), which are also important for determining neuronal identity. Thus, the cascade of tTFs in neuroblasts creates the diversity necessary for forming precise neural circuits. Although prior research shows that TFs generate variety, few studies have examined how these TFs influence circuit establishment. My study focuses on the Pair1 neurons, which initiate pausing in larvae through neural circuits. Prior research in our laboratory showed that the Pair1s derive from the tTF Hunchback (Hb), which activates the HDTF Bicoid (Bcd). Therefore, I hypothesized that manipulating Hb’s and Bcd’s expressions in Pair1 would alter the Pair1 circuit. To investigate this,

I expressed the green fluorescent protein (GFP) in the Pair1s and knocked down Hb and Bcd levels individually to assay circuit morphology and behavior. I found that decreasing Hb resulted in more Pair1 axonal connections, affecting behavior. Interestingly, I saw similar results when Bcd was knocked down in Pair1, but the phenotypes are weaker than those experienced with decreased Hb levels. These results suggest that tTF activation of HDTFs is vital for circuit establishment in the central nervous system.

Enhancer-GAL4 Database Yields Subsets of Gene Expression

Presenter: David Bauer, Kris Haenderson, Allison Wilbur, Biology

Poster: A-2

Mentor: Chris Doe, Biology

Enhancers and suppressors are major controls in the tempro-spatial expression of a gene. Prior work has shown that enhancers may be made up of small modules, each driving only a small subset of expression in an overall pattern that we associate with a gene. The Rubin lab at Janelia Farms Research Center created over 70,000 fragment sequences from more than 1,100 neural gene regions. These fragment enhancer sequences were used to generate site-specific transgenic GAL4 expressing Drosophila lines. These lines are currently being screened by several collaborating labs for their ability to function as transcriptional enhancers to drive limited subsets of UAS-GFP expression. Our lab’s role is to screen Drosophila embryos for neural expression patterns of the transgenic enhancer fragment lines. As a test analysis, we took a subset of 82 individual fragment enhancer lines derived from seven gene regions in embryonic nerve cords (VNC). These 82 lines produced 77% of fragments with expression in the VNC. Within that expressing set, 62% drove small patterns of less than ten cells per hemisegment. This data complements the screening results collaborative labs are finding in Drosophila larval brains. Together, these results show that the theory of enhancers being a set of modules acting in concert to yield an overall pattern of gene expression seems quite likely. The end result should be a valuable tool that can be used for studies of gene control, patterning, and neuronal circuitry.