How axons in the developing nervous system successfully navigate to their correct targets is a fundamental problem in neurobiology. Axons are guided by both attractive and repulsive cues, which are members of evolutionarily conserved protein families. We are interested in the signaling mechanisms that function during attractive and repulsive axon guidance. The midline of the Drosophila embryonic CNS provides an ideal system to address these questions. Like its structural analog, the vertebrate floor plate, the fly midline is an intermediate target for many classes of navigating axons, which must decide whether or not to cross the midline. In the Drosophila CNS, the conserved guidance cue, Slit, functions to prevent axons from abnormally crossing the midline. Slit repulsion is mediated by the conserved family of Roundabout (Robo) receptors. The major aims of this application are: 1) to delimit and characterize the regions of the Robo receptor's cytoplasmic domain that are necessary and sufficient for Robo-mediated axon repulsion in response to the Slit ligand, 2) to assess the potential role of the SH3-SH2 adaptor protein Dreadlocks (Dock) and associated proteins in contributing to Robo repulsion, 3) to identify additional components involved in Slit and Robo signaling using a Drosophila genetic screen. A well established Drosophila transgenic approach will be used to determine which regions of Robo's cytoplasmic domain are required for repulsion. Classical genetic and biochemical techniques, including genetic interaction tests, mutant analysis, yeast two hybrid and co-immunoprecipitation will be used to investigate the potential role of Dock and associated proteins in Robo repulsion. In addition, established cell culture techniques will be used to address whether Slit stimulation of the Robo receptor regulates Dock/Robo interactions. To identify additional molecules involved in Robo function, a sensitized genetic screen will be performed. The proposed genetic screening strategy has already been successfully used on a small scale to identify genes that may play important roles during Robo repulsion. Deciphering the mechanisms that mediate Robo repulsive axon guidance will give important insight into how the nervous system is correctly wired during development and may have implications for nerve regeneration.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS046333-05
Application #
7262460
Study Section
Molecular, Cellular and Developmental Neurosciences 2 (MDCN)
Program Officer
Mamounas, Laura
Project Start
2003-08-01
Project End
2008-07-31
Budget Start
2007-08-01
Budget End
2008-07-31
Support Year
5
Fiscal Year
2007
Total Cost
$351,385
Indirect Cost
Name
University of Pennsylvania
Department
Neurosciences
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Santiago, Celine; Bashaw, Greg J (2017) Islet Coordinately Regulates Motor Axon Guidance and Dendrite Targeting through the Frazzled/DCC Receptor. Cell Rep 18:1646-1659
Hernandez-Fleming, Melissa; Rohrbach, Ethan W; Bashaw, Greg J (2017) Sema-1a Reverse Signaling Promotes Midline Crossing in Response to Secreted Semaphorins. Cell Rep 18:174-184
Neuhaus-Follini, Alexandra; Bashaw, Greg J (2015) Crossing the embryonic midline: molecular mechanisms regulating axon responsiveness at an intermediate target. Wiley Interdiscip Rev Dev Biol 4:377-89
Evans, Timothy A; Santiago, Celine; Arbeille, Elise et al. (2015) Robo2 acts in trans to inhibit Slit-Robo1 repulsion in pre-crossing commissural axons. Elife 4:e08407
Chance, Rebecca K; Bashaw, Greg J (2015) Slit-Dependent Endocytic Trafficking of the Robo Receptor Is Required for Son of Sevenless Recruitment and Midline Axon Repulsion. PLoS Genet 11:e1005402
Neuhaus-Follini, Alexandra; Bashaw, Greg J (2015) The Intracellular Domain of the Frazzled/DCC Receptor Is a Transcription Factor Required for Commissural Axon Guidance. Neuron 87:751-63
Santiago, Celine; Bashaw, Greg J (2014) Transcription factors and effectors that regulate neuronal morphology. Development 141:4667-80
Zarin, Aref Arzan; Asadzadeh, Jamshid; Hokamp, Karsten et al. (2014) A transcription factor network coordinates attraction, repulsion, and adhesion combinatorially to control motor axon pathway selection. Neuron 81:1297-1311
Santiago, Celine; Labrador, Juan-Pablo; Bashaw, Greg J (2014) The homeodomain transcription factor Hb9 controls axon guidance in Drosophila through the regulation of Robo receptors. Cell Rep 7:153-65
O'Donnell, Michael P; Bashaw, Greg J (2013) Distinct functional domains of the Abelson tyrosine kinase control axon guidance responses to Netrin and Slit to regulate the assembly of neural circuits. Development 140:2724-33

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