Determining how neurons are correctly specified and assembled into functional circuits will provide critical insight into developmental disorders of th nervous system and may suggest therapeutic approaches to promote nerve regeneration. To achieve this goal it is important to understand how axon responses to conserved families of axon guidance cues are regulated. Slit and Netrin, and their Robo and Fra/DCC receptors, are highly conserved signaling molecules that regulate multiple aspects of circuit development. Here, we propose to investigate how responses to Slit and Netrin are regulated by defining functional and molecular links between conserved transcriptional regulators that impart neuronal subtype identity and the cell surface axon guidance receptors for Slit and Netrin that coordinate motor and midline axon guidance. In addition, we propose to explore a newly discovered mechanism through which the Frazzled/DCC receptor intracellular domain (ICD) itself can regulate transcription to negatively regulate responses to the midline repellant Slit. Te developing Drosophila embryonic CNS is an ideal system to explore transcriptional mechanisms that regulate axon guidance because of the availability of powerful genetic approaches and the evolutionary conservation of the transcription factors and cell surface receptors that coordinate circuit assembly.
The aims of this proposal are to determine how the Slit receptor Robo2 and the Netrin receptor Frazzled (Fra) are regulated by transcription factors, including Hb9, Nkx6 and Islet, and how, in turn, this regulation instructs pathway selection in defined populations of motor and interneurons. We will also use FACs sorting of defined subsets of motor neurons, together with transcript profiling in wild type and mutant backgrounds, to systematically identify additional effectors of these transcriptional programs. Finally, using a combination of robust in vitro and in vivo genetic and biochemical strategies, we will evaluate the hypothesis that in order to promote midline crossing, the Fra receptor undergoes gamma-secretase dependent proteolysis to release an intracellular domain (ICD) fragment that translocates to the nucleus to regulate its target gene commissureless (a key negative regulator of midline repulsion). Our proposed research will inform studies of homologous proteins in mammalian systems and could provide pharmacologic and genetic strategies to manipulate the specification of neuronal subtypes and receptor signaling. Furthermore, the results of our research may suggest new therapeutic targets for diverse disorders of the nervous system.

Public Health Relevance

Understanding how neurons are correctly specified and assembled into functional circuits will provide critical insight into developmental disorders of th nervous system. In addition, the ability to control the acquisition of specific neural cell fatesin cultured stem cells is prerequisite to the development of cell replacement therapies for numerous neurodegenerative disorders, including Alzheimer's and Parkinson's disease. Our current knowledge of the molecular genetic pathways that control differentiation and connectivity are incomplete. The research in this proposal aims to define molecular mechanisms that translate neuronal identity into wiring specificity in the nervous system.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS054739-06
Application #
8741994
Study Section
Neurodifferentiation, Plasticity, and Regeneration Study Section (NDPR)
Program Officer
Riddle, Robert D
Project Start
2006-04-01
Project End
2017-07-31
Budget Start
2014-09-01
Budget End
2015-07-31
Support Year
6
Fiscal Year
2014
Total Cost
$332,213
Indirect Cost
$116,149
Name
University of Pennsylvania
Department
Neurosciences
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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
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 81:1297-311
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
O'Donnell, Michael P; Bashaw, Greg J (2013) Src inhibits midline axon crossing independent of Frazzled/Deleted in Colorectal Carcinoma (DCC) receptor tyrosine phosphorylation. J Neurosci 33:305-14
Evans, Timothy A; Bashaw, Greg J (2012) Slit/Robo-mediated axon guidance in Tribolium and Drosophila: divergent genetic programs build insect nervous systems. Dev Biol 363:266-78
Coleman, Hope A; Labrador, Juan-Pablo; Chance, Rebecca K et al. (2010) The Adam family metalloprotease Kuzbanian regulates the cleavage of the roundabout receptor to control axon repulsion at the midline. Development 137:2417-26
Evans, Timothy A; Bashaw, Greg J (2010) Functional diversity of Robo receptor immunoglobulin domains promotes distinct axon guidance decisions. Curr Biol 20:567-72
Yang, Long; Garbe, David S; Bashaw, Greg J (2009) A frazzled/DCC-dependent transcriptional switch regulates midline axon guidance. Science 324:944-7