Heparan sulfate (HS) and heparan sulfate proteoglycans (HSPGs) play significant roles in various biological processes. The long-term objective of this project is to understand the physiological roles of HS in mammalian brain development and function. Toward this goal, we propose to use conditional gene disruption of enzymes involved in HS synthesis. We have created conditional knockout of the HS polymerizing enzyme EXT1 targeted to the developing brain (Nes-EXT1 mice). Preliminary analysis of Nes- EXT1 mice has revealed multiple brain phenotypes that suggest functional defects in morphogen signaling, axon guidance, and synapse maturation. Based on these prior achievements, 3 aims are proposed.
Aim 1. Role of HS in CNS morphogenesis: The defects observed in Nes-EXT1 mutant mice strongly suggest the essential role of HS in multiple morphogen signaling pathways. Based on these preliminary observations, we will perform molecular and cellular analysis of CNS patterning in three key brain regions. Furthermore, we will compare morphological and molecular phenotypes between Nes-EXT1 mutant mice and conditional knockout mice of the key HS-modifying enzyme N-deacetylase/N-sulfotransferase. By these studies, we will define and dissect the physiological functions of HS in CNS patterning.
Aim 2. Role of HS in axon guidance: Another remarkable phenotype in Nes-EXT1 mutant mice is severe defects in commissural fiber tracts in the forebrain. The phenotype is similar to that of the mutant mice of netrin-1, an important axon guidance molecule that has strong affinity to heparin. Here we will combine histological, cell biological, and genetic studies to elucidate the role of HS in netrin-mediated axon guidance.
Aim 3. Role of HS in CNS synapses: HS has been suggested to be involved in the development and function of CNS synapses. Moreover, there are reports that hereditary multiple exostosis, a human bone syndrome caused by mutations of EXT genes, sometimes associates with mental disorders. To define the role of HS in synapses and the higher bran function, we will generate another EXT1 conditional knockout mice targeted to mature neurons using CaMKII-Cre mice. We will perform histological, cell biological, and physiological analyses to determine the physiological role of HS in CNS synapses.
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| Kantor, David B; Chivatakarn, Onanong; Peer, Katherine L et al. (2004) Semaphorin 5A is a bifunctional axon guidance cue regulated by heparan and chondroitin sulfate proteoglycans. Neuron 44:961-75 |
