Receptor protein tyrosine phosphatases in vertebrate axon outgrowth
Date of Award
Doctor of Philosophy (Ph.D.)
First Committee Member
John L. Bixby - Committee Chair
Axon pathfinding is regulated by a diverse group of environmental guidance cues that bind to receptors on the growth cone, which in turn regulate the growth cone's movement. Receptor protein tyrosine phosphatases (RPTPs) have been shown to be involved in axon outgrowth in a variety of contexts. They have extracellular domains (ECDs) containing motifs found in cell adhesion molecules, and intracellular phosphatase domains. The most striking evidence for RPTP function in axon growth and guidance comes from Drosophila loss-of-function mutants for type IIa and type III RPTPs. These mutants have defects in fasciculation and axon targeting in motor neurons, as well as aberrant midline crossing. We examined the function of type IIa and type III RPTPs in axon outgrowth in the chick embryo. In vitro experiments using the ectodomain of the type III RPTP, PTPRO, demonstrated that PTPRO arts as an anti-adhesive, neurite-inhibitory factor for retinal neurons. The soluble PTPRO ECD can collapse retinal growth cones, and is a chemorepellant guidance cue modulated by cGMP. Using this PTPRO-Fc fusion protein, we immunoprecipitated at least one specific binding partner for PTPRO from the chick forebrain and tectum. We tested the involvement of PTPRO as well as two type IIa RPTPs (PTP-delta and PTP-sigma) in axon outgrowth in vivo using in ovo electroporation of dsRNA to produce targeted knock down of RPTP levels in the embryonic chick spinal cord. We found that knock down of PTPRO, PTP-delta, or PTP-sigma produced aberrant growth of the anterior iliotibialis, a superficial dorsal motor nerve. The nerve was decreased in size and exhibited changes in fasciculation in embryos electroporated with dsRNA targeting RPTPs, but was not affected in control embryos electroporated with dsRNA targeting choline acetyltransferase or axonin-1. The nerve was most strongly affected by decreased PTPRO expression. Surprisingly, coelectroporation with dsRNA targeting PTP-delta and PTPRO, or all three RPTPs, produced less severe phenotypes than electroporation with PTPRO dsRNA alone. These data indicate that there must be a balance in type IIa and type III phosphatase activity to regulate correct motor axon pathfinding, suggesting that the function of RPTPs has been conserved through evolution.
Stepanek, Laurie, "Receptor protein tyrosine phosphatases in vertebrate axon outgrowth" (2004). Dissertations from ProQuest. 2083.