Doctor of Philosophy

dissertationDuring development, axons are generated by specialized structures called growth cones in response to chemical guidance cues. When a growth cone reaches its target, a synapse is formed, the growth cone collapses, and the axon is stable. In mature neurons, the architecture and synaptic connections are then maintained as the neuron is subjected to growth, physical forces and age. I am studying this developmental transition: How do neurons switch from an embryonic state to a mature differentiated state? And how is the mature state maintained? To identify new genes required for axon stability we screened for C. elegans mutants with phenotypes in which growth cones do not collapse after synapse formation, but continue to sprout into adulthood. We found that mutations in casein kinase 1δ (kin- 20 in C. elegans) disrupt nervous system architecture as a result of continued growth cone sprouting; thus, a function of casein kinase 1δ is to stabilize neuronal architecture and inhibit growth cone sprouting. The identification of a kinase in this process suggests that the stabilization of axon morphology requires a signaling pathway. To identify the signaling pathway downstream of casein kinase 1δ, we performed a kin-20 suppressor screen. These screens identified 11 genes that when mutated restore stable mature neurons in the kin-20 mutant. Most of the suppressor mutations are in 9 genes associated with transcriptional pausing and termination: one mutation likely disinhibits the closely related CK1α and two of the suppressor mutations directly disrupt the termination site for the short form of unc-44. We found that casein kinase 1δ is required iv for the expression of the giant isoform of ankyrin / UNC-44, and that second-site mutations in the RNA polymerase-II termination complex restore expression of ankyrin and rescue the kin-20 casein kinase mutants. We found that CK1δ can phosphorylate the RNA polymerase II phosphatase SSUP-72 (found as a suppressor of kin-20) directly and based on genetics experiments, phosphorylation of SSUP-72 is sufficient for expression of long ankyrin. In summary, my dissertation led to the discovery of a novel axon maturation signaling pathway (Chapter 2). This pathway begins with a conserved kinase, which acts through an RNA-polymerase II termination phosphatase, to express a neuron-specific giant isoform of ankyrin, required for axon maturation and maintenance