Eaters of the Dead: How Glial Cells Respond to and Engulf Degenerating Axons in the CNS: A Dissertation

Abstract

Glia, whose name derives from the original Greek word, meaning “glue,” have long been understood to be cells that play an important functional role in the nutritive and structural support of the central nervous system, yet their full involvement has been historically undervalued. Despite the strong evidence that glial reactions to cellular debris govern the health of the nervous system, the specific properties of damaged axonal debris and the mechanisms by which glia sense them, morphologically adapt to their presence, and initiate phagocytosis for clearance, have remained poorly understood. The work presented in this thesis was aimed at addressing this fundamental gap in our understanding of the role for glia in neurodegenerative processes. I demonstrate that the cellular machinery responsible for the phagocytosis of apoptotic cell corpses is well conserved from worms to mammals. Draper is a key component of the glial response machinery and I am able to show here, for the first time, that it signals through Drosophila Shark, a non-receptor tyrosine kinase similar to mammalian Syk and Zap-70. Shark binds Draper through an immunoreceptor tyrosine-based activation motif (ITAM) in the Draper intracellular domain. I show that Shark activity is essential for Draper-mediated signaling events in vivo, including the recruitment of glial membranes to axons undergoing Wallerian degeneration. I further show that the Src family kinase (SFK) Src42A can markedly increase Draper phosphorylation and is essential for glial phagocytic activity. Therefore I propose that ligand-dependent Draper receptor activation initiates the Src42A-dependent tyrosine phosphorylation of Draper, the association of Shark and the subsequent downstream activation of the Draper pathway. I observed that these Draper-Src42A-Shark interactions are strikingly similar to mammalian immunoreceptor-SFK-Syk signaling events in myeloid and lymphoid cells. Thus, Draper appears to be an ancient immunoreceptor with an extracellular domain tuned to modified-self antigens and an intracellular domain that promotes phagocytosis through an ITAM domain-SFK-Syk-mediated signaling cascade. I have further identified the Drosophila guanine-nucleotide exchange factor (GEF) complex Crk/Mbc/dCed-12, and the small GTPase Rac1 as novel modulators of glial clearance of axonal debris. I am able to demonstrate that Crk/Mbc/dCed-12 and Rac1 function in a non-redundant fashion with the Draper pathway to promote a distinct step in the clearance of axonal debris. Whereas Draper signaling is required early during glial responses, promoting glial activation and extension of glial membranes to degenerating axons, the Crk/Mbc/dCed-12 complex functions at later stages of glial response, promoting the actual phagocytosis of axonal debris. Finally, many interesting mutants have been identified in primary screens for genes active in neurons that are required for axon fragmentation or clearance by glia, and genes potentially active in glia that orchestrate…