Organization of Postsynaptic Assembly by Filamin at the Drosophila Neuromuscular Junction

Abstract

Proper postsynaptic assembly is a fundamental requirement for circuit function, yet our understanding of its molecular mechanism is incomplete. The Drosophila neuromuscular junction is a powerful system for genetically dissecting synapse organization at the molecular level. Taking advantage of its features, I sought to identify novel molecular mechanisms governing synapse assembly. The Ral pathway for activity-dependent membrane addition is highly localized at the postsynapse and is essential to proper postsynaptic development, thus offering a tractable ideal starting point. Through a candidate-based genetic approach using Ral localization as the initial readout, I initially discovered that filamin is required for localizing the Ral pathway at the postsynapse. Filamin is a scaffolding and actin-binding protein not previously known to be required for synaptic development. My findings now show that the Drosophila ortholog filamin/cheerio localizes key signaling proteins that govern morphological plasticity and the composition of glutamate receptor clusters at the neuromuscular junction (NMJ). Genetic and biochemical analysis revealed that the short FLN90 isoform of filamin, which lacks the actin-binding domain, is required in the muscle and present surrounding boutons. FLN90 is required for dPak localization and, downstream of dPak, for localization of the Ral pathway for activity-dependent membrane addition. Consequently filamin is needed for growth of the subsynaptic reticulum, the postsynaptic membrane structure at the NMJ. In addition, in the absence of filamin, GluRIIA subunits are lacking though GluRIIB subunits cluster correctly. The shift in subunit composition is likely dependent on dPak, but independent of the Ral pathway. Based on these findings, I present a model in which filamin orchestrates a pathway governing two distinct aspects of synapse formation, morphological plasticity and subtype-specific receptor clustering. In Chapter 1 I provide general introduction to the relevant background topics. In Chapter 2 I present and discuss the major findings of my thesis research. Based on these, I discuss some of the remaining questions and future directions in Chapters 3 and 4.