Membrane Tubulation Coupled with Fission by a Minimal Two-component Module

Abstract

Membrane tubulation coupled to fission is fundamental to numerous cellular processes. It manifests in all known forms of vesicular trafficking and its precise regulation is critical for inter and intra-cellular cross-talk. Over the years, research has significantly advanced our understanding of these processes separately and has identified candidate proteins capable of either membrane tubulation or fission. However, mechanisms regulating their coupling have so far remained unclear, mainly due to a dearth of assay systems allowing us to unambiguously probe early intermediates formed during the process. In this thesis, I describe our efforts at reconstituting a coupled membrane tubulation and fission reaction using a mixture of two proteins, the membrane tubulator BIN1 and the fission catalyst dynamin2. Using cushioned planar bilayer islands as novel membrane templates that allow real-time monitoring of membrane tubulation, we show how the dual ability of BIN1 to recruit dynamin2 to tubules, but prevent its engagement with the membrane allows for tubulation and fission to be coupled. As a result, fission manifests in a manner dependent on relative dynamin2 amounts. We uncover mechanistic insights into the process and discuss its significance in understanding muscle cell physiology.