The role of membrane protein phase separation in nuclear membrane sealing and fusion

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Membrane remodeling is essential to life. The self-assembly of the nucleus at the end of vertebrate mitosis exemplifies how efficiently and robustly cells reshape membranes, even up to the point of building an entire organelle from solubilized pieces. Our recent work suggests that nuclear membrane reformation at the end of mitosis proceeds via the complete fusion of incoming sheets and tubules of the endoplasmic reticulum (ER) on the surface of decondensing chromatin – a prerequisite for nuclear compartmentalization – by coupling membrane protein phase separation to the work of the cell’s membrane fusion machinery endosomal sorting complexes required for transport-III (ESCRT-III). In this proposal, we aim to investigate the yet undefined structural and biophysical principles by which the condensed intrinsically disordered domain (IDD)-containing membrane proteins LEM2 and MAN1 recruit the structured ESCRT-III machinery protein CHMP7 to form filaments and seal and finally fuse the nuclear membrane to complete nuclear reformation. We will reconstitute the LEM2/MAN1-ESCRT-III pathway components on membranes to study their properties using cryo electron microscopy (cryo-EM) and biophysics. Established assays monitoring nuclear self-assembly in mitosis will allow us to validate our models in vivo. A growing body of work links the LEM2-ESCRT-III pathway, and potentially MAN1, to a variety of age-related pathologies underscoring its critical role for cellular physiology and its therapeutic potential. Generating a first mechanistic framework of how protein phase separation controls nuclear membrane assembly will hence not only address fundamental questions of membrane organelle self-organization but will also be of medical relevance.