Pucadyil Lab
Mechanistic and regulatory aspects of membrane tubulation, fission and fusion
Cells are compartmentalised into various membrane-bound organelles, which are highly dynamic in shape and composition. They are formed and maintained by the constant exchange of material between donor and acceptor compartments via vesicular transport carriers. These carriers are formed by membrane tubulation of the donor membrane. Membrane fission releases them from the donor compartment, and their fusion with the acceptor compartment completes the transport cycle.
The throughput and fidelity of vesicular transport pathways relies on a diverse and complex set of proteins, as well as an intricate coordination between the various step of the transport cycle. Remarkably, many of the participant proteins are conserved from yeast to humans, thus highlighting their importance to sustain life. Furthermore, mutations in many of the participant proteins are linked to various developmental disorders and several pathogens subvert the intrinsic coordination in vesicular transport pathways during infection.
Movie credit: Soumya Bhattacharyya
At the Pucadyil lab, we focus on discovering proteins that orchestrate membrane tubulation, fission and fusion and understand their mechanism.
Evolution of membrane intermediates during vesicular transport
Membrane tubulation, fission and fusion are extremely dynamic processes that involve a myriad of proteins in cells.
But at the core of these processes lies the ability of specific proteins to have evolved to overcome the hydrophobic effect, which otherwise ensures that the membrane retains its planar topology. In other words, these proteins manage to bend membrane out of shape - a process collectively referred to as membrane remodelling.
Original image credit: Himani Khurana
Reconstituting membrane remodelling ex vivo
Cellular complexity, while fascinating, frequently comes in the way of assign specific functions to proteins and building testable models. To get around these problems, we use a bottom-up reconstitution approach that allows recreating membrane tubulation, fission and fusion with specific proteins or their combinations on a define membrane template outside of the cell - a process that is referred to as reconstitution.
Reconstituted tubulation and fission of Supported Membrane Templates. Movie credits: Soumya Bhattacharyya and Srishti Dar
For our reconstitution efforts, we utilise Supported Membrane Templates (SMrT) that represent a versatile assay system displaying a wide range of topologies, from planar bilayers to highly curved membrane tubes, resting on a passive glass surface (see schematic above). Our experimental workflow is simple and typically involves the real-time monitoring of membrane tubulation, fission and fusion using fluorescence microscopy.
Read more here
"Gearing" up for dynamin-catalyzed membrane fission
Khurana and Pucadyil (2023) Current Opinions in Cell Biology
Membrane contacts, lipid flux, and fission
Pucadyil (2023) Molecular Cell
Function and regulation of the divisome for mitochondrial fission
Kraus and Roy et al. (2021) Nature
Cellular functions and intrinsic attributes of the ATP-binding Eps15 homology domain-containing (EHD) proteins
Bhattacharyya and Pucadyil (2020) Protein Science