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de Sautu, M.

Publications and source records attributed to de Sautu, M..

2 recordsLinked to original sources
bioRxiv2023

Deep learning assisted single particle tracking for automated correlation between diffusion and function

Sub-cellular diffusion in living systems reflects cellular processes and interactions. Recent advances in optical microscopy allow the tracking of this nanoscale diffusion of individual objects with an unprecedented level of precision. However, the agnostic and automated extraction of functional information from the diffusion of molecules and organelles within the sub-cellular environment, is labor-intensive and poses a significant challenge. Here we introduce DeepSPT, a deep learning framework to interpret the diffusional 2D or 3D temporal behavior of objects in a rapid and efficient manner, agnostically. Demonstrating its versatility, we have applied DeepSPT to automated mapping of the early events of viral infections, identifying distinct types of endosomal organelles, and clathrin-coated pits and vesicles with up to 95% accuracy and within seconds instead of weeks. The fact that DeepSPT effectively extracts biological information from diffusion alone indicates that besides structure, motion encodes function at the molecular and subcellular level.

biophysics↗
bioRxiv2023

The rotavirus VP5*/VP8* conformational transition permeabilizes membranes to Ca2+

Rotaviruses infect cells by delivering into the cytosol a transcriptionally active inner capsid particle (a "double-layer particle": DLP). Delivery is the function of a third, outer layer, which drives uptake from the cell surface into small vesicles from which the DLPs escape. In published work, we followed stages of rhesus rotavirus (RRV) entry by live-cell imaging and correlated them with structures from cryogenic electron microscopy and tomography (cryo-EM and cryo-ET). The virus appears to wrap itself in membrane, leading to complete engulfment and loss of Ca2+ from the vesicle produced by the wrapping. One of the outer-layer proteins, VP7, is a Ca2+-stabilized trimer; loss of Ca2+ releases both outer-layer proteins from the particle. The other outer-layer protein, VP4, activated by cleavage into VP8* and VP5*, is a trimer that undergoes a large-scale conformational rearrangement, reminiscent of the transition that viral fusion proteins undergo to penetrate a membrane. The rearrangement of VP5* thrusts a 250-residue, C-terminal segment of each of the three subunits outward, while allowing the protein to remain attached to the virus particle and to the cell being infected. We proposed that this segment inserts into the membrane of the target cell, enabling Ca2+ to cross. In the work reported here, we show the validity of key aspects of this proposed sequence. By cryo-EM studies of liposome-attached virions ("triple-layer particles": TLPs) and single-particle fluorescence imaging of liposome-attached TLPs, we confirm insertion of the VP4 C-terminal segment into the membrane and ensuing generation of a Ca2+ "leak". The results allow us to formulate a molecular description of early events in entry. We also discuss our observations in the context of other work on double-strand RNA virus entry.

microbiology↗