Biology subjects

Hamon, Y.

Publications and source records attributed to Hamon, Y..

4 recordsLinked to original sources

bioRxiv2023

Dia2 formin controls receptor activity by organizing plasma membrane lipid partitioning at the nanoscale

Activation of JAK/STAT signaling by IFN-{gamma} requires partitioning of IFN-{gamma}R into specific lipid nanodomains at the plasma membrane. Using IFN-{gamma}R as a proxy, we investigated the role of actin dynamics in the formation and organization of lipid nanodomains, a process that remains poorly understood. We identified formin Dia2/DIAPH3 as a specific and RhoA -dependent regulator of IFN-{gamma}-induced JAK/STAT signaling. Based on lipidomics and specific probes enabling membrane lipid imaging by super resolution microscopy, we demonstrate that Dia2 is required for proper assembly of sphingomyelin and cholesterol lipid complexes. Finally, we show that the disorganization of lipid nanodomains induced by Dia2 depletion results in drastic changes in nano-partitioning and activity of other membrane proteins, such as Thy1 and PD-L1. Our data establish, therefore, the central role of the RhoA-Dia2 axis in the regulation of IFN-{gamma} induced JAK/STAT signaling, and more broadly, in the nanoscale organization of the plasma membrane.

cell biology↗

bioRxiv2020

Controlling T cells shape, mechanics and activation by micropatterning.

We designed a strategy, based on a careful examination of the activation capabilities of proteins and antibodies used as substrates for adhering T cells, coupled to protein microstamping. This allowed us to control at the same time the position, shape, mechanics and activation state of T cells. Once adhered on shaped patterns we examined the capacities of T cells to be activated with soluble aCD3, in comparison to T cells adhered to a continuously decorated substrate with the same density of ligands. We show that, in our hand, adhering onto an anti CD45 (aCD45) antibody decorated surface is not affecting T cell calcium fluxes, even adhered on variable size micro-patterns. We further demonstrate this by expressing MEGF10 as a non immune adhesion receptor in T cells to obtain the very same spreading area on PLL substrates and Young modulus than immobilized cells on aCD45, while retaining similar activation capabilities using soluble aCD3 or through model APC contacts. We propose that our system is a way to test activation or anergy of T cells with defined adhesion and mechanical characteristics, and may allow to dissect fine details of these mechanisms since it allows to observe homogenised populations in standardized T cell activation assays.

biophysics↗

bioRxiv2019

Single-cell immuno-mechanics: rapid viscoelastic changes are a hallmark of early leukocyte activation.

To accomplish their critical task of removing infected cells and fighting pathogens, leukocytes activate by forming specialized interfaces with other cells. Using an innovative micropipette rheometer, we show in three different cell types that when stimulated by microbeads mimicking target cells, leukocytes become up to ten times stiffer and more viscous. These mechanical changes initiate within seconds after contact and evolve rapidly over minutes. Remarkably, leukocyte elastic and viscous properties evolve in parallel, preserving a well-defined ratio that constitutes a mechanical signature specific to each cell type. The current results indicate that simultaneously tracking both elastic and viscous properties during an active cell process provides a new way to investigate cell mechanical processes. Our findings also suggest that dynamic immuno-mechanical measurements provide an identifier of leukocyte type and an indicator of the cells state of activation.

biophysics↗

bioRxiv2018

A theoretical high-density nanoscopy study leads to the design of UNLOC, an unsupervised algorithm

Among the superresolution microscopy techniques, the ones based on serially imaging sparse fluorescent particles enable the reconstruction of high-resolution images by localizing single molecules. Although challenging, single-molecule localization microscopy (SMLM) methods aim at listing the position of individual molecules leading a proper quantification of the stoichiometry and spatial organization of molecular actors. However, reaching the precision requested to localize accurately single molecules is mainly constrained by the signal-to-noise ratio (SNR) but also the density (Dframe), i.e., the number of fluorescent particles per m2 per frame. Of central interest, we establish here a comprehensive theoretical study relying on both SNR and Dframe to delineate the achievable limits for accurate SMLM observations. We demonstrate that, for low-density hypothesis (i.e. one-Gaussian fitting hypothesis), any fluorescent particle biases the localization of a particle of interest when they are distant by less than {approx} 600 nm. Unexpectedly, we also report that even dim fluorescent particles should be taken into account to ascertain unbiased localization of any surrounding particles. Therefore, increased Dframe quickly deteriorates the localization precision, the image reconstruction and more generally the quantification accuracy. The first outcome is a standardized density-SNR space diagram to determine the achievable SMLM resolution expected with experimental data. Additionally, this study leads to the identification of the essential requirements for implementing UNLOC (UNsupervised particle LOCalization), an unsupervised and fast computing algorithm approaching the Cramer-Rao bound for particles at high-density per frame and without any prior on their intensity. UNLOC is available as an ImageJ plugin.

biophysics↗