The plasma membrane is a complex medium where transmembrane proteins diffuse and interact to facilitate cell function. Membrane protein mobility is affected by multiple mechanisms, including crowding, trapping, medium elasticity and structure, thus limiting our ability to distinguish them in intact cells. Here we characterize the mobility and organization of a short transmembrane protein at the plasma membrane of live T cells, using single particle tracking and photoactivated-localization microscopy. Protein mobility is highly heterogeneous, subdiffusive and ergodic-like. Using mobility characteristics, we segment individual trajectories into subpopulations with distinct Gaussian step-size distributions. Particles of low-to-medium mobility consist of clusters, diffusing in a viscoelastic and fractal-like medium and are enriched at the centre of the cell footprint. Particles of high mobility undergo weak confinement and are more evenly distributed. This study presents a methodological approach to resolve simultaneous mixed subdiffusion mechanisms acting on polydispersed samples and complex media such as cell membranes.
Bibliographical noteFunding Information:
We thank Yasmine Meroz (Harvard University) and Ralf Metzler (Potsdam University) for fruitful discussions and Roland Schwarzer (Gladstone Institute) and AndreasHerrmann (Humboldt University) for providing gp41 plasmids with fluorescent tags. This research was supported by Grant no. 321993 from the Marie Sklodowska-Curie actions of the European Commission, and Grants no. 1417/13 and no. 1937/13 from the Israeli Science Foundation.