Weak ergodicity breaking of receptor motion in living cells stemming from random diffusivity

Molecular transport in living systems regulates numerous processes underlying biological function. Although many cellular components exhibit anomalous diffusion, only recently has the subdiffusive motion been associated with nonergodic behavior. These findings have stimulated new questions for their implications in statistical mechanics and cell biology. Is nonergodicity a common strategy shared by living systems? Which physical mechanisms generate it? What are its implications for biological function? Here, we use single particle tracking to demonstrate that the motion of DC-SIGN, a receptor with unique pathogen recognition capabilities, reveals nonergodic subdiffusion on living cell membranes. In contrast to previous studies, this behavior is incompatible with transient immobilization and therefore it can not be interpreted according to continuous time random walk theory. We show that the receptor undergoes changes of diffusivity, consistent with the current view of the cell membrane as a highly dynamic and diverse environment. Simulations based on a model of ordinary random walk in complex media quantitatively reproduce all our observations, pointing toward diffusion heterogeneity as the cause of DC-SIGN behavior. By studying different receptor mutants, we further correlate receptor motion to its molecular structure, thus establishing a strong link between nonergodicity and biological function. These results underscore the role of disorder in cell membranes and its connection with function regulation. Due to its generality, our approach offers a framework to interpret anomalous transport in other complex media where dynamic heterogeneity might play a major role, such as those found, e.g., in soft condensed matter, geology and ecology.Comment: 27 pages, 5 figure

The actin cytoskeleton modulates the activation of iNKT cells by segregating CD1d nanoclusters on antigen-presenting cells

Invariant natural killer T (iNKT) cells recognize endogenous and exogenous lipid antigens presented in the context of CD1d molecules. The ability of iNKT cells to recognize endogenous antigens represents a distinct immune recognition strategy, which underscores the constitutive memory phenotype of iNKT cells and their activation during inflammatory conditions. However, the mechanisms regulating such “tonic” activation of iNKT cells remain unclear. Here, we show that the spatiotemporal distribution of CD1d molecules on the surface of antigen-presenting cells (APCs) modulates activation of iNKT cells. By using superresolution microscopy, we show that CD1d molecules form nanoclusters at the cell surface of APCs, and their size and density are constrained by the actin cytoskeleton. Dual-color single-particle tracking revealed that diffusing CD1d nanoclusters are actively arrested by the actin cytoskeleton, preventing their further coalescence. Formation of larger nanoclusters occurs in the absence of interactions between CD1d cytosolic tail and the actin cytoskeleton and correlates with enhanced iNKT cell activation. Importantly and consistently with iNKT cell activation during inflammatory conditions, exposure of APCs to the Toll-like receptor 7/8 agonist R848 increases nanocluster density and iNKT cell activation. Overall, these results define a previously unidentified mechanism that modulates iNKT cell autoreactivity based on the tight control by the APC cytoskeleton of the sizes and densities of endogenous antigen-loaded CD1d nanoclusters

Mrp1 is involved in lipid presentation and iNKT cell activation by Streptococcus pneumoniae

The CD1d pathway present lipid antigens resulting in the activation of iNKT cells but the complete pathway remains to be fully elucidated. Here, Chandra et al. use an siRNA screen and identify Mrp1 as crucial for CD1d lipid presentation and activation of iNKT in the context of Streptococcus pneumoniae infection

Nanoclustering as a dominant feature of plasma membrane organization

Contains fulltext : 139177.pdf (publisher's version ) (Open Access)Early studies have revealed that some mammalian plasma membrane proteins exist in small nanoclusters. The advent of super-resolution microscopy has corroborated and extended this picture, and led to the suggestion that many, if not most, membrane proteins are clustered at the plasma membrane at nanoscale lengths. In this Commentary, we present selected examples of glycosylphosphatidyl-anchored proteins, Ras family members and several immune receptors that provide evidence for nanoclustering. We advocate the view that nanoclustering is an important part of the hierarchical organization of proteins in the plasma membrane. According to this emerging picture, nanoclusters can be organized on the mesoscale to form microdomains that are capable of supporting cell adhesion, pathogen binding and immune cell-cell recognition amongst other functions. Yet, a number of outstanding issues concerning nanoclusters remain open, including the details of their molecular composition, biogenesis, size, stability, function and regulation. Notions about these details are put forth and suggestions are made about nanocluster function and why this general feature of protein nanoclustering appears to be so prevalent

Healing Of Apicectomy Wounds In Dogs

Glycan-protein interactions are emerging as important modulators of membrane protein organization and dynamics, regulating multiple cellular functions. In particular, it has been postulated that glycan-mediated interactions regulate surface residence time of glycoproteins and endocytosis. How this precisely occurs is poorly understood. Here we applied single-molecule-based approaches to directly visualize the impact of glycan-based interactions on the spatiotemporal organization and interaction with clathrin of the glycosylated pathogen recognition receptor dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN). We find that cell surface glycan-mediated interactions do not influence the nanoscale lateral organization of DC-SIGN but restrict the mobility of the receptor to distinct micrometer-size membrane regions. Remarkably, these regions are enriched in clathrin, thereby increasing the probability of DC-SIGN-clathrin interactions beyond random encountering. N-glycan removal or neutralization leads to larger membrane exploration and reduced interaction with clathrin, compromising clathrin-dependent internalization of virus-like particles by DC-SIGN. Therefore, our data reveal that cell surface glycan-mediated interactions add another organization layer to the cell membrane at the microscale and establish a novel mechanism of extracellular membrane organization based on the compartments of the membrane that a receptor is able to explore. Our work underscores the important and complex role of surface glycans regulating cell membrane organization and interaction with downstream partners