Monitoring the Size and Lateral Dynamics of ErbB1 Enriched Membrane Domains through Live Cell Plasmon Coupling Microscopy

To illuminate the role of the spatial organization of the epidermal growth factor receptor (ErbB1) in signal transduction quantitative information about the receptor topography on the cell surface, ideally on living cells and in real time, are required. We demonstrate that plasmon coupling microscopy (PCM) enables to detect, size, and track individual membrane domains enriched in ErbB1 with high temporal resolution. We used a dendrimer enhanced labeling strategy to label ErbB1 receptors on epidermoid carcinoma cells (A431) with 60 nm Au nanoparticle (NP) immunolabels under physiological conditions at 37°C. The statistical analysis of the spatial NP distribution on the cell surface in the scanning electron microscope (SEM) confirmed a clustering of the NP labels consistent with a heterogeneous distribution of ErbB1 in the plasma membrane. Spectral shifts in the scattering response of clustered NPs facilitated the detection and sizing of individual NP clusters on living cells in solution in an optical microscope. We tracked the lateral diffusion of individual clusters at a frame rate of 200 frames/s while simultaneously monitoring the configurational dynamics of the clusters. Structural information about the NP clusters in their membrane confinements were obtained through analysis of the electromagnetic coupling of the co-confined NP labels through polarization resolved PCM. Our studies show that the ErbB1 receptor is enriched in membrane domains with typical diameters in the range between 60–250 nm. These membrane domains exhibit a slow lateral diffusion with a diffusion coefficient of  = |0.0054±0.0064| µm2/s, which is almost an order of magnitude slower than the mean diffusion coefficient of individual NP tagged ErbB1 receptors under identical conditions

Role of the endocytosis of caveolae in intracellular signaling and metabolism

Caveolae are 60-80 nm invaginated plasma membrane (PM) nanodomains, with a specific lipid and protein composition, which assist and regulate multiple processes in the plasma membrane-ranging from the organization of signalling complexes to the mechanical adaptation to changes in PM tension. However, since their initial descriptions, these structures have additionally been found tightly linked to internalization processes, mechanoadaptation, to the regulation of signalling events and of endosomal trafficking. Here, we review caveolae biology from this perspective, and its implications for cell physiology and disease

Microbial efflux systems and inhibitors: Approaches to drug discovery and the challenge of clinical implementation

Conventional antimicrobials are increasingly ineffective due to the emergence of multidrug-resistance among pathogenic microorganisms. The need to overcome these deficiencies has triggered exploration for novel and unconventional approaches to controlling microbial infections. Multidrug efflux systems (MES) have been a profound obstacle in the successful deployment of antimicrobials. The discovery of small molecule efflux system blockers has been an active and rapidly expanding research discipline. A major theme in this platform involves efflux pump inhibitors (EPIs) from natural sources. The discovery methodologies and the available number of natural EPI-chemotypes are increasing. Advances in our understanding of microbial physiology have shed light on a series of pathways and phenotypes where the role of efflux systems is pivotal. Complementing existing antimicrobial discovery platforms such as photodynamic therapy (PDT) with efflux inhibition is a subject under investigation. This core information is a stepping stone in the challenge of highlighting an effective drug development path for EPIs since the puzzle of clinical implementation remains unsolved. This review summarizes advances in the path of EPI discovery, discusses potential avenues of EPI implementation and development, and underlines the need for highly informative and comprehensive translational approaches