Microtubule-mitochondrial attachment facilitates cell division symmetry and mitochondrial partitioning in fission yeast

Association with microtubules inhibits the fission of mitochondria in Schizosaccharomyces pombe. Here, we show that this attachment of mitochondria to microtubules is an important cell-intrinsic factor in determining cell division symmetry. By comparing mutant cells that exhibited enhanced attachment and no attachment of mitochondria to microtubules (Dnm1Δ and Mmb1Δ, respectively), we show that microtubules in these mutants displayed aberrant dynamics compared to wild-type cells, which resulted in errors in nuclear positioning. This translated to cell division asymmetry in a significant proportion of both Dnm1Δ and Mmb1Δ cells. Asymmetric division in Dnm1Δ and Mmb1Δ cells resulted in unequal distribution of mitochondria, with the daughter cell that received more mitochondria growing faster than the other daughter cell. Taken together, we show the existence of homeostatic feedback controls between mitochondria and microtubules in fission yeast, which directly influence mitochondrial partitioning and, thereby, cell growth. This article has an associated First Person interview with the first author of the paper

Statistical predictions on the encapsulation of single molecule binding pairs into sized-dispersed nanocontainers

Single molecule experiments have recently attracted enormous interest. Many of these studies involve the encapsulation of a single molecule into nanoscale containers (such as vesicles, droplets and nanowells). In such cases, the single molecule encapsulation efficiency is a key parameter to consider in order to get a statistically significant quantitative information. It has been shown that such encapsulation typically follows a Poisson distribution and such theory of encapsulation has only been applied to the encapsulation of single molecules into perfectly sized monodispersed containers. However, experimentally nanocontainers are usually characterized by a size distribution, and often just a single binding pair (rather than a single molecule) is required to be encapsulated. Here the use of Poisson distribution is extended to predict the encapsulation efficiency of two different molecules in an association equilibrium. The Poisson distribution is coupled with a log-normal distribution in order to consider the effect of the container size distribution, and the effect of adsorption to the container is also considered. This theory will allow experimentalists to determine what single molecule encapsulation efficiency can be expected as a function of the experimental conditions. Two case studies, based on experimental data, are given to support the theoretical predictions

Retromer has a selective function in cargo sorting via endosome transport carriers

Retromer is a peripheral membrane protein complex that coordinates multiple vesicular trafficking events within the endolysosomal system. Here, we demonstrate that retromer is required for the maintenance of normal lysosomal morphology and function. The knockout of retromer subunit Vps35 causes an ultrastructural alteration in lysosomal structure and aberrant lysosome function, leading to impaired autophagy. At the whole-cell level, knockout of retromer Vps35 subunit reduces lysosomal proteolytic capacity as a consequence of the improper processing of lysosomal hydrolases, which is dependent on the trafficking of the cation-independent mannose 6-phosphate receptor (CI-M6PR). Incorporation of CI-M6PR into endosome transport carriers via a retromer-dependent process is restricted to those tethered by GCC88 but not golgin-97 or golgin-245. Finally, we show that this retromer-dependent retrograde cargo trafficking pathway requires SNX3, but not other retromer-associated cargo binding proteins, such as SNX27 or SNX-BAR proteins. Therefore, retromer does contribute to the retrograde trafficking of CI-M6PR required for maturation of lysosomal hydrolases and lysosomal function.Yi Cui, Julian M. Carosi, Zhe Yang, Nicholas Ariotti, Markus C. Kerr, Robert G. Parton, Timothy J. Sargeant, and Rohan D. Teasdal

Imaging Immune and Metabolic Cells of Visceral Adipose Tissues with Multimodal Nonlinear Optical Microscopy

Visceral adipose tissue (VAT) inflammation is recognized as a mechanism by which obesity is associated with metabolic diseases. The communication between adipose tissue macrophages (ATMs) and adipocytes is important to understanding the interaction between immunity and energy metabolism and its roles in obesity-induced diseases. Yet visualizing adipocytes and macrophages in complex tissues is challenging to standard imaging methods. Here, we describe the use of a multimodal nonlinear optical (NLO) microscope to characterize the composition of VATs of lean and obese mice including adipocytes, macrophages, and collagen fibrils in a label-free manner. We show that lipid metabolism processes such as lipid droplet formation, lipid droplet microvesiculation, and free fatty acids trafficking can be dynamically monitored in macrophages and adipocytes. With its versatility, NLO microscopy should be a powerful imaging tool to complement molecular characterization of the immunity-metabolism interface

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

Mechanisms of T cell organotropism

F.M.M.-B. is supported by the British Heart Foundation, the Medical Research Council of the UK and the Gates Foundation

Cancer stem cell drugs target K-ras signaling in a stemness context

Cancer stem cells (CSCs) are considered to be responsible for treatment relapse and have therefore become a major target in cancer research. Salinomycin is the most established CSC inhibitor. However, its primary mechanistic target is still unclear, impeding the discovery of compounds with similar anti-CSC activity. Here, we show that salinomycin very specifically interferes with the activity of K-ras4B, but not H-ras, by disrupting its nanoscale membrane organization. We found that caveolae negatively regulate the sensitivity to this drug. On the basis of this novel mechanistic insight, we defined a K-ras-associated and stem cell-derived gene expression signature that predicts the drug response of cancer cells to salinomycin. Consistent with therapy resistance of CSC, 8% of tumor samples in the TCGA-database displayed our signature and were associated with a significantly higher mortality. Using our K-ras-specific screening platform, we identified several new candidate CSC drugs. Two of these, ophiobolin A and conglobatin A, possessed a similar or higher potency than salinomycin. Finally, we established that the most potent compound, ophiobolin A, exerts its K-ras4B-specific activity through inactivation of calmodulin. Our data suggest that specific interference with the K-ras4B/calmodulin interaction selectively inhibits CSC.Peer reviewe