A Fast Automatic Colocalization Method for 3D Live Cell and Super-Resolution Microscopy

Colocalizing two fluorescent-labeled proteins remains an open issue in diffraction-limited micro-scopy and raises new challenges with the emergence of super-resolution imaging, single molecule tagging (PALM, dSTORM...) and high content screening. Two distinct colocalization approaches are usually considered to address this problem : the intensity-based methods are very popular but are known to be sensitive to high intensity backgrounds and provide errors if the signal-to-noise ratio (SNR) is low ; the object-based methods analyze the spatial distribution of the two sets of detected spots by using point process statistics but unfortunately get rid of valuable information by reducing objects to points. We propose a unique method (GcoPS : Geo-coPositioning System) that reconciles intensity-based and object-based methods for various applications in both conventional diffraction-limited and super-resolution microscopy. Unlike previous methods, GcoPS is very fast, robust-to-noise and versatile since it efficiently handles 2D and 3D images, variable signal-to-noise ratios (SNR) and any kind of cell shapes and sizes. The experimental results demonstrate that GcoPS unequivocally outperforms the best competitive methods in adverse situations (noise, chromatic aberrations, ...). The method is able to automatically evaluate the colocalization between large regions and small dots and to detect significant negative colocalization. Since the one-parameter (p-value) GcoPS procedure is very fast in 2D and 3D, it should greatly facilitate objective analysis in large-scale high-content screening experiments

QuantEv: quantifying the spatial distribution of intracellular events

Analysis of the spatial distribution of endomembrane trafficking is fundamental to understand the mechanisms controlling cellular dynamics, cell homeostasy, and cell interaction with its external environment in normal and pathological situations. The development of automated methods to visualize and quantify the spatial distribution of intracellular events is essential to process the ever-increasing amount of data generated with modern light mi-croscopy. We present a generic and non-parametric framework to quantitatively analyze and visualize the spatio-temporal distribution of intracellular events from different conditions in fluorescence microscopy. From the spatial coordinates of intracellular features such as segmented subcellular structures or dynamic processes like vesicle trajectories, QuantEv automatically estimates weighted densities for each dimension of the 3D cylindrical coordinate system and performs a comprehensive statistical analysis from distribution distances. We apply this approach to study the spatio-temporal distribution of moving Rab6 fluorescently labeled membranes with respect to their direction of movement in cells constrained in crossbow-and disk-shaped fibronectin patterns. We also investigate the position of the generating hub of Rab11 positive membranes and the effect of actin disruption on Rab11 trafficking in coordination with cell shape. An Icy plugin and a tutorial are available athttp://icy.bioimageanalysis.org/plugin/QuantEv

Triptolide Inhibits the Proliferation of Prostate Cancer Cells and Down-Regulates SUMO-Specific Protease 1 Expression

Recently, traditional Chinese medicine and medicinal herbs have attracted more attentions worldwide for its anti-tumor efficacy. Celastrol and Triptolide, two active components extracted from the Chinese herb Tripterygium wilfordii Hook F (known as Lei Gong Teng or Thunder of God Vine), have shown anti-tumor effects. Celastrol was identified as a natural 26 s proteasome inhibitor which promotes cell apoptosis and inhibits tumor growth. The effect and mechanism of Triptolide on prostate cancer (PCa) is not well studied. Here we demonstrated that Triptolide, more potent than Celastrol, inhibited cell growth and induced cell death in LNCaP and PC-3 cell lines. Triptolide also significantly inhibited the xenografted PC-3 tumor growth in nude mice. Moreover, Triptolide induced PCa cell apoptosis through caspases activation and PARP cleavage. Unbalance between SUMOylation and deSUMOylation was reported to play an important role in PCa progression. SUMO-specific protease 1 (SENP1) was thought to be a potential marker and therapeutical target of PCa. Importantly, we observed that Triptolide down-regulated SENP1 expression in both mRNA and protein levels in dose-dependent and time-dependent manners, resulting in an enhanced cellular SUMOylation in PCa cells. Meanwhile, Triptolide decreased AR and c-Jun expression at similar manners, and suppressed AR and c-Jun transcription activity. Furthermore, knockdown or ectopic SENP1, c-Jun and AR expression in PCa cells inhibited the Triptolide anti-PCa effects. Taken together, our data suggest that Triptolide is a natural compound with potential therapeutic value for PCa. Its anti-tumor activity may be attributed to mechanisms involving down-regulation of SENP1 that restores SUMOylation and deSUMOyaltion balance and negative regulation of AR and c-Jun expression that inhibits the AR and c-Jun mediated transcription in PCa

Testing independence between two random sets for the analysis of colocalization in bioimaging

International audienc

Optimization of a Traffic Control Scheme for a Post-Disaster Urban Road Network

Traffic control of urban road networks during emergency rescues is conducive to rapid rescue in the affected areas. However, excessive control will lead to negative impacts on the normal traffic order. We propose a novel model to optimize the traffic control scheme during the post-disaster emergency rescue period named PD-TCM (post-disaster traffic control model). In this model, the vertex and edge betweenness indexes of urban road networks are introduced to evaluate the controllability of the road sections. The gravity field model is also used to adjust the travel time function of different road sections in the control and diverging domains. Experimental results demonstrate that the proposed model can obtain the optimal traffic control scheme efficiently, which gives it the ability to meet the demand of emergency rescues as well as reducing the disturbances caused by controls

Vinculin head–tail interaction defines multiple early mechanisms for cell substrate rigidity sensing

International audienc

Mechanical checkpoint for persistent cell polarization in adhesion-naive fibroblasts.

International audienceCell polarization is a fundamental biological process implicated in nearly every aspect of multicellular development. The role of cell-extracellular matrix contacts in the establishment and the orientation of cell polarity have been extensively studied. However, the respective contributions of substrate mechanics and biochemistry remain unclear. Here we propose a believed novel single-cell approach to assess the minimal polarization trigger. Using nonadhered round fibroblast cells, we show that stiffness sensing through single localized integrin-mediated cues are necessary and sufficient to trigger and direct a shape polarization. In addition, the traction force developed by cells has to reach a minimal threshold of 56 ± 1.6 pN for persistent polarization. The polarization kinetics increases with the stiffness of the cue. The polarized state is characterized by cortical actomyosin redistribution together with cell shape change. We develop a physical model supporting the idea that a local and persistent inhibition of actin polymerization and/or myosin activity is sufficient to trigger and sustain the polarized state. Finally, the cortical polarity propagates to an intracellular polarity, evidenced by the reorientation of the centrosome. Our results define the minimal adhesive requirements and quantify the mechanical checkpoint for persistent cell shape and organelle polarization, which are critical regulators of tissue and cell development