Fusion of secretory vesicles isolated from rat liver

Secretory vesicles isolated from rat liver were found to fuse after exposure to Ca2+. Vescle fusion is characterized by the occurrence of twinned vesicles with a continuous cleavage plane between two vesicles in freeze-fracture electron microscopy. The number of fused vesicles increases with increasing Ca2+-concentrations and is half maximal around 10–6 m. Other divalent cations (Ba2+, Sr2+, and Mg2+) were ineffective. Mg2+ inhibits Ca2+-induced fusion. Therefore, the fusion of secretory vesiclesin vitro is Ca2+ specific and exhibits properties similar to the exocytotic process of various secretory cells. Various substances affecting secretionin vivo (microtubular inhibitors, local anethetics, ionophores) were tested for their effect on membrane fusion in our system. The fusion of isolated secretory vesicles from liver was found to differ from that of pure phospholipid membranes in its temperature dependence, in its much lower requirement for Ca2+, and in its Ca2+-specificity. Chemical and enzymatic modifications of the vesicle membrane indicate that glycoproteins may account for these differences

NET1-mediated RhoA activation facilitates lysophosphatidic acid-induced cell migration and invasion in gastric cancer

The most lethal aspects of gastric adenocarcinoma (GA) are its invasive and metastatic properties. This aggressive phenotype remains poorly understood. We have recently identified neuroepithelial cell transforming gene 1 (NET1), a guanine exchange factor (GEF), as a novel GA-associated gene. Neuroepithelial cell transforming gene 1 expression is enhanced in GA and it is of functional importance in cell invasion. In this study, we demonstrate the activity of NET1 in driving cytoskeletal rearrangement, a key pathological mechanism in gastric tumour cell migration and invasion. Neuroepithelial cell transforming gene 1 expression was increased 10-fold in response to treatment with lysophosphatidic acid (LPA), resulting in an increase in active levels of RhoA and a 2-fold increase in cell invasion. Lysophosphatidic acid-induced cell invasion and migration were significantly inhibited using either NET1 siRNA or a RhoA inhibitor (C3 exoenzyme), thus indicating the activity of both NET1 and RhoA in gastric cancer progression. Furthermore, LPA-induced invasion and migration were also significantly reduced in the presence of cytochalasin D, an inhibitor of cytoskeletal rearrangements. Neuroepithelial cell transforming gene 1 knockdown resulted in AGS cell rounding and a loss of actin filament organisation, demonstrating the function of NET1 in actin organisation. These data highlight the importance of NET1 as a driver of tumour cell invasion, an activity mediated by RhoA activation and cytoskeletal reorganisation

Systems analysis of apoptosis protein expression allows the case-specific prediction of cell death responsiveness of melanoma cells.

Many cancer entities and their associated cell line models are highly heterogeneous in their responsiveness to apoptosis inducers and, despite a detailed understanding of the underlying signaling networks, cell death susceptibility currently cannot be predicted reliably from protein expression profiles. Here, we demonstrate that an integration of quantitative apoptosis protein expression data with pathway knowledge can predict the cell death responsiveness of melanoma cell lines. By a total of 612 measurements, we determined the absolute expression (nM) of 17 core apoptosis regulators in a panel of 11 melanoma cell lines, and enriched these data with systems-level information on apoptosis pathway topology. By applying multivariate statistical analysis and multi-dimensional pattern recognition algorithms, the responsiveness of individual cell lines to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) or dacarbazine (DTIC) could be predicted with very high accuracy (91 and 82% correct predictions), and the most effective treatment option for individual cell lines could be pre-determined in silico. In contrast, cell death responsiveness was poorly predicted when not taking knowledge on protein-protein interactions into account (55 and 36% correct predictions). We also generated mathematical predictions on whether anti-apoptotic Bcl-2 family members or x-linked inhibitor of apoptosis protein (XIAP) can be targeted to enhance TRAIL responsiveness in individual cell lines. Subsequent experiments, making use of pharmacological Bcl-2/Bcl-xL inhibition or siRNA-based XIAP depletion, confirmed the accuracy of these predictions. We therefore demonstrate that cell death responsiveness to TRAIL or DTIC can be predicted reliably in a large number of melanoma cell lines when investigating expression patterns of apoptosis regulators in the context of their network-level interplay. The capacity to predict responsiveness at the cellular level may contribute to personalizing anti-cancer treatments in the future

Chance mechanisms affecting the burden of metastases

BACKGROUND: The burden of cancer metastases within an individual is commonly used to clinically characterize a tumor's biological behavior. Assessments like these implicitly assume that spurious effects can be discounted. Here the influence of chance on the burden of metastasis is studied to determine whether or not this assumption is valid. METHODS: Monte Carlo simulations were performed to estimate tumor burdens sustained by individuals with cancer, based upon empirically derived and validated models for the number and size distributions of metastases. Factors related to the intrinsic metastatic potential of tumors and their host microenvironments were kept constant, to more clearly demonstrate the contribution from chance. RESULTS: Under otherwise identical conditions, both the simulated numbers and the sizes of metastases were highly variable. Comparable individuals could sustain anywhere from no metastases to scores of metastases, and the sizes of the metastases ranged from microscopic to macroscopic. Despite the marked variability in the number and sizes of the metastases, their respective growth times were rather more narrowly distributed. In such situations multiple occult metastases could develop into fully overt lesions within a comparatively short time period. CONCLUSION: Chance can have a major effect on the burden of metastases. Random variability can be so great as to make individual assessments of tumor biology unreliable, yet constrained enough to lead to the apparently simultaneous appearance of multiple overt metastases