1,721,001 research outputs found
Compartmentalization of signaling by vesicular trafficking: a shared building design for the immune synapse and the primary cilium
No full textAccumulating evidence underscores the immune synapse (IS) of naive T cells as a site of intense vesicular trafficking. At variance with helper and cytolytic effectors, which use the IS as a secretory platform to deliver cytokines and/or lytic granules to their cellular targets, this process is exploited by naive T cells as a means to regulate the assembly and maintenance of the IS, on which productive signaling and cell activation crucially depend. We have recently identified a role of the intraflagellar transport (IFT) system, which is responsible for the assembly of the primary cilium, in the non-ciliated T-cell, where it controls IS assembly by promoting polarized T-cell receptor recycling. This unexpected finding not only provides new insight into the mechanisms of IS assembly but also strongly supports the notion that the IS and the primary cilium, which are both characterized by a specialized membrane domain highly enriched in receptors and signaling mediators, share architectural similarities and are homologous structures. Here, we review our current understanding of vesicular trafficking in the regulation of the assembly and maintenance of the naive T-cell IS and the primary cilium, with a focus on the IFT system
Bidimentional in vitro angiogenic assays to study CCM pathogenesis: endothelial cell proliferation and migration
No full textCerebral cavernous malformation (CCM) is a cerebrovascular disorder of proven genetic origin characterized by abnormally dilated and leaky capillaries occurring mainly in the central nervous system, with a prevalence of 0.3–0.5% in the general population. Genetic studies have identified three genes associated to CCMs: KRIT1 (CCM1), MGC4607 (CCM2), and PDCD10 (CCM3), which account for about 50%, 20%, and 10% of the cases, respectively. The great advances in the knowledge of the physiopathological functions of CCM genes, such as their involvement in the angiogenic process, have allowed to propose distinct putative therapeutic compounds, which showed to be effective at least in limiting some pathological phenotypes in cellular and animal models of the disease. However, despite numerous efforts, targeted pharmacological therapies that improve the outcome of CCM disease are currently lacking. Here we describe simply and low-cost assays as in vitro endothelial cell proliferation and migration assays that can be used to better understand the role of CCM genes on endothelial cell functions and to screen potential new compounds for CCM therapy
Study of Molecular Interactions of CCM Proteins by Using a GAL4-Based Yeast Two-Hybrid Screening
No full textThe yeast two-hybrid system was originally designed to detect protein–protein interactions using yeast transcriptional activators. Since its original description, this technique has been extensively used to identify protein–protein interactions from many different organisms, thus providing a convenient mean to both screen for proteins that interact with a protein of interest and to characterize the known interaction between two proteins. Nowadays, the yeast two-hybrid screen remains one of the leading molecular tools to study protein–protein interactions in native intracellular conditions. In these years, the technique has improved to overcome the limitations of the original assay, and many efforts have been made to scale up the technique and to adapt it to large-scale studies. In addition, variations have been introduced to enlarge the range of proteins and interactors that can be assayed by hybrid-based approaches. Several groups studying molecular mechanisms underlying the Cerebral Cavernous Malformation disease have successfully used the yeast two-hybrid system or related methods to isolate, identify, and characterize molecular interactions involved in the onset and progression of the pathology. Here we describe general principles, strengths, and limits of the yeast two-hybrid technology, and the basic protocol for a yeast two-hybrid library screening and for a small-scale yeast two-hybrid assay by using a GAL4-based system
Disease models in cerebral cavernous malformations
No full textCerebral cavernous malformation (CCM) is a rare disease of genetic origin characterized by dilated and leaky capillaries occurring mainly in the central nervous system. CCM can arise sporadically or may be inherited as an autosomal dominant condition with incomplete penetrance and variable clinical expressivity. The sporadic form accounts for up to 80% of cases, whereas the familial form accounts for at least 20% of cases. Genetic studies have identified three genes associated with CCMs: KRIT1 (CCM1), MGC4607 (CCM2) and PDCD10 (CCM3). Recently, great advances in understanding the pathophysiology of CCM disease have been obtained thanks to the use of animal and cellular models displaying all or some of the pathological characteristics that are observed in the human disease. Despite interspecies differences and the difficulty in creating animal models that completely recapitulate the human CCM disease onset and progression, these models have been helpful in identifying new molecular mechanisms underlying CCM development and in testing novel pharmacological therapies
Non-autonomous effects of CCM genes loss
Full text linkCerebral cavernous malformation (CCM) is a rare disease of proven genetic origin characterized by vascular lesions affecting capillaries and small vessels of the central nervous system. CCM lesions occur in a range of different phenotypes, including wide differences in lesion number, size, and susceptibility to intracerebral hemorrhage. CCM lesion genesis requires loss of function of any of three genes, namely KRIT1 (CCM1), MGC4607 (CCM2), and PDCD10 (CCM3). These genes exert pleiotropic effects regulating multiple mechanisms involved in angiogenesis, cellular response, cell-cell and cell-matrix adhesion, cytoskeleton dynamics, and oxidative damage protection. Familial CCM is an autosomal-dominantly inherited disease in which the loss of any of the three CCM genes follows a two-hit mechanism. The heterozygous loss-of-function germline variants in one of the involved genes seems to be associated with a second postzygotic mutation, according to Knudson’s two-hit model of tumor suppressor genes. This review is an overview of very recent literature on CCM onset and progression focused on the novel concept that the loss of a CCM gene in a single cell is sufficient to induce vascular lesions. Mutated cells undergo clonal expansion and become able to promote the recruitment of non-mutated cells and to induce their angiogenic switch through the increased production of angiogenic factors and downregulation of antiangiogenic factors. A deep understanding of this process and the knowledge of unbalanced secreted factors will be useful to design new pharmacological strategies for CCM patients
Emerging Roles of the Intraflagellar Transport System in the Orchestration of Cellular Degradation Pathways
Full text linkCiliated cells exploit a specific transport system, the intraflagellar transport (IFT) system, to ensure the traffic of molecules from the cell body to the cilium. However, it is now clear that IFT activity is not restricted to cilia-related functions. This is strikingly exemplified by the observation that IFT proteins play important roles in cells lacking a primary cilium, such as lymphocytes. Indeed, in T cells the IFT system regulates the polarized transport of endosome-associated T cell antigen receptors and signaling mediators during assembly of the immune synapse, a specialized interface that forms on encounter with a cognate antigen presenting cell and on which T cell activation and effector function crucially depend. Cellular degradation pathways have recently emerged as new extraciliary functions of the IFT system. IFT proteins have been demonstrated to regulate autophagy in ciliated cells through their ability to recruit the autophagy machinery to the base of the cilium. We have now implicated the IFT component IFT20 in another central degradation process that also controls the latest steps in autophagy, namely lysosome function, by regulating the cation-independent mannose-6-phosphate receptor (CI-MPR)-dependent lysosomal targeting of acid hydrolases. This involves the ability of IFT20 to act as an adaptor coupling the CI-MPR to dynein for retrograde transport to the trans-Golgi network. In this short review we will discuss the emerging roles of IFT proteins in cellular degradation pathways
CCM proteins are key players in redox signaling and oxidative stress regulation in Cerebral Cavernous Malformations
No full textSynthesis mechanism of SiC–SiO2 core/shell nanowires grown by chemical vapor deposition
No full textCore-shell SiC-SiO2 nanowires were grown on silicon substrate with the chemical vapor deposition method using iron nitrate as promoter for the growth and CO as a carbon precursor, under atmospheric pressure and at a temperature of 1100 degrees C. The whole process involves two main stages: promoter conditioning (dewetting) and growth, by reaction with CO. The dewetting phase has been characterized by SEM and TEM techniques, x-ray diffraction and Raman spectroscopy. The results show that at the operating temperature, a solid-state reaction between the substrate and the promoter takes place with the formation of alpha-FeSi2. The growth of the nanowires begins after an induction time of about 5 min from the introduction of CO. The experimental data have been interpreted by considering a nucleation process involving a reaction between FeSi2 and CO. For the nanowires growth phase, a mechanism based on the Vapor-Liquid-Solid theory is proposed, compatible with the morphology of the drop-shaped particles present on the tip of the nanowires
Regulation of Selective B Cell Autophagy by the Pro-oxidant Adaptor p66SHC
Full text linkp66SHC is a pro-oxidant member of the SHC family of protein adaptors that acts as a negative regulator of cell survival. In lymphocytes p66SHC exploits both its adaptor and its reactive oxygen species (ROS)-elevating function to antagonize mitogenic and survival signaling and promote apoptosis. As a result, p66SHC deficiency leads to the abnormal expansion of peripheral T and B cells and lupus-like autoimmunity. Additionally, a defect in p66SHC expression is a hallmark of B cell chronic lymphocytic leukemia, where it contributes to the accumulation of long-lived neoplastic cells. We have recently provided evidence that p66SHC exerts a further layer of control on B cell homeostasis by acting as a new mitochondrial LC3-II receptor to promote the autophagic demise of dysfunctional mitochondria. Here we discuss this finding in the context of the autophagic control of B cell homeostasis, development, and differentiation in health and disease
Cooperation between Prostaglandin E2 and Epidermal Growth Factor Receptor in Cancer Progression: A Dual Target for Cancer Therapy
Full text linkIt is recognized that prostaglandin E2 (PGE2) is one key lipid mediator involved in chronic inflammation, and it is directly implicated in tumor development by regulating cancer cell growth and migration, apoptosis, epithelial–mesenchymal transition, angiogenesis, and immune escape. In addition, the expression of the enzymes involved in PGE2 synthesis, cyclooxygenase 2 (COX-2) and microsomal prostaglandin E synthase 1 (mPGES1), positively correlates with tumor progression and aggressiveness, clearly indicating the crucial role of the entire pathway in cancer. Moreover, several lines of evidence suggest that the COX2/mPGES1/PGE2 inflammatory axis is involved in the modulation of epidermal growth factor receptor (EGFR) signaling to reinforce the oncogenic drive of EGFR activation. Similarly, EGFR activation promotes the induction of COX2/mPGES1 expression and PGE2 production. In this review, we describe the interplay between COX2/mPGES1/PGE2 and EGFR in cancer, and new therapeutic strategies that target this signaling pathway, to outline the importance of the modulation of the inflammatory process in cancer fighting. © 2023 by the authors
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