This study shows that the activity of neurons can trigger shedding of a protein, NG2, from the surface of oligodendrocyte precursor cells; this protein in turn modulates synaptic transmission, revealing a two-way conversation between neurons and glia.

The contribution of microglia to ischemic cortical stroke is of particular therapeutic interest because of the impact on the survival of brain tissue in the ischemic penumbra, a region that is potentially salvable upon a brain infarct. Whether or not tissue in the penumbra survives critically depends on blood flow and vessel perfusion. To study the role of microglia in cortical stroke and blood vessel stability, CX3CR1(+/GFP) mice were subjected to transient middle cerebral artery occlusion and then microglia were investigated using time-lapse two-photon microscopy in vivo. Soon after reperfusion, microglia became activated in the stroke penumbra and started to expand cellular protrusions towards adjacent blood vessels. All microglia in the penumbra were found associated with blood vessels within 24 h post reperfusion and partially fully engulfed them. In the same time frame blood vessels became permissive for blood serum components. Migration assays in vitro showed that blood serum proteins leaking into the tissue provided molecular cues leading to the recruitment of microglia to blood vessels and to their activation. Subsequently, these perivascular microglia started to eat up endothelial cells by phagocytosis, which caused an activation of the local endothelium and contributed to the disintegration of blood vessels with an eventual break down of the blood brain barrier. Loss-of-microglia-function studies using CX3CR1(GFP/GFP) mice displayed a decrease in stroke size and a reduction in the extravasation of contrast agent into the brain penumbra as measured by MRI. Potentially, medication directed at inhibiting microglia activation within the first day after stroke could stabilize blood vessels in the penumbra, increase blood flow, and serve as a valuable treatment for patients suffering from ischemic stroke.

The transmembrane proteoglycan NG2 is expressed by oligodendrocyte precursor cells (OPC), which migrate to axons during developmental myelination and remyelinate in the adult after migration to injured sites. Highly invasive glial tumors also express NG2. Despite the fact that NG2 has been implicated in control of OPC migration, its mode of action remains unknown. Here, we show in vitro and in vivo that NG2 controls migration of OPC through the regulation of cell polarity. In stab wounds in adult mice we show that NG2 controls orientation of OPC toward the wound. NG2 stimulates RhoA activity at the cell periphery via the MUPP1/Syx1 signaling pathway, which favors the bipolar shape of migrating OPC and thus directional migration. Upon phosphorylation of Thr-2256, downstream signaling of NG2 switches from RhoA to Rac stimulation. This triggers process outgrowth through regulators of front-rear polarity and we show using a phospho-mimetic form of NG2 that indeed NG2 recruits proteins of the CRB and the PAR polarity complexes to stimulate Rac activity via the GEF Tiam1. Our findings demonstrate that NG2 is a core organizer of Rho GTPase activity and localization in the cell, which controls OPC polarity and directional migration. This work also reveals CRB and PAR polarity complexes as new effectors of NG2 signaling in the establishment of front-rear polarity.

p190RhoGAP (p190A) is a negative regulator of RhoA and localizes to membrane protrusions, where its GAP activity is required for directional migration. Here, Binamé et al. identify the protrusion-localization sequence in p190A and show that cancer-associated mutations in this region affect p190A localization and function as well as tumor cell migration.

Movement of individual cells and of cellular cohorts, chains or sheets requires physical forces that are established through interactions of cells with their environment. In vivo, migration occurs extensively during embryonic development and in adults during wound healing and tumorigenesis. In order to identify the molecular events involved in cell movement, in vitro systems have been developed. These have contributed to the definition of a number of molecular pathways put into play in the course of migratory behaviours, such as mesenchymal and amoeboid movement. More recently, our knowledge of migratory modes has been enriched by analyses of cells exploring and moving through three-dimensional (3D) matrices. While the cells' morphologies differ in 2D and 3D environments, the basic mechanisms that put a cellular body into motion are remarkably similar. Thus, in both 2D and 3D, the polarity of the migrating cell is initially defined by a specific subcellular localization of signalling molecules and components of molecular machines required for motion. While the polarization can be initiated either in response to extracellular signalling or be a chance occurrence, it is reinforced and sustained by positive feedback loops of signalling molecules. Second, adhesion to a substratum is necessary to generate forces that will propel the cell engaged in either mesenchymal or ameboid migration. For collective cell movement, intercellular coordination constitutes an additional requirement: a cell cohort remains stationary if individual cells pull in opposite directions. Finally, the availability of space to move into is a general requirement to set cells into motion. Lack of free space is probably the main obstacle for migration of most healthy cells in an adult multicellular organism. Thus, the requirements for cell movement are both intrinsic to the cell, involving coordinated signalling and interactions with molecular machines, and extrinsic, imposed by the physicochemical nature of the environment. In particular, the geometry and stiffness of the support act on a range of signalling pathways that induce specific cell migratory responses. These issues are discussed in the present review in the context of published work and our own data on collective migration of hepatocyte cohorts.

Highlights d Rac1 immobilization at the lamellipodium tip correlates with its activation d Rac1 immobilization depends on effector binding, including WRC d RhoA does not display selective immobilization at the lamellipodium tip d Local Rac1 activation at the lamellipodium tip triggers membrane protrusion

An unresolved question regarding the physiopathology of hepatitis C virus (HCV) infection is the remarkable efficiency with which host defenses are neutralized to establish chronic infection. Modulation of an apoptotic response is one strategy used by viruses to escape immune surveillance. We previously showed that HCV proteins down-regulate expression of BH3-only Bcl2 interacting domain (Bid) in hepatocytes of HCV transgenic mice. As a consequence, cells acquire resistance to Fas-mediated apoptosis, which in turn leads to increased persistence of experimental viral infections in vivo. This mechanism might participate in the establishment of chronic infections and the resulting pathologies, including hepatocellular carcinoma. We now report that Bid is also down-regulated in patients in the context of noncirrhotic HCV-linked tumorigenesis and in the HCV RNA replicon system. We show that the nonstructural HCV viral protein NS5A is sufficient to activate a calpain cysteine protease, leading to degradation of Bid. Moreover, pharmacological inhibitors of calpains restore both the physiological levels of Bid and the sensitivity of cells toward a death receptor-mediated apoptotic signal. Finally, human HCV-related tumors and hepatocytes from HCV transgenic mice that display low Bid expression contain activated calpains. Conclusion: Calpains activated by HCV proteins degrade Bid and thus dampen apoptotic signaling. These results suggest that inhibiting calpains could lead to an improved efficiency of immune-mediated elimination of HCV-infected cells. (HEPATOLOGY 2009;50:1370-1379.)P ersistent infection with hepatitis C virus (HCV) is among the most common infectious causes of chronic liver disease. The majority of patients fail to clear the virus and become chronic carriers, with a persistent presence of detectable virus in the serum. 1 Patients with chronic hepatitis C are at risk for hepatic fibrosis, frequently culminating in hepatic cirrhosis and hepatocellular carcinoma (HCC). 2 FL-N/35 transgenic mice, with hepatocyte-targeted expression of the entire open reading frame of the genotype 1b HCV, are at risk for steatosis and hepatocellular adenoma and carcinoma. 3 We previously showed that the FL-N/35 hepatocytes are resistant to apoptosis induced by the Fas/CD95 death receptor stimulation. The lack of sensitivity to apoptotic stimulation was related to decreased expression of BH3-only Bcl2 interacting domain (Bid), a BH3-only member of the Bcl-2 family of apoptosis regulators. 4 Importantly, Bid-deficient hepatocytes are refractory to T lymphocyte-induced cell death, suggesting that apoptosis contributes to HCV persistence and, as a consequence, to liver pathologies characteristic of chronic HCV infection. 4 Here, we report that Bid is also down-regulated in the context of the subgenomic and genome-length HCV replicons and in a subset of HCV-linked human HCC.

Transforming growth factor ␤ (TGF-␤) has a strong impact on liver development and physiopathology, exercised through its pleiotropic effects on growth, differentiation, survival, and migration. When exposed to TGF-␤, the mhAT3F cells, immortalized, highly differentiated hepatocytes, maintained their epithelial morphology and underwent dramatic alterations of adhesion, leading to partial or complete detachment from a culture plate, followed by readhesion and spreading. These alterations of adhesive behavior were caused by sequential changes in expression of the ␣5␤1 integrin and of its ligand, the fibronectin. The altered specificity of anchorage to the extracellular matrix gave rise to changes in cells' collective motility: cohorts adhering to fibronectin maintained a persistent, directional motility, with ezrin-rich pathfinder cells protruding from the tips of the cohorts. The absence of adhesion to fibronectin prevented the appearance of polarized pathfinders and lead to random, oscillatory motility. Our data suggest a novel role for TGF-␤ in the control of collective migration of epithelial cohorts. INTRODUCTIONThe cytokines of the transforming growth factor ␤ (TGF-␤) family have essential and pleiotropic roles in embryonic development, adult homeostasis, and pathology (reviewed in Massague et al., 2000;Bachman and Park, 2005). Although the outcome of the TGF-␤ signaling is strongly dependent on its cross-talk with other signal transduction pathways, as a general rule it is cytostatic and cytotoxic for epithelia and the immune system, while it stimulates differentiation and migration of endothelial and mesenchymal cells (Siegel and Massague, 2003).Liver is one of the organs dramatically affected by the TGF-␤ signaling (reviewed in Bissell et al., 2001). Ectopic expression of the active form of TGF-␤1 gives rise to liver fibrosis through the differentiation of hepatic stellate cells into myofibroblasts and stimulation of collagen synthesis and deposition (Sanderson et al., 1995). During liver regeneration, TGF-␤ acts to control the transient stimulation of stellate cells and, through its growth inhibitory action on hepatocytes, the extent of hepatocyte regenerative proliferation (Romero-Gallo et al., 2005). Primary hepatocytes in culture are also sensitive to TGF-␤: in this setting the cytokine provokes a strong apoptotic response of both mature and fetal hepatocytes (Fabregat et al., 1996;Gressner et al., 1997), whereas early hepatic precursors survive the same treatment . The resistance to the apoptotic effects of TGF-␤ signaling can be achieved by simultaneous activation of MAPK Erk and PI3K survival signaling (Fabregat et al., 1996;Janda et al., 2002;Valdes et al., 2004), which likely accounts for continued survival and proliferation of many hepatoma cell lines in the presence of TGF-␤ (Gressner et al., 1997). Cross-talk with other signal transduction pathways (Janda et al., 2002;Cordenonsi et al., 2003;Kamaraju and Roberts, 2005;Mishra et al., 2005;Katuri et al., 2006), acquisition of mutations invalidati...