Disruption of cell-matrix interactions can lead to anoikis -apoptosis due to loss of matrix contacts. Altered fibronectin (FN) induces anoikis of primary human fibroblasts by a novel signaling pathway characterized by reduced phosphorylation of focal adhesion kinase (FAK). However, the receptors involved are unknown. FAK phosphorylation is regulated by nerve/glial antigen 2 (NG2) receptor signaling through PKCa a point at which signals from integrins and proteoglycans may converge. We found that an altered FN matrix induced anoikis in fibroblasts by upregulating NG2 and downregulating integrin a4. Suppressing NG2 expression or overexpressing a4 rescued cells from anoikis. NG2 overexpression alone induced apoptosis and, by reducing FAK phosphorylation, increased anoikis induced by an altered matrix. NG2 overexpression or an altered matrix also suppressed PKCa expression, but overexpressing integrin a4 enhanced FAK phosphorylation independently of PKCa. Cotransfection with NG2 cDNA and integrin a4 siRNA did not lower PKCa and pFAK levels more than transfection with either alone. PKCa was upstream of FAK phosphorylation, as silencing PKCa decreased FAK phosphorylation. PKCa overexpression reversed this behavior and rescued cells from anoikis. Thus, NG2 is a novel proapoptotic receptor, and NG2 and integrin a4 oppositely regulate anoikis in fibroblasts. NG2 and integrin a4 regulate FAK phosphorylation by PKCa-dependent and -independent pathways, respectively. Cell Death and Differentiation (2008) The extracellular matrix (ECM) glycoprotein fibronectin (FN) affects adhesion, migration, survival, and other cellular functions. FN fragments found in vivo and associated with disease or comparable recombinant fragments trigger apoptosis/anoikis in primary human fibroblasts via a novel pathway regulated by decreases in focal adhesion kinase (FAK) phosphorylation and downregulation of p53.1-3 However, the receptors involved are not known. Two classes of cellsurface receptors, integrins and proteoglycans, interact with FN and could mediate the anoikis. This process is regulated by FAK, an integrin-dependent non-receptor tyrosine kinase, consistent with a role for integrins. Also proteoglycans are involved in this apoptotic mechanism. 1Integrins are a and b heterodimeric cell-surface receptors that mediate cell-ECM interactions and regulate cell adhesion, migration, proliferation, and survival. Interactions between FN and integrins promote the survival of many cell types. We showed that blocking antibodies against the a4 integrin subunit induced apoptosis-like cell rounding in human fibroblasts, similar to that induced by anoikis in response to an altered FN matrix fragment.2 Integrins mediate those responses by binding ECM ligands and activating signaling cascades that promote these actions. The a4 integrin subunit can bind to at least three interaction sites on FN. These include the arginine-glycine-aspartic acid site on the central cell-binding domain, the V region, and the heparin-binding domain. Once engaged, integrins ...
Gene transfer in the lung holds promise for the treatment for 5 days thereafter. When DNA was delivered in a 50% of diseases such as pulmonary fibrosis, cystic fibrosis and suspension of Exosurf, the expression of either CAT or asthma. Pulmonary surfactant has been reported to Luc was significantly reduced by 89.6 ± 1.4% and enhance expression from endobronchial, adenovirus-82.7 ± 10.5%, respectively. The decrease in Luc mediated gene transfer in experimental animals. This study expression was closely correlated (r = 0.99, P Ͻ 0.001) to examines the effect of exogenous synthetic surfactant log concentration of surfactant in the plasmid buffer sol-(Exosurf) on gene expression from naked plasmid DNA ution (IC 50 = 8.6%). CAT expression was not altered when administered endobronchially to adult mice. Transfection surfactant was administered either 2 h before or after plasefficiency was evaluated by quantifying the expression of mid DNA instillation. Examination of the components of chloramphenicol acetyltransferase (CAT) and luciferase Exosurf revealed that two compounds, DPPC and tylox-(Luc) genes in the lung. Endobronchial administration of apol, showed inhibitory effects on CAT expression. Howeither CAT or Luc expression plasmid DNA resulted in ever, the inhibition caused by Exosurf appeared greater detectable concentrations of each reporter protein. CAT than that of either component. Our results suggest that the expression from plasmid DNA was monitored after endolung surfactant is a barrier to transfection of the endobronchial administration with the maximal expression bronchial airway and may be partly responsible for the low observed at 3-5 days after administration and decreasing expression of exogenous DNA in vivo in the bronchial tree.
Direct three-dimensional image transmission through one single-mode or multimode fiber is demonstrated. Image transmission is carried out with a grating interferometer under monochromatic, spatially incoherent illumination.
The use of a grating interferometer under spatially incoherent illumination for direct three-dimensional image transmission through optical fibers is analyzed. The issues of resolution, image depth, and signal-to-noise ratio are addressed. Experimental results are presented.
SummarySwitch-like activation of the Spindle Assembly Checkpoint (SAC) is critical for accurate chromosome segregation during cell division. To determine the mechanisms that implement it, we engineered an ectopic, kinetochore-independent SAC activator, the "eSAC". The eSAC stimulates the SAC signaling cascade by artificially dimerizing the Mps1 kinase domain and a cytosolic KNL1 phosphodomain, the signaling scaffold in the kinetochore. Quantitative analyses and mathematical modeling of the eSAC reveal that the recruitment of multiple SAC proteins by the KNL1 phosphodomain stimulates synergistic signaling, which enables a small number of KNL1 molecules produce a disproportionately strong anaphase-inhibitory signal. However, when multiple KNL1 molecules signal concurrently, they compete for a limited cellular pool of SAC proteins. This frustrates synergistic signaling and modulates signal output. Together, these mechanisms institute automatic gain control -inverse, non-linear scaling between the signal output per kinetochore and the unattached kinetochore number, and thus enact the SAC switch.
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