Stabilized beta-catenin potentiates Fas-mediated T cell apoptosis

In response to Ag stimulation, Ag-specific T cells proliferate and accumulate in the peripheral lymphoid tissues. To avoid excessive T cell accumulation, the immune system has developed mechanisms to delete clonally expanded T cells. Fas/FasL-mediated apoptosis plays a critical role in the deletion of activated peripheral T cells, which is clearly demonstrated by superantigen (staphylococcal enterotoxin B)-induced deletion of Vbeta8(+) T cells. Using transgenic mice expressing a stabilized beta-catenin (beta-cat(Tg)), we show here that beta-catenin was able to enhance apoptosis of activated T cells by up-regulating Fas. In response to staphylococcal enterotoxin B stimulation, beta-cat(Tg) mice exhibited accelerated deletion of CD4(+)Vbeta8(+) T cells compared with wild type mice. Surface Fas levels were significantly higher on activated T cells obtained from beta-cat(Tg) mice than that from wild type mice. Additionally, T cells from beta-cat(Tg) mice were more sensitive to apoptosis induced by crosslinking Fas, activation-induced cell death, and to apoptosis induced by cytokine withdrawal. Lastly, beta-catenin bound to and stimulated the Fas promoter. Therefore, our data demonstrated that the beta-catenin pathway was able to promote the apoptosis of activated T cells in part via up-regulation of Fas.     

T cell-specific adapter protein inhibits T cell activation by modulating Lck activity

We previously reported the isolation of a cDNA encoding a T cell-specific adapter protein (TSAd). Its amino acid sequence contains an SH2 domain, tyrosines in protein binding motifs, and proline-rich regions. In this report we show that expression of TSAd is induced in normal peripheral blood T cells stimulated with anti-CD3 mAbs or anti-CD3 plus anti-CD28 mAbs. Overexpression of TSAd in Jurkat T cells interfered with TCR-mediated signaling by down-modulating anti-CD3/PMA-induced IL-2 promoter activity and anti-CD3 induced Ca2+ mobilization. The TCR-induced tyrosine phosphorylation of phospholipase C-gamma1, SH2-domain-containing leukocyte-specific phosphoprotein of 76kDa, and linker for activation of T cells was also reduced. Furthermore, TSAd inhibited Zap-70 recruitment to the CD3zeta-chains in a dose-dependent manner. Consistent with this, Lck kinase activity was reduced 3- to 4-fold in COS-7 cells transfected with both TSAd and Lck, indicating a regulatory effect of TSAd on Lck. In conclusion, our data strongly suggest an inhibitory role for TSAd in proximal T cell activation.     

A cryptic frizzled module in cell surface collagen 18 inhibits Wnt/beta-catenin signaling

Collagens contain cryptic polypeptide modules that regulate major cell functions, such as cell proliferation or death. Collagen XVIII (C18) exists as three amino terminal end variants with specific amino terminal polypeptide modules. We investigated the function of the variant 3 of C18 (V3C18) containing a frizzled module (FZC18), which carries structural identity with the extracellular cysteine-rich domain of the frizzled receptors. We show that V3C18 is a cell surface heparan sulfate proteoglycan, its topology being mediated by the FZC18 module. V3C18 mRNA was expressed at low levels in 21 normal adult human tissues. Its expression was up-regulated in fibrogenesis and in small well-differentiated liver tumors, but decreased in advanced human liver cancers. Low FZC18 immunostaining in liver cancer nodules correlated with markers of high Wnt/beta-catenin activity. V3C18 (M(r) = 170 kD) was proteolytically processed into a cell surface FZC18-containing 50 kD glycoprotein precursor that bound Wnt3a in vitro through FZC18 and suppressed Wnt3a-induced stabilization of beta-catenin. Ectopic expression of either FZC18 (35 kD) or its 50 kD precursor inhibited Wnt/beta-catenin signaling in colorectal and liver cancer cell lines, thus downregulating major cell cycle checkpoint gatekeepers cyclin D1 and c-myc and reducing tumor cell growth. By contrast, full-length V3C18 was unable to inhibit Wnt signaling. In summary, we identified a cell-surface signaling pathway whereby FZC18 inhibits Wnt/beta-catenin signaling. The signal, encrypted within cell-surface C18, is released by enzymatic processing as an active frizzled cysteine-rich domain (CRD) that reduces cancer cell growth. Thus, extracellular matrix controls Wnt signaling through a collagen-embedded CRD behaving as a cell-surface sensor of proteolysis, conveying feedback cues to control cancer cell fate.     

Heparin inhibits enhancing factor/phospholipase A2 activity and its binding to the cell surface

Enhancing factor (EF), a mouse intestinal phospholipase A₂ (PLA₂), has been isolated and characterized. EF increases the binding of epidermal growth factor (EGF) to A431 cells almost two-fold by interacting with EGF. EF binds to a 100 kDa cell surface receptor and brings about an increase in the binding of EGF. In the present study we demonstrate that EF is a heparin binding protein and at the time of iodination of EF, the heparin binding site of EF has to be protected. Heparin inhibits the enhancing activity of EF as well as the binding of labelled EF to A431 cells. Inhibition of binding of EF to cells by heparin indicates that heparin binding region forms at least part of the receptor binding domain. These data suggest that the receptor for EF on the cell surface could be a heparin sulphate proteoglycan.     

RBM47/SNHG5/FOXO3 axis activates autophagy and inhibits cell proliferation in papillary thyroid carcinoma

Papillary thyroid carcinoma (PTC) is the main type of thyroid carcinoma. Despite the good prognosis, some PTC patients may deteriorate into more aggressive diseases, leading to poor survival. Molecular technology has been increasingly used in the diagnosis and treatment of thyroid carcinoma. In this study, we identified that RNA Binding Motif Protein 47 (RBM47) was downregulated in PTC tissues and cells, and overexpression of RBM47 could activate autophagy and inhibit proliferation in PTC cells. RBM47 promotes but can not bind directly to Forkhead Box O3 (FOXO3). FOXO3 activates Autophagy Related Gene 3 (ATG3), ATG5, and RBM47 to form a loop and promote autophagy. RBM47 can bind directly to and stabilized lncRNA Small Nucleolar RNA Host Gene 5 (SNHG5) to inhibit PTC cells proliferation and activate autophagy in vitro and in vivo. SNHG5 inhibits ubiquitination and degradation of FOXO3 by recruiting Ubiquitin Specific Peptidase 21 (USP21), then promotes the translocation of FOXO3 from cytoplasm to nucleus. Our study revealed the regulatory mechanism of RBM47/SNHG5/FOXO3 axis on cell proliferation and autophagy in PTC, which may provide valuable insight for the treatment of PTC.Subject terms: Oncogenes, Head and neck cancer     

Transcriptional regulation of CD4 gene expression by T cell factor-1/beta-catenin pathway

By interacting with MHC class II molecules, CD4 facilitates lineage development as well as activation of Th cells. Expression of physiological levels of CD4 requires a proximal CD4 enhancer to stimulate basic CD4 promoter activity. T cell factor (TCF)-1/beta-catenin pathway has previously been shown to regulate thymocyte survival via up-regulating antiapoptotic molecule Bcl-xL. By both loss and gain of function studies, in this study we show additional function of TCF-1/beta-catenin pathway in the regulation of CD4 expression in vivo. Mice deficient in TCF-1 displayed significantly reduced protein and mRNA levels of CD4 in CD4+ CD8+ double-positive (DP) thymocytes. A transgene encoding Bcl-2 restored survival but not CD4 levels of TCF-1(-/-) DP cells. Thus, TCF-1-regulated survival and CD4 expression are two separate events. In contrast, CD4 levels were restored on DP TCF-1(-/-) cells by transgenic expression of a wild-type TCF-1, but not a truncated TCF-1 that lacks a domain required for interacting with beta-catenin. Furthermore, forced expression of a stabilized beta-catenin, a coactivator of TCF-1, resulted in up-regulation of CD4. TCF-1 or stabilized beta-catenin greatly stimulated activity of a CD4 reporter gene driven by a basic CD4 promoter and the CD4 enhancer. However, mutation of a potential TCF binding site located within the enhancer abrogated TCF-1 and beta-catenin-mediated activation of CD4 reporter. Finally, recruitment of TCF-1 to CD4 enhancer was detected in wild-type but not TCF-1 null mice by chromatin-immunoprecipitation analysis. Thus, our results demonstrated that TCF/beta-catenin pathway enhances CD4 expression in vivo by recruiting TCF-1 to stimulate CD4 enhancer activity.     

Protein-tyrosine phosphatase PCP-2 inhibits beta-catenin signaling and increases E-cadherin-dependent cell adhesion

beta-Catenin is a key molecule involved in both cell adhesion and Wnt signaling pathway. However, the exact relationship between these two roles has not been clearly elucidated. Tyrosine phosphorylation of beta-catenin was shown to decrease its binding to E-cadherin, leading to decreased cell adhesion and increased beta-catenin signaling. We have previously shown that receptor-like protein-tyrosine phosphatase PCP-2 localizes to the adherens junctions and directly binds and dephosphorylates beta-catenin, suggesting that PCP-2 might regulate the balance between signaling and adhesive beta-catenin. Here we demonstrate that PCP-2 can inhibit both the wild-type and constitutively active forms of beta-catenin in activating target genes such as c-myc. The phosphatase activity of PCP-2 is required for this effect since loss of catalytic activity attenuates its inhibitory effect on beta-catenin activation. Expression of PCP-2 in SW480 colon cancer cells can lead to stabilization of cytosolic pools of beta-catenin perhaps, by virtue of their physical interaction. PCP-2 expression also leads to increased membrane-bound E-cadherin and greater stabilization of adherens junctions by dephosphorylation of beta-catenin, which could further sequester cytosolic beta-catenin and thus inhibit beta-catenin mediated nuclear signaling. Furthermore, SW480 cells stably expressing PCP-2 have a reduced ability to proliferate and migrate. Thus, PCP-2 may play an important role in the maintenance of epithelial integrity, and a loss of its regulatory function may be an alternative mechanism for activating beta-catenin signaling.     

HIV Nef inhibits T cell migration

Nef is a viral regulatory protein of the human immunodeficiency virus (HIV) that has been shown to contribute to disease progression. Among its putative effects on T cell functions are the down-regulation of CD4 and major histocompatibility class I surface molecules. These effects occur in part via Nef interactions with intracellular signaling molecules. We sought to better characterize the effects of HIV Nef on T cell function by examining chemotaxis in response to stromal cell-derived factor-1alpha (SDF-1alpha) as well as CXCR4 signaling molecules. Here, we report the novel observation that HIV Nef inhibited chemotaxis in response to SDF-1alpha in both Jurkat T cells and primary peripheral CD4+ T lymphocytes. Our data indicate that HIV Nef altered critical downstream molecules in the CXCR4 pathway, including focal adhesion kinases. These findings suggest that HIV Nef may blunt the T cell response to chemokines. Because T lymphocyte migration is an integral component of host defense, HIV Nef may thereby contribute to the pathogenesis of AIDS.     

Detection in a murine T lymphoma of a lipid-like factor that inhibits cell growth

In this paper we show that MCG3 cells, a murine T lymphoma, contain a factor(s) that inhibits the proliferation of cells of different histological origin. The lack of sensitivity of this cell proliferation-inhibiting factor (CPIF) to the treatment with proteolytic enzymes and its solubility in organic solvent demonstrated that it is a lipid-like substance. Separation by thin-layer chromatography showed it migrates before the prostaglandins with activity on cell proliferation. CPIF activity was reversible and more intense on bone marrow cells than on tumor cells, suggesting that it can play a role in cell growth regulation.