Aoki, K. et al. Extracellular matrix interacts with soluble CD95L: retention and enhancement of cytotoxicity. Nat. Immunol. 2, 333-337
Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 40 Convent Drive, Bethesda, MD 20892-3005, USA. Nature Immunology
(Impact Factor: 20).
05/2001; 2(4):333-7. DOI: 10.1038/86336
Fas ligand (CD95L) is synthesized both on the cell surface membrane and in a soluble form. Although CD95L contributes to immune privilege in the cornea and testis, the functions of these alternatively processed proteins are not well understood. Some reports suggest that the cytotoxicity of soluble CD95L is insignificant, whereas others show potent responses in vivo, including hepatocyte apoptosis that causes liver failure. We show here that extracellular matrix proteins interact with soluble CD95L and potentiate its pro-apoptotic activity. The cytotoxicity of supernatants from CD95L-expressing cells was increased by incubation on tissue culture plates coated with these matrix proteins; this effect was mediated by trimeric soluble CD95L. With the use of immunoprecipitation, it was found that CD95L binds directly to fibronectin. In addition, immunohistochemical analysis of the cornea revealed that soluble CD95L binds primarily to extracellular matrix. The retention of soluble CD95L on extracellular matrices is likely to play an important role in the development of peripheral tolerance in immune-privileged sites.
Available from: Taiyang Ma
- "It has also been reported that naturally cleaved FasL may dampen apoptotic activity by competing with mFasL or blocking Fas receptors (Gregory et al., 2011; Knox et al., 2003; O' Reilly et al., 2009; Suda et al., 1996; Tanaka et al., 1998). When binding to the matrix proteins or cross-linking by antibodies, the cytotoxicity of naturally cleaved soluble FasL was potentiated in vitro and in vivo (Aoki et al., 2001; Schneider et al., 1998). Recombinant soluble FasL corresponding to the entire extracellular domain of FasL shows considerable cytotoxicity compared with naturally cleaved sFasL, although lower than membrane-bound FasL (Hohlbaum et al., 2000; Suda et al., 1996). "
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ABSTRACT: FasL is the most extensively studied apoptosis ligand. In 2000, tilapia FasL was identified using anti-human FasL monoclonal antibody by Evans's research group. Recently, a tilapia FasL-like protein of smaller molecule weight was predicted in Genbank (XM_003445156.2). Based on several clues drawn from previous studies, we cast doubt on the authenticity of the formerly identified tilapia FasL. Conversely, using reverse transcription polymerase chain reaction (RT-PCR), the existence of the predicted FasL-like was verified at the mRNA level. Through multiple alignments, this FasL-like protein was found to be highly similar to the FasL of the Japanese flounder. Moreover, we artificially expressed the functional region of the predicted protein and later confirmed its apoptosis-inducing activity using a methyl thiazolyl tetrazolium (MTT) assay, Annexin-V / Propidium iodide (PI) double staining, and DNA fragment detection. Supported by these evidences, we suggest that the predicted protein is the authentic tilapia FasL. To advance this research further, tilapia FasL mRNA and its protein across different tissues were quantified. High expression levels were identified in the tilapia immune system and sites where active cell turnover conservatively occurs. In this regard, FasL may assume an active role in the immune system and cell homeostasis maintenance in tilapia, similar to that shown in other species. In addition, because the distribution pattern of FasL mRNA did not synchronize with that of the protein, post-transcriptional expression regulation is suggested. Such regulation may be dominated by potential adenylate- and uridylate-rich elements (AREs) featuring AUUUA repeats found in the 3' untranslated region (UTR) of tilapia FasL mRNA.
Available from: Andrea Giani
- "This is in contrast to an experimental form of FasL that corresponds to the entire extracellular domain , . On the other hand, sFasL bound to extracellular matrix proteins is cytotoxic and FasL has been localized to the extracellular matrix in the anterior chamber of the eye . Thus, whether FasL accumulates in the ocular environment as full-length mFasL or truncated sFasL, matrix-associated or not, could influence its functional consequences. "
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ABSTRACT: Glaucoma, the most frequent optic neuropathy, is a leading cause of blindness worldwide. Death of retinal ganglion cells (RGCs) occurs in all forms of glaucoma and accounts for the loss of vision, however the molecular mechanisms that cause RGC loss remain unclear. The pro-apoptotic molecule, Fas ligand, is a transmembrane protein that can be cleaved from the cell surface by metalloproteinases to release a soluble protein with antagonistic activity. Previous studies documented that constitutive ocular expression of FasL maintained immune privilege and prevented neoangeogenesis. We now show that FasL also plays a major role in retinal neurotoxicity. Importantly, in both TNFα triggered RGC death and a spontaneous model of glaucoma, gene-targeted mice that express only full-length FasL exhibit accelerated RGC death. By contrast, FasL-deficiency, or administration of soluble FasL, protected RGCs from cell death. These data identify membrane-bound FasL as a critical effector molecule and potential therapeutic target in glaucoma.
- "Any reconsideration of CD95 as a therapeutic target therefore first of all has to account for and solve the problem of unwanted toxic side effects. Although the soluble, trimeric CD95L exerts little bioactivity per se, it has been observed that, when bound to the extracellular matrix, it is bioactive . Thus, the latent CD95-stimulating capability of soluble CD95L becomes apparent after immobilization. "
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ABSTRACT: Due to their strong apoptosis-inducing capacity, the death receptor ligands CD95L, TNF and TRAIL have been widely viewed as potential cancer therapeutics. While clinical data with CD95L and TRAIL are not yet available, TNF is a registered drug, albeit only for loco-regional application in a limited number of indications. The TNF experience has told us that specific delivery and restricted action is a major challenge in the development of multifunctional, pleiotropically acting cytokines into effective cancer therapeutics. Thus, gene-therapeutic approaches and new cytokine variants have been designed over the last 10 years with the aim of increasing anti-tumoral activity and reducing systemic side effects. Here, we present our current view of the therapeutic potential of the death receptor ligands TNF, CD95L and TRAIL and of the progress made towards improving their efficacy by tumor targeting, use of gene therapy and genetic engineering. Results generated with newly designed fusion proteins suggest that enhanced tumor-directed activity and prevention of undesirable actions of death receptor ligands is possible, thereby opening up a useful therapeutic window for all of the death receptor ligands, including CD95L.
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