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Distinct adapter proteins mediate acid versus neutral sphingomyelinase activation through the p55 receptor for tumor
S Adam-Klages1, R Schwandner, D Adam
1Institut für Immunologie, Christian-Albrechts-Universität Kiel, Germany. sabine@immunologie.uni-kiel.de
Abstract:
Ceramide, generated by the enzymatic function of sphingomyelinases (SMases) has emerged as an important signaling pathway transducing diverse biological effects of various cytokine receptors. The 55-kDa receptor for tumor necrosis factor (TNF-R55) activates two types of SMases through distinct cytoplasmic domains. The death domain that is responsible for the initiation of the apoptotic pathway also signals for the activation of an acid SMase (A-SMase). The adapter protein TRADD binds to TNF-R55 in a ligand-dependent manner and serves as anchor for the subsequent recruitment of other proteins into the signaling complex that directly lead to cell death or nuclear factor-kappaB (NF-kappaB) induction. Notably, the two pro-apoptotic adapter proteins TRADD and FADD are also involved in the activation of A-SMase. In contrast, the NF-kappaB-inducing adapters TRAF2 and RIP do not signal for A-SMase. Thus, activation of A-SMase appears to belong to signals leading to TNF-induced cell death. A second signaling domain (NSD) is located upstream of the death domain and directly links the TNF-R55 to the activation of a neutral SMase (N-SMase). A novel adapter protein, FAN, has been identified that specifically binds to the NSD. FAN contains five WD repeats at its carboxy terminus, while it shows significant sequence homology with the mouse beige protein and its human homolog, the CHS protein, in the center portion of the protein. Overexpression of full-length FAN enhanced N-SMase activity in TNF-treated cells, whereas truncated mutants of FAN produced dominant negative effects. FAN, however, did not interfere with any of the TNF responses signaled for by the death domain. Taken together, our data suggest that distinct cytoplasmic domains of TNF-R55 initiate independent signaling pathways by binding different adapter proteins.
Insights
Tumor necrosis factor receptor 55 (TNF-R55) utilizes distinct domains to activate sphingomyelinases (SMases), influencing cell death and NF-kappaB pathways. A novel protein, FAN, specifically activates neutral SMase (N-SMase) via a separate domain.
Area of Science:
- Cellular Biology
- Molecular Signaling
- Biochemistry
Background:
- Ceramide, produced by sphingomyelinases (SMases), is a key signaling molecule involved in cytokine receptor pathways.
- The 55-kDa tumor necrosis factor receptor (TNF-R55) mediates diverse cellular responses through distinct cytoplasmic domains.
Purpose of the Study:
- To elucidate the distinct signaling mechanisms by which TNF-R55 activates different SMase isoforms.
- To identify adapter proteins involved in TNF-R55-mediated ceramide generation and downstream signaling.
Main Methods:
- Investigated TNF-R55 signaling complexes using ligand-dependent binding assays.
- Analyzed the role of adapter proteins TRADD, FADD, TRAF2, RIP, and FAN in SMase activation.
- Utilized overexpression and dominant-negative mutant approaches to assess FAN function.
Main Results:
- The TNF-R55 death domain recruits TRADD and FADD, activating acid SMase (A-SMase), linked to apoptosis.
- The non-death signaling domain (NSD) recruits FAN, activating neutral SMase (N-SMase).
- FAN overexpression enhances N-SMase activity, while mutants inhibit it, without affecting death domain signaling.
Conclusions:
- Distinct cytoplasmic domains of TNF-R55 initiate independent signaling pathways.
- Adapter proteins like TRADD, FADD, and FAN differentially regulate SMase activation and downstream cellular outcomes.
- TNF-R55 signaling involves compartmentalized activation of ceramide production pathways.