A protein disulfide isomerase/thioredoxin-1 complex is physically attached to exofacial membrane TNF-receptors: overexpression in chronic lymphocytic leukemia

• Aims: The 3D structures and functions of cysteine-rich receptors such as tumor necrosis factor receptors (TNFRs) are redox-modulated by dithiol–disulfide exchange. TNFR superfamily members participate in growth regulation in B-cell chronic lymphocytic leukemia (CLL), and tissue stromal cells interact with leukemia cells, profoundly affecting their viability via release of redox-active components, including cysteine, thioredoxin-1 (Trx1), and Trx reductase. Trx1 was previously shown to enhance release of TNF, which acts as an autocrine/paracrine growth factor in CLL. The nature of the mechanism is not known, however. Here, we investigated whether Trx1 and protein disulfide isomerase (PDI), a chaperone and Trx-family member, may interact with TNFRs. Results: We found direct physical association between PDI and TNFR1 or TNFR2 by coclustering and affinity isolation. PDI (57 kDa) formed covalent/reduction-sensitive 69-kDa complexes with Trx1 (12 kDa) in a majority of CLL cell samples, detected at low levels only in control B-cells. Functionally, the TNF/TNFR signaling via the nuclear factor kappa B-driven autocrine loop was disrupted in a dose-dependent fashion by PDI-inhibitors bacitracin, anti-PDI, or anti-Trx1 antibodies, resulting in reduced viability. PDI was significantly overexpressed in immunoglobulin heavy-chain variable (IGHV) unmutated versus mutated CLL (p=0.0102), and amplified TNF release was observed in the former group. Innovation: This study points out a previously unrecognized physical and functional association of TNFRs with the redox-active proteins PDI and Trx1. Conclusion: We describe here a new level of TNF regulation, in which membrane TNFRs are redox controlled at the exofacial surface by PDI/Trx1. These findings shed new light on the observed survival benefit in CLL B-cells exerted by TNFR-superfamily ligands and point at potential therapeutic strategies

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