Full-length, Membrane-anchored TWEAK Can Function as a Juxtacrine Signaling Molecule and Activate the NF-κB Pathway

Abstract

Tumor necrosis factor (TNF) family members are initially synthesized as type II transmembrane proteins, but some of these proteins are substrates for proteolytic enzymes that generate soluble cytokines with biological activity. TWEAK (TNFlike weak inducer of apoptosis), a member of the TNF family, is a multifunctional cytokine that acts via binding to a cell surface receptor named Fn14 (fibroblast growth factor-inducible 14). Studies conducted to date indicate that TWEAK-producing cells can co-express both membrane-anchored and soluble TWEAK isoforms, but there is little information on TWEAK proteolytic processing. Also, it is presently unclear whether membrane-anchored TWEAK, like soluble TWEAK, is biologically active. Here we show that full-length human TWEAK is processed intracellularly by the serine protease furin and identify TWEAK amino acid residues 90 -93 as the predominant furin recognition site. In addition, we report that full-length, membrane-anchored TWEAK can bind the Fn14 receptor on neighboring cells and activate the NF-B signaling pathway. Thus, TWEAK can act in a juxtacrine manner to initiate cellular responses, and this property may be important for TWEAK function during physiological wound repair and disease pathogenesis.TWEAK (tumor necrosis factor-like weak inducer of apoptosis) is a member of the TNF 2 superfamily of structurally related cytokines that together are involved in many critical biological processes, including development, organogenesis, tissue repair, and the innate and adaptive immune responses (1-3). TWEAK was initially described as a proapoptotic factor for certain tumor cell lines (4), but subsequent studies revealed that it can stimulate many other cellular responses, including cell proliferation, survival, and differentiation (5, 6). This cytokine acts on responsive cells via binding to the Fn14 (fibroblast growth factor-inducible 14) cell surface receptor, a 102-amino acid type I transmembrane protein originally described as a serum-and growth factor-inducible gene product in fibroblasts and smooth muscle cells (7-9). The TWEAK/Fn14 axis has been implicated in physiological wound repair and in the pathogenesis of ischemic stroke and several chronic inflammatory diseases (5, 6), including rheumatoid arthritis (10, 11) and inflammatory bowel disease (12).The human TWEAK gene encodes a 249-amino acid type II transmembrane protein, and when this full-length form of TWEAK was overexpressed in transfected HEK293-EBNA cells it was detected on the cell surface by FACS analysis, as expected (4). However, when a metabolic labeling and immunoprecipitation experiment was performed using the transfected cells, a smaller TWEAK form was found in conditioned medium (4). These results indicated that HEK293-EBNA cells could produce two TWEAK isoforms: a full-length, membraneanchored form and a smaller secreted form that is probably generated by TWEAK proteolytic processing. Subsequent studies have shown that other cell types can also co-express membrane-anchored and soluble TWEAK, indica...

Tumor Necrosis Factor-Like Weak Inducer of Apoptosis Induces Astrocyte Proliferation through the Activation of Transforming-Growth Factor-α/Epidermal Growth Factor Receptor Signaling Pathway

“…Unlike other studies in which various cell populations have been investigated (Marsters et al, 1998;Brown et al, 2003Brown et al, , 2010Dai et al, 2009;Kumar et al, 2009;Sanz et al, 2010;Moreno et al, 2011), TWEAK stimulation of primary astrocytes was not associated with NF-B but rather with ERK and p38 MAPK transduction pathway activation. This observation emphasizes the differences that exist between cell types in transducing signals from common receptors, thereby mediating distinct biological functions.…”

Structural Basis and Targeting of the Interaction between Fibroblast Growth Factor-inducible 14 and Tumor Necrosis Factor-like Weak Inducer of Apoptosis

“…The THD functions in ligand trimerization and receptor binding causing TWEAK to signal as a trimerized molecule (4,5). Importantly, both membrane-bound and -soluble TWEAK (sTWEAK) proteins are fully functional and can mediate similar cellular signaling effects by binding to cellular receptors (6).…”

A Homogeneous HTRF Assay for the Identification of Inhibitors of the TWEAK-Fn14 Protein Interaction

“…TWEAK forms homotrimers in solution 2 and mediates its biological activities mainly by binding to the Fn14 (fibroblast growth factor inducible 14) receptor, 3,4 although full-length, membrane-anchored TWEAK is also capable of signaling through Fn14. 5 Fn14 itself is a type I membrane protein with a cysteine-rich extracellular domain required for TWEAK binding while a small cytoplasmic tail contains one TNFR-associated factor (TRAF) binding domain. 6 Binding of TWEAK trimers induces trimerization also of the Fn14 receptor, followed by activation of the proinflammatory NF-κB pathway.…”