TRAF2 Phosphorylation Modulates Tumor Necrosis Factor Alpha-Induced Gene Expression and Cell Resistance to Apoptosis

Abstract

TRAF2 is an adaptor protein that regulates the activation of the c-Jun N-terminal kinase (JNK) and IB kinase (IKK) signaling cascades in response to tumor necrosis factor alpha (TNF-␣) stimulation. Although the downstream events in TNF-␣ signaling are better understood, the membrane-proximal events are still elusive. Here, we demonstrate that TNF-␣ and cellular stresses induce TRAF2 phosphorylation at serine 11 and that this phosphorylation is required for the expression of a subset of NF-B target genes. Although TRAF2 phosphorylation had a minimal effect on the TNF-␣-induced rapid and transient IKK activation, it was essential for secondary and prolonged IKK activation. Consistent with this, TRAF2 phosphorylation is not required for its recruitment to the TNFR1 complex in response to TNF-␣ stimulation but is required for its association with a cytoplasmic complex containing RIP1 and IKK. In addition, TRAF2 phosphorylation was essential for the full TNF-␣-induced activation of JNK. Notably, TRAF2 phosphorylation increased both basal and inducible c-Jun and NF-B activities and rendered cells resistant to stress-induced apoptosis. Moreover, TRAF2 was found to be constitutively phosphorylated in some lymphomas. These results unveil a new, finely tuned mechanism for TNF-␣-induced IKK activation modulated by TRAF2 phosphorylation and suggest that TRAF2 phosphorylation contributes to elevated levels of basal NF-B activity in certain human cancers.Tumor necrosis factor (TNF) receptor (TNFR)-associated factors (TRAFs) are characterized by the presence of a TRAF domain at the C terminus. Currently, six members of this family are known (TRAR1 to TRAR6), and all TRAFs, except TRAF1, contain N-terminal RING finger domains followed by five or seven zinc finger motifs (5, 31). TRAF2 is a prototypical member of the TRAF family and regulates signals from TNFR superfamily members.One of the well-characterized members of the TNFR superfamily is TNFR1. Currently, it is believed that TNFR1 activation by soluble TNF-␣ induces the sequential formation of two complexes with opposing effects on cell fate. In the first step, TNFR1 recruits the TNFR1-associated death domain (TRADD) protein, which in turn recruits TRAF2 and receptor-interacting protein 1 (RIP1) to form the membrane-bound prosurvival complex I. This leads to the sequential activation of mitogen-activated protein kinase (MAPK) kinase kinase (such as MEKK1/3), MAPK kinase (e.g., MKK4/7), and MAPK (e.g., c-Jun N-terminal kinase [JNK]) as well as in activation of transforming growth factor ␤-activated kinase 1 (TAK1), RIP1, and IB kinase (IKK). In the next step, the TRADD/ RIP1/TRAF2 complex dissociates from TNFR1 and associates with the Fas-associated death domain protein and caspase-8 to form the cytoplasmic proapoptotic complex II (23). The balance between these pathways is usually tipped by two events. One is the expression of cFLIP in response to NF-B activation by complex I; cFLIP then binds to complex II and inhibits apoptosis by interfering with caspase-8 activation. T...

IκB Kinase ε Phosphorylates TRAF2 To Promote Mammary Epithelial Cell Transformation

“…Recent work indicates that TRAF2 is phosphorylated at various serine residues important for NF-B activation (3,21,31,35). In particular, TRAF2 Ser11 phosphorylation has been previously reported, and this activity occurs constitutively in some cancer cell lines (3). Here, we demonstrate that IKKε phosphorylates TRAF2 both in vitro and in vivo.…”

“…TRAF2 is an adaptor molecule that mediates a series of ubiquitination events that facilitate NF-B activation (2) and is itself modified by ubiquitination (7). Recent work indicates that TRAF2 is phosphorylated at various serine residues important for NF-B activation (3,21,31,35). In particular, TRAF2 Ser11 phosphorylation has been previously reported, and this activity occurs constitutively in some cancer cell lines (3).…”

Two Coordinated Mechanisms Underlie Tumor Necrosis Factor Alpha-Induced Immediate and Delayed IκB Kinase Activation

“…The mean Ϯ standard error (SE) of four independent experiments was considered to be statistically significant at a P value of Ͻ0.05. For PCR analysis of mouse genes, the same primer pairs reported in a previous publication were used (27). For RT-PCR of human genes in Jurkat T cells, the following primer pairs were used: GAPDH, 5=-ACCACAGTCCATGCCATCAC-3= (forward) and 5=-TCCACCACCCTGTTGCTGTA-3=; IP-10, 5=-GCATCAG CATTAGTAATCAACC-3= (forward) and 5=-GCTCCCCTCTGGTTTTA AGGAG-3=.…”

Phosphorylation of Ribosomal Protein S3 and Antiapoptotic TRAF2 Protein Mediates Radioresistance in Non-small Cell Lung Cancer Cells

“…Moreover, inhibition of TRAF2 down-regulates G 2 -M phase control proteins, resulting in increased G 2 -M cell cycle arrest, growth inhibition, and radiosensitization (15). These regulatory effects of TRAF2 are mainly dependent on interactions with other proteins: TRADD, FAS-associated death domain protein (FADD), RIP1, TGF␤-associated kinase 1 (TAK1), TAK1-binding protein (TAB), NF-B-inducing kinase (NIK), A20, cIAPs, and TRAF5 (14,38). Thus, the functional complexity of TRAF2 signaling is considered to be associated with recruitment and the combination of the numerous TRAF2-interacting proteins.…”