NAK Is Recruited to the TNFR1 Complex in a TNFα-dependent Manner and Mediates the Production of RANTES: IDENTIFICATION OF ENDOGENOUS TNFR-INTERACTING PROTEINS BY A PROTEOMIC APPROACH

Jun Kuai,Joe Wooters,J Perry Hall,Vikram R Rao,Elliott Nickbarg,Bilian Li,Moitreyee Chatterjee-Kishore,Yongchang Qiu,Lih-Ling Lin

doi:10.1074/jbc.m411037200

Abstract

Tumor necrosis factor alpha (TNFalpha) is a proinflammatory cytokine with pleiotropic immunological and biological activities. TNFalpha signaling is triggered by the engagement of soluble TNFalpha to two types of cell surface receptors, TNFR1 and TNFR2. This recruits cytosolic proteins to the intracellular domains of the receptors and initiates signaling to downstream effectors. In this study, we used a proteomic approach to identify these cytosolic proteins from affinity-purified, endogenous TNFalpha.TNFR complexes in human myelomonocytic U937 cells. Seven proteins were identified, including TRADD, TRAP2, and TRAF2, which are three proteins known to be recruited to TNFalpha receptors. NAK, RasGAP3, TRCP1, and TRCP2 were also identified. We further showed that NAK is recruited to TNFR1 in a temporally regulated and TNFalpha-dependent manner and that it mediates the TNFalpha-induced production of the chemokine RANTES (regulated on activation normal T cell expressed and secreted). These data demonstrate that NAK is a component of the TNFalpha.TNFR1 signaling complex and confirm the physiological role of NAK in the TNFalpha-mediated response.

Highlights

Tumor necrosis factor ␣ (TNF␣) is a proinflammatory cytokine with pleiotropic immunological and biological activities
We further showed that NAK is recruited to TNFR1 in a temporally regulated and TNF␣-dependent manner and that it mediates the TNF␣-induced production of the chemokine RANTES
We demonstrated that recruitment of NAK to TNFR1 is temporally regulated and TNF␣-dependent and that endogenous NAK is required for the TNF␣-induced production of the chemokine RANTES

Summary

Introduction

Tumor necrosis factor ␣ (TNF␣) is a proinflammatory cytokine with pleiotropic immunological and biological activities. We used a proteomic approach to identify these cytosolic proteins from affinity-purified, endogenous TNF␣1⁄7TNFR complexes in human myelomonocytic U937 cells. Seven proteins were identified, including TRADD, TRAP2, and TRAF2, which are three proteins known to be recruited to TNF␣ receptors.

Results

Conclusion