Nuclear Tumor Necrosis Factor Receptor-associated Factor 6 in Lymphoid Cells Negatively Regulates c-Myb-mediated Transactivation through Small Ubiquitin-related Modifier-1 Modification

Lan V Pham,Hai-Jun Zhou,Yen-Chiu Lin-Lee,Archito T Tamayo,Linda C Yoshimura,Lingchen Fu,Bryant G Darnay,Richard J Ford

doi:10.1074/jbc.m706307200

Abstract

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is an adaptor/scaffold protein that mediates several important signaling pathways, including the tumor necrosis factor-R:NF-kappaB pathway, involved in immune surveillance, inflammation, etc. Because most studies of TRAF6 function have focused primarily on its role as an adaptor molecule in signaling pathways in the cytoplasm, the potential functions of TRAF6 in other cellular compartments has not been previously investigated. Here, we demonstrate that TRAF6 resides not only in the cellular cytoplasm but is also found in the nuclei of both normal and malignant B lymphocytes. TRAF6 does not possess a nuclear localization signal but enters the nucleus through the nuclear pore complex containing RanGap1. Chromatin immunoprecipitation cloning experiments demonstrated that nuclear TRAF6 associates with c-Myb within the 5'-end of the c-Myb promoter. Further analysis showed that nuclear TRAF6 is modified by small ubiquitin-related modifier-1, interacts with histone deacetylase 1, and represses c-Myb-mediated transactivation. Thus, TRAF6 negatively regulates c-Myb through a novel repressor function in the nuclei of both normal and malignant B-lymphocytes that could represent a novel control mechanism that maintains cell homeostasis and immune surveillance.

Highlights

B-cell growth and survival [1, 2]
tumor necrosis factor (TNF) receptor engagement results in the assembly of a cascade of signaling molecules composed of critical proteins, including adaptor molecules such as TNF receptor-associated factor 6 (TRAF6) that are recruited to the cell membrane and into lipid raft microdomains [3]
Identification of Nuclear TRAF6 in Lymphoma B Cells—While studying the nuclear role of CD40 in lymphoma B cells, we identified the presence of TRAF6 protein in the nuclei of cells from five different aggressive B-cell lymphoma cell lines by nuclear cell fractionation and Western blotting (Fig. 1A)

Summary

Introduction

B-cell growth and survival [1, 2]. TNF receptor engagement results in the assembly of a cascade of signaling molecules composed of critical proteins, including adaptor molecules such as TNF receptor-associated factor 6 (TRAF6) that are recruited to the cell membrane and into lipid raft microdomains [3]. In vivo sumoylation assays were performed by transfecting lymphoma cells with FLAG-TRAF6 and HA-SUMO-1 expression vectors as indicated.

Results

Conclusion