The N-terminal domains target TNF receptor-associated factor-2 to the nucleus and display transcriptional regulatory activity.
The subcellular localization of the TNF receptor-associated factor-2 (TRAF2) adaptor protein in human endothelial cells, which mediates proinflammatory responses of TNF, has been analyzed by confocal immunofluorescence microscopy and by Western blotting of fractionated cell extracts. Rabbit antisera reactive with either amino- or carboxyl-terminal TRAF2 peptides frequently but not uniformly stain nuclei of cultured HUVEC or the established human endothelial cell line, ECV304. However, Western blotting reveals significant heterogeneity in the reactivities of these polyclonal Abs. Transiently transfected HUVEC expressing FLAG epitope-tagged TRAF2 consistently show prominent nuclear localization, and deletion mutants of TRAF2 identify the portion of the molecule responsible for nuclear localization as the amino-terminal ring finger domain. TNF treatment does not appear to influence the localization of endogenous or transfected TRAF2 protein. Transfection of the amino-terminal half of the TRAF2 molecule, containing the ring and zinc finger domains, which localizes to the nucleus, results in activation of E-selectin but not of NF-kappaB promoter-reporter gene transcription or of c-Jun N-terminal kinase activation. These observations suggest that TRAF2 may reside in the nucleus and directly regulate transcription, independent of its role in cytoplasmic signal transduction.
Min
W
W
Boyer Center for Molecular Medicine, Yale University School of Medicine, New Haven, CT 06536, USA.
Bradley
J R
JR
Galbraith
J J
JJ
Jones
S J
SJ
Ledgerwood
E C
EC
Pober
J S
JS
D001692:Biological Transport / Q000235:genetics / Q000276:immunology; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000276:immunology / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D010446:Peptide Fragments / Q000235:genetics / Q000502:physiology; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism*; D047992:TNF Receptor-Associated Factor 2; D014158:Transcription, Genetic / Q000276:immunology*; D014162:Transfection / Q000276:immunology; D014471:Umbilical Veins
D010446:Peptide Fragments; D011506:Proteins; D018124:Receptors, Tumor Necrosis Factor; D047992:TNF Receptor-Associated Factor 2