Structure-function analysis of NADE: identification of regions that mediate nerve growth factor-induced apoptosis.
Nerve growth factor (NGF) can induce apoptosis in neural cells via activation of the low affinity neurotrophin receptor p75NTR. NADE (p75NTR-associated cell death executor) is a p75NTR-associated protein that mediates apoptosis in response to NGF by interacting with the death domain of p75NTR in 293T, PC12, and nnr5 cells (Mukai, J., Hachiya, T., Shoji-Hoshino, S., Kimura, M. T., Nadano, D., Suvanto, P., Hanaoka, T., Li, Y., Irie, S., Greene, L. A., and Sato, T. A. (2000) J. Biol. Chem. 275, 17566-17570). We performed extensive mutational analysis on NADE, to better characterize its structural and functional features. Truncation of a minimal region, including amino acid residues 41-71 of NADE, was found to be sufficient to induce apoptosis. The designated regulatory region includes the C-terminal amino acid residues (72-112) and is essential for NGF-dependent regulation of NADE-induced apoptosis. Furthermore, the mutants with amino acid substitutions in the leucine-rich nuclear export signal (NES) sequence (residues 90-100) abolished the export of NADE from the nucleus to the cytoplasm. Mutation of the NES also abolished self-association of NADE, its interaction with p75NTR, and NGF-dependent apoptosis. Expression of a fragment of NADE (amino acid residues 81-124) blocked NGF-induced apoptosis in oligodendrocytes, suggesting that this region has a dominant negative effect on NGF/p75NTR-induced apoptosis. These studies identify distinct regions of NADE that are involved in regulating specific functions involved in p75NTR signal transduction.
Mukai
Jun
J
Division of Molecular Oncology, Department of Otolaryngology/Head & Neck Surgery and Pathology, College of Physicians & Surgeons, Columbia University, New York, New York 10032, USA.
Shoji
Shisako
S
Kimura
Makoto T
MT
Okubo
Shuichi
S
Sano
Hajime
H
Suvanto
Petro
P
Li
Yin
Y
Irie
Shinji
S
Sato
Taka-Aki
TA
D000256:Adenoviridae / Q000235:genetics; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D002460:Cell Line; D004252:DNA Mutational Analysis; D018014:Gene Transfer Techniques; D005799:Genes, Dominant; D049452:Green Fluorescent Proteins; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D007930:Leucine / Q000737:chemistry; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D016716:PC12 Cells; D017354:Point Mutation; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry* / Q000502:physiology*; D051381:Rats; D020800:Receptor, Nerve Growth Factor; D017475:Receptors, Nerve Growth Factor / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection; D020798:Two-Hybrid System Techniques
D051017:Apoptosis Regulatory Proteins; C410727:BEX3 protein, human; C485843:Bex3 protein, rat; D008164:Luminescent Proteins; C485842:Ngfrap1 protein, mouse; D011506:Proteins; D020800:Receptor, Nerve Growth Factor; D017475:Receptors, Nerve Growth Factor; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D007930:Leucine