Genotoxic stress-induced nuclear localization of oncoprotein YB-1 in the absence of proteolytic processing.
Y-box-binding protein 1 (YB-1) is an oncogenic transcription factor whose overexpression and nuclear localization is associated with tumor progression and drug resistance. Transcriptional activation of YB-1 in response to genotoxic stress is believed to occur in the cytoplasm through sequence-specific endoproteolytic cleavage by the 20S Proteasome, followed by nuclear translocation of cleaved YB-1. To study the proteolysis model, we developed a two-step affinity purification of endogenous YB-1 protein species and characterized the products using mass spectrometry. Whereas full-length YB-1 was readily identified, the smaller protein band thought to be activated YB-1 was identified as hnRNP A1. An antibody specific for YB-1 was generated, which revealed only one YB-1 species, even after genotoxic stress-induced nuclear YB-1 translocation. These findings warrant re-evaluation of the mechanism of YB-1 nuclear translocation and transcriptional activation. The relationship between nuclear YB-1 and tumor progression may also have to re-evaluated in some cases.
Cohen
S B
SB
Cell Transformation Unit, Children's Medical Research Institute, Westmead, New South Wales, 2145, Australia.
Ma
W
W
Valova
V A
VA
Algie
M
M
Harfoot
R
R
Woolley
A G
AG
Robinson
P J
PJ
Braithwaite
A W
AW
D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000528:radiation effects; D000970:Antineoplastic Agents / Q000494:pharmacology; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism*; D002853:Chromatography, Liquid; D002945:Cisplatin / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism*; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003609:Dactinomycin / Q000494:pharmacology; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D021381:Protein Transport / Q000187:drug effects / Q000528:radiation effects; D034622:RNA Interference; D053719:Tandem Mass Spectrometry; D015533:Transcriptional Activation / Q000187:drug effects / Q000528:radiation effects; D014466:Ultraviolet Rays; D051840:Y-Box-Binding Protein 1
D000970:Antineoplastic Agents; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D051840:Y-Box-Binding Protein 1; C070856:YBX1 protein, human; D003609:Dactinomycin; D002945:Cisplatin