Proteomic analysis of ubiquitin-proteasome effects: insight into the function of eukaryotic initiation factor 5A.
The global effect of ubiquitin-proteasome (UP) inhibitors on leukemic cell proteome was analysed. A total of 39 protein spots, affected by UP inhibitors, were identified, including 11 new apoptosis-associated proteins. They are involved in different cellular functions and four were associated with caspase-3 activation. Eukaryotic initiation factor 5A (eIF-5A) was identified in two spots; however, the peptide mass-fingerprinting for the accumulated one included a peptide with lysine50, indicating that hypusine formation was suppressed during UP inhibitor-induced apoptosis. Hypusine modification ensues immediately following translation of eIF-5A precursor, unless cells are treated with the modification inhibitors diaminoheptane. However, UP inhibitors induced a much stronger accumulation of unmodified eIF-5A compared to the effect of diaminoheptane. We further showed the unmodified eIF-5A was regulated in a proteasome-dependent manner. Inhibition of hypusine formation by diaminoheptane triggered apoptosis, but of particular interest is the finding that eIF-5A expression inhibition by antisense oligodeoxynucleotides significantly enhanced the stimulating effect of GM-CSF on cell growth. Therefore, the eIF-5A accumulation played important roles in the apoptosis induced by UP inhibitors. Moreover, hypusine inhibition in apoptosis was further revealed to be associated with the subcellular localization of eIF-5A. Our data pave the way to a better understanding of the mechanisms by which UP system has been linked to apoptosis.
Jin
Bao-Feng
BF
Institute of Basic Medical Sciences, National Center of Biomedical Analysis, 27 Tai-Ping Road, Beijing 100850, China.
He
Kun
K
Wang
Hong-Xia
HX
Wang
Jie
J
Zhou
Tao
T
Lan
Yu
Y
Hu
Mei-Ru
MR
Wei
Kai-Hua
KH
Yang
Song-Cheng
SC
Shen
Bei-Fen
BF
Zhang
Xue-Min
XM
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000502:physiology*; D003959:Diamines / Q000494:pharmacology; D015180:Electrophoresis, Gel, Two-Dimensional; D004789:Enzyme Activation; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D007947:Leukemia, Megakaryoblastic, Acute / Q000473:pathology; D007976:Leupeptins / Q000494:pharmacology*; D008239:Lysine / Q000031:analogs & derivatives* / Q000378:metabolism; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000502:physiology*; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology; D010448:Peptide Initiation Factors / Q000235:genetics / Q000502:physiology*; D010449:Peptide Mapping; D011480:Protease Inhibitors / Q000494:pharmacology*; D046988:Proteasome Endopeptidase Complex; D040901:Proteomics; D016601:RNA-Binding Proteins; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013347:Subcellular Fractions / Q000737:chemistry; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D025801:Ubiquitin / Q000502:physiology*; D000097574:Eukaryotic Translation Initiation Factor 5A
D000111:Acetylcysteine; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases; D003959:Diamines; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D007976:Leupeptins; D008239:Lysine; D009097:Multienzyme Complexes; D020319:Oligodeoxyribonucleotides, Antisense; D010448:Peptide Initiation Factors; D011480:Protease Inhibitors; D046988:Proteasome Endopeptidase Complex; D016601:RNA-Binding Proteins; D025801:Ubiquitin; D000097574:Eukaryotic Translation Initiation Factor 5A; C067713:lactacystin; C100028:hypusine; C015127:1,7-diaminoheptane; C505407:CASP3 protein, human; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde