benzyloxycarbonylleucyl-leucyl-leucine-aldehyde and 1-7-diaminoheptane

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.

Topics: Acetylcysteine; Apoptosis; Caspase 3; Caspases; Cysteine Endopeptidases; Diamines; Electrophoresis, Gel, Two-Dimensional; Enzyme Activation; Eukaryotic Translation Initiation Factor 5A; Gene Expression Profiling; Gene Expression Regulation; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Leukemia, Megakaryoblastic, Acute; Leupeptins; Lysine; Multienzyme Complexes; Oligodeoxyribonucleotides, Antisense; Peptide Initiation Factors; Peptide Mapping; Protease Inhibitors; Proteasome Endopeptidase Complex; Proteomics; RNA-Binding Proteins; Spectrometry, Mass, Electrospray Ionization; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Subcellular Fractions; Tumor Cells, Cultured; Ubiquitin