| Research Abstract |
To clarify the role of Bcl-2 phosphorylation, we established the wild-type and several mutant-form of Bcl-2-expressing cell lines. We found that transmembrane region ?-deleted Bcl-2 (Bcl-2ΔTM), which was not localized at mitochondria, was constitutively phosphorylated in the cells. We identified Ser-87 residue within Bcl-2ΔTM protein as the phosphorylation site. To identify the responsible kinase for phosphorylation of Bcl-2ΔTM, we used several kinase inhibitors, and found that PD98059 and U0126, MEK1 inhibitors, suppressed phosphorylation of Bcl-2ΔTM. Moreover, in vitro kinase assay showed that ERK, a downstream kinase of MEK1, phosphorylated at Ser-87 residue of Bcl-2ΔTM. Both wild-type Bcl-2 and Bcl-2ΔTM could bind to ERK. Wild-type Bcl-2 could associate with PP2A, thus, wild-type Bcl-2 was not phosphorylated constitutively. On the other hand, Bcl-2ΔTM could interact with ERK, but not with PP2A. Moreover, Bcl-2 protein was constitutively phosphorylated in normal human blood cells, whereas Bcl-2 was not phosphorylated in human tumor cell lines.
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