PP1 phosphatase is involved in Bcl-2 dephosphorylation after prolonged mitotic arrest induced by paclitaxel.
During mitotic arrest induced by paclitaxel, most of the mitochondrial Bcl-2 is phosphorylated. This mitotic arrest is transient; exit from mitosis, due to mitotic slippage, occurs and Bcl-2 is rapidly dephosphorylated. In the present study, we characterized PP1 as the cytosolic phosphatase involved in Bcl-2 dephosphorylation. When mitochondria and cytosol prepared from mitotic arrested cells were incubated in vitro, the proportion of phosphorylated forms of Bcl-2 in mitochondria remained unchanged. In contrast, cytosol prepared from cells during mitotic slippage led to a dose-dependent loss of phosphorylated forms of Bcl-2. Depletion of these cytosol extracts by microcystin-Sepharose maintained Bcl-2 phosphorylated forms, indicating that this cytosol possessed phosphatase activity. Furthermore, the dephosphorylation of Bcl-2 by cytosol prepared from cells exiting mitotic block was inhibited by okadaic acid, at a dose known to inhibit PP1, and by inhibitor 2, a specific inhibitor of PP1 and by immunodepletion of PP1. Finally, we showed that PP1 is associated with mitochondrial Bcl-2 in vivo. Taken together, these results demonstrate that PP1 is directly involved in Bcl-2 dephosphorylation during mitotic slippage.
Brichese
Laetitia
L
LBCMCP, UMR CNRS 5088, Université Paul Sabatier, 31 062 Toulouse Cedex, France.
Valette
Annie
A
D000970:Antineoplastic Agents / Q000494:pharmacology; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D003600:Cytosol / Q000737:chemistry / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006801:Humans; D052998:Microcystins; D008928:Mitochondria / Q000737:chemistry / Q000378:metabolism; D008938:Mitosis / Q000187:drug effects; D017239:Paclitaxel / Q000494:pharmacology*; D010456:Peptides, Cyclic / Q000737:chemistry / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011233:Precipitin Tests; D011485:Protein Binding; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D012685:Sepharose / Q000737:chemistry; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D052998:Microcystins; D010456:Peptides, Cyclic; D019253:Proto-Oncogene Proteins c-bcl-2; C078588:microcystin; D012685:Sepharose; D010749:Phosphoprotein Phosphatases; D017239:Paclitaxel