Microtubule-targeting drugs induce bcl-2 phosphorylation and association with Pin1.
Bcl-2 is a critical suppressor of apoptosis that is overproduced in many types of cancer. Phosphorylation of the Bcl-2 protein is induced on serine residues in tumor cells arrested by microtubule-targeting drugs (paclitaxel, vincristine, nocodazole) and has been associated with inactivation of antiapoptotic function through an unknown mechanism. Comparison of a variety of pharmacological inhibitors of serine/threonine-specific protein kinases demonstrated that the cyclin-dependent kinase inhibitor, flavopiridol, selectively blocks Bcl-2 phosphorylation induced by antimicrotubule drugs. Bcl-2 could also be coimmunoprecipitated with the kinase Cdc2 in M-phase-arrested cells, suggesting that Cdc2 may be responsible for phosphorylation of Bcl-2 in cells treated with microtubule-targeting drugs. Examination of several serine-->alanine substitution mutants of Bcl-2 suggested that serine 70 and serine 87 represent major sites of Bcl-2 phosphorylation induced in response to microtubule-targeting drugs. Both these serines are within sequence contexts suitable for proline-directed kinases such as Cdc2. Phosphorylated Bcl-2 protein was discovered to associate in M-phase-arrested cells with Pin1, a mitotic peptidyl prolyl isomerase (PPIase) known to interact with substrates of Cdc2 during mitosis. In contrast, phosphorylation of Bcl-2 induced by microtubule-targeting drugs did not alter its ability to associate with Bcl-2 (homodimerization), Bax, BAG1, or other Bcl-2-binding proteins. Since the region in Bcl-2 containing serine 70 and serine 87 represents a proline-rich loop that has been associated with autorepression of its antiapoptotic activity, the discovery of Pin1 interactions with phosphorylated Bcl-2 raises the possibility that Pin1 alters the conformation of Bcl-2 and thereby modulates its function in cells arrested with antimicrotubule drugs.
Pathan
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The Burnham Institute, 10901 N. Torrey Pines Rd., La Jolla, CA 92037, USA.
Aime-Sempe
C
C
Kitada
S
S
Basu
A
A
Haldar
S
S
Reed
J C
JC
D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D016203:CDC2 Protein Kinase / Q000378:metabolism; D002460:Cell Line / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D019169:Jurkat Cells / Q000187:drug effects / Q000378:metabolism; D007668:Kidney; D008301:Maleimides / Q000494:pharmacology; D008677:Metaphase; D008870:Microtubules / Q000187:drug effects*; D008938:Mitosis / Q000187:drug effects; D016297:Mutagenesis, Site-Directed; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D015739:Nocodazole / Q000494:pharmacology*; D019696:Peptidylprolyl Isomerase / Q000378:metabolism*; D010766:Phosphorylation; D010880:Piperidines / Q000494:pharmacology; D011392:Proline / Q000737:chemistry; D025941:Protein Interaction Mapping; D047428:Protein Kinase Inhibitors; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D011725:Pyridines / Q000494:pharmacology; D013329:Structure-Activity Relationship; D014162:Transfection
D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D005419:Flavonoids; D007093:Imidazoles; D007211:Indoles; D008301:Maleimides; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D010880:Piperidines; D047428:Protein Kinase Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D011725:Pyridines; C077990:alvocidib; D011392:Proline; D016203:CDC2 Protein Kinase; C000606846:PIN1 protein, human; D019696:Peptidylprolyl Isomerase; C070515:bisindolylmaleimide I; C093642:SB 203580; D015739:Nocodazole; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one