title: Caspase-dependent Mcl-1 cleavage and effect of Mcl-1 phosphorylation in ABT-737-induced apoptosis in human acute lymphoblastic leukemia cell lines.
abstract: ABT-737 is a BH3-mimetic that has a wide spectrum of single-agent activity against acute lymphoblastic leukemia (ALL) cell lines and xenografts. Previously, we reported that in response to ABT-737, ABT-737-resistant ALL cell lines showed an apparent increase in Mcl-1 (an anti-apoptotic Bcl-2 family protein that is not effectively inhibited by ABT-737) while ABT-737-sensitive ALL cell lines showed decreased Mcl-1 levels. Here we explored the mechanism of Mcl-1 cleavage by ABT-737 and the effect of adjacent phosphorylation sites on Mcl-1 cleavage and apoptosis induced by ABT-737 in a human B-lineage ALL cell line. Caspase cleavage sites in Mcl-1 and the effect of mutation in Mcl-1 phosphorylation sites were determined by transducing Mcl-1 variants tagged with the V5 epitope into human ALL cells. Cytotoxicity was by fluorescence-based DIMSCAN, and changes in protein by immunoblotting. ABT-737 induced a caspase-dependent cleavage of Mcl-1. Of the two Mcl-1 caspase cleavage sites (D127 and D157), D157 was the site of ABT-737-induced cleavage in ALL cells. Cells with exogenously expressed Mcl-1 Δ157 fragment showed greater caspase-3 and caspase-9 activation when they were treated with ABT-737 compared with cells expressing wild-type or D157A mutant Mcl-1. Cells with mutated phosphorylation sites on Mcl-1 (S159A and T163A) were less susceptible to Mcl-1 cleavage and apoptosis induced by ABT-737. Our data showed that Mcl-1 is post-translationally regulated in response to ABT-737 treatment, primarily via a caspase-dependent cleavage that generates a pro-apoptotic Mcl-1 fragment.
authors.affiliation: Cancer Center, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA Cell Biology & Biochemistry, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
authors.forename: YongKu
authors.initials: Y
authors.lastname: Ryu
authors.affiliation: Cancer Center, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA Neuroscience and Pharmacology, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
authors.forename: Connor P
authors.initials: CP
authors.lastname: Hall
authors.affiliation: Cancer Center, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA Cell Biology & Biochemistry, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA Neuroscience and Pharmacology, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
authors.forename: C Patrick
authors.initials: CP
authors.lastname: Reynolds
authors.affiliation: Cancer Center, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA Cell Biology & Biochemistry, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA Neuroscience and Pharmacology, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA min.kang@ttuhsc.edu.
authors.forename: Min H
authors.initials: MH
authors.lastname: Kang
mesh_terms: D017209:Apoptosis; D001402:B-Lymphocytes / Q000187:drug effects* / Q000502:physiology*; D001713:Biphenyl Compounds / Q000633:toxicity*; D020169:Caspases / Q000378:metabolism*; D045744:Cell Line, Tumor; D003584:Cytological Techniques; D006801:Humans; D006868:Hydrolysis; D015151:Immunoblotting; D064549:Myeloid Cell Leukemia Sequence 1 Protein / Q000378:metabolism*; D009596:Nitrophenols / Q000633:toxicity*; D010766:Phosphorylation; D010879:Piperazines / Q000633:toxicity; D013449:Sulfonamides / Q000633:toxicity*
chemical_list: C501332:ABT-737; D001713:Biphenyl Compounds; C579095:MCL1 protein, human; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009596:Nitrophenols; D010879:Piperazines; D013449:Sulfonamides; D020169:Caspases
keywords: ABT-737; Mcl-1; acute lymphoblastic leukemia; caspase-dependent cleavage