Damaged microtubules can inactivate BCL-2 by means of the mTOR kinase

Calastretti, Angela; Bevilacqua, Annamaria; Ceriani, Cristina; Viganò, Simona; Zancai, Paola; Capaccioli, S.; Nicolin, A

doi:10.1038/sj.onc.1204751

Cited by 50 publications

(43 citation statements)

References 63 publications

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“…Identification of this kinase is considered to be of critical importance, since it will provide insight into the molecular links between mitotic arrest and cell death, as well as the molecular mechanism of action of antimitotic drugs. Several candidates have been proposed, including Raf-1 (3), Jun N-terminal protein kinase (JNK) (2,11,36), protein kinase A (PKA) (32), cyclindependent kinase 1 (CDK1) (24), and mammalian target of rapamycin (mTOR) (4). In general, however, conclusions have been correlative or have been based on the use of kinase inhibitors tested under conditions that precluded mitotic arrest and thus indirectly blocked the effects of MTIs.…”

mentioning

confidence: 99%

“…To prepare whole-cell extracts for kinase reactions or size exclusion chromatography, KB-3 cells were harvested, washed, pelleted twice with cold phosphate-buffered saline (PBS) (5 min at 500 ϫ g), and resuspended in lysis buffer (25 mM HEPES [pH 7.5], 300 mM NaCl, 0.1% Triton X-100, 1.5 mM MgCl 2 , 0.2 mM EDTA, 0.5 mM dithiothreitol [DTT], EDTA-free complete protease inhibitor tablets [Roche], 20 g/ml aprotinin, 50 g/ml leupeptin, 10 M pepstatin, 1 mM phenylmethylsulfonyl fluoride, 20 mM ␤-glycerophosphate, 1 mM Na 3 VO 4 , and 1 M okadaic acid). The suspension was incubated for 15 min on ice with occasional mixing; insoluble material was removed by centrifugation (20 min at 100,000 ϫ g); and the supernatant was retained as the whole-cell extract.…”

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confidence: 99%

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Cyclin-Dependent Kinase 1-Mediated Bcl-x_L/Bcl-2 Phosphorylation Acts as a Functional Link Coupling Mitotic Arrest and Apoptosis

Terrano

Upreti

Chambers

2010

Molecular and Cellular Biology

207

205

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Despite detailed knowledge of the components of the spindle assembly checkpoint, a molecular explanation of how cells die after prolonged spindle checkpoint activation, and thus how microtubule inhibitors and other antimitotic drugs ultimately elicit their lethal effects, has yet to emerge. Mitotically arrested cells typically display extensive phosphorylation of two key antiapoptotic proteins, Bcl-x L and Bcl-2, and evidence suggests that phosphorylation disables their antiapoptotic activity. However, the responsible kinase has remained elusive. In this report, evidence is presented that cyclin-dependent kinase 1 (CDK1)/cyclin B catalyzes mitoticarrest-induced Bcl-x L /Bcl-2 phosphorylation. Furthermore, we show that CDK1 transiently and incompletely phosphorylates these proteins during normal mitosis. When mitosis is prolonged in the absence of microtubule inhibition, Bcl-x L and Bcl-2 become highly phosphorylated. Transient overexpression of nondegradable cyclin B1 caused apoptotic death, which was blocked by a phosphodefective Bcl-x L mutant but not by a phosphomimetic Bcl-x L mutant, confirming Bcl-x L as a key target of proapoptotic CDK1 signaling. These findings suggest a model whereby a switch in the duration of CDK1 activation, from transient during mitosis to sustained during mitotic arrest, dramatically increases the extent of Bcl-x L /Bcl-2 phosphorylation, resulting in inactivation of their antiapoptotic function. Thus, phosphorylation of antiapoptotic Bcl-2 proteins acts as a sensor for CDK1 signal duration and as a functional link coupling mitotic arrest to apoptosis.

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confidence: 99%

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confidence: 99%

Cyclin-Dependent Kinase 1-Mediated Bcl-x_L/Bcl-2 Phosphorylation Acts as a Functional Link Coupling Mitotic Arrest and Apoptosis

Terrano

Upreti

Chambers

2010

Molecular and Cellular Biology

207

205

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“…mTOR phosphorylates p70S6K (p70 ribosomal protein S6 kinase) on Thr389, which correlates with the activation of p70S6kinase [24][25][26] , while over-expression of TSC-2 suppresses phosphorylation and activation of p70S6K on residue Thr389 [14][15][16] . In addition, several studies have shown that Akt/mTOR pathway is activated in diabetes and this activation is redox dependent in different cell types [27][28][29] including renal cells [13] . Previous reports have shown that the serine/threonine kinase, mTOR to be involved in the phosphorylation/inactivation of Bcl-2 in microtubules treated with apoptotic agents [30] .…”

Section: Apoptosismentioning

confidence: 99%

Diabetes and renal tubular cell apoptosis

Habib

2013

WJD

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Apoptosis contributes to the development of diabetic nephropathy, but the mechanism by which high glucose induces apoptosis is not fully understood. Apoptosis of tubular epithelial cells is a major feature of diabetic kidney disease, and hyperglycemia triggers the generation of free radicals and oxidant stress in tubular cells. Hyperglycemia and high glucose in vitro also lead to apoptosis, a form of programmed cell death. High glucose similar to those seen with hyperglycemia in people with diabetes mellitus, lead to accelerated apoptosis, a form of programmed cell death characterized by cell shrinkage, chromatin condensation and DNA fragmentation, in variety of cell types, including renal proximal tubular epithelial cells.

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“…Response to mTOR inhibition may differ by cell type, with some investigators reporting induction of apoptosis 11,26 (as we have previously demonstrated) and others G 1 arrest. 27,28 Based on our prior work with murine ALL, we hypothesized that CCI-779-treated human ALL cells would undergo apoptosis. A total of 10 6 lymphoblasts from the 4 patient samples were cultured on bone marrow stroma for 24 hours.…”

Section: Efficacy Of Mtis In Bone Marrow Stroma-supported Culturementioning

confidence: 99%

The mTOR inhibitor CCI-779 induces apoptosis and inhibits growth in preclinical models of primary adult human ALL

Teachey

Obzut

Cooperman

et al. 2006

Blood

158

128

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Acute lymphoblastic leukemia (ALL) in adult patients is often resistant to current therapy, making the development of novel therapeutic agents paramount. We investigated whether mTOR inhibitors (MTIs), a class of signal transduction inhibitors, would be effective in primary human ALL. Lymphoblasts from adult patients with precursor B ALL were cultured on bone marrow stroma and were treated with CCI-779, a second generation MTI. Treated cells showed a dramatic decrease in cell proliferation and an increase in apoptotic cells, compared to untreated cells. We also assessed the effect of CCI-779 in a NOD/SCID xenograft model. We treated a total of 68 mice generated from the same patient samples with CCI-779 after establishment of disease. Animals treated with CCI-779 showed a decrease in peripheralblood blasts and in splenomegaly. In dramatic contrast, untreated animals continued to show expansion of human ALL. We performed immunoblots to validate the inhibition of the mTOR signaling intermediate phospho-S6 in human ALL, finding down-regulation of this target in xenografted human ALL exposed to CCI-779. We conclude that MTIs can inhibit the growth of adult human ALL and deserve close examination as therapeutic agents against a disease that is often not curable with current therapy. IntroductionWhile children with precursor B-cell acute lymphoblastic leukemia (ALL) are often cured, children with relapsed ALL and adults with ALL usually succumb to their disease with current therapy. Even with aggressive therapy, these patient groups have 5-year diseasefree survival rates of only 28% to 39%. 1 Thus, the development of novel therapeutic agents is crucial.One potential class of novel therapeutics is mTOR inhibitors (MTIs). MTIs are a class of signal transduction inhibitors with anticancer activity that were initially developed as immunosuppressive agents. [2][3][4][5] Rapamycin, a macrocyclic lactone produced by Streptomyces hydroscopicus, 6 was the first MTI to be used in a clinical setting. Rapamycin is well tolerated in humans. 7 MTIs also have been shown to be active against a wide variety of tumor types. [8][9][10] We have previously shown that rapamycin induces apoptosis in precursor B ALL lines in vitro and has in vivo activity in transgenic mice with pre-B leukemia/lymphoma. 11 Second generation MTIs, CCI-779 and RAD-001, are currently in phase 1 to phase 3 clinical trials in patients with various cancers, 12-15 but preclinical studies have not previously been performed in primary human ALL.Preclinical testing of chemotherapeutic agents often involves using transformed tumor lines and transgenic mouse models. While these are valuable tools, they may not be representative of human disease. More clinically relevant data may be obtained from systems using primary human ALL cells. Two of these systems, bone marrow stroma-supported culture 16 and xenografting human ALL in nonobese diabetic/severe combined immunodeficient (NOD/ SCID) mice, have recently been developed. 17 Both of these systems allow for direct testing...

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Damaged microtubules can inactivate BCL-2 by means of the mTOR kinase

Cited by 50 publications

References 63 publications

Cyclin-Dependent Kinase 1-Mediated Bcl-x_L/Bcl-2 Phosphorylation Acts as a Functional Link Coupling Mitotic Arrest and Apoptosis

Cyclin-Dependent Kinase 1-Mediated Bcl-x_L/Bcl-2 Phosphorylation Acts as a Functional Link Coupling Mitotic Arrest and Apoptosis

Diabetes and renal tubular cell apoptosis

The mTOR inhibitor CCI-779 induces apoptosis and inhibits growth in preclinical models of primary adult human ALL

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Damaged microtubules can inactivate BCL-2 by means of the mTOR kinase

Cited by 50 publications

References 63 publications

Cyclin-Dependent Kinase 1-Mediated Bcl-xL/Bcl-2 Phosphorylation Acts as a Functional Link Coupling Mitotic Arrest and Apoptosis

Cyclin-Dependent Kinase 1-Mediated Bcl-xL/Bcl-2 Phosphorylation Acts as a Functional Link Coupling Mitotic Arrest and Apoptosis

Diabetes and renal tubular cell apoptosis

The mTOR inhibitor CCI-779 induces apoptosis and inhibits growth in preclinical models of primary adult human ALL

Contact Info

Product

Resources

About

Cyclin-Dependent Kinase 1-Mediated Bcl-x_L/Bcl-2 Phosphorylation Acts as a Functional Link Coupling Mitotic Arrest and Apoptosis

Cyclin-Dependent Kinase 1-Mediated Bcl-x_L/Bcl-2 Phosphorylation Acts as a Functional Link Coupling Mitotic Arrest and Apoptosis