Cleavage of Cdc6 by caspase-3 promotes ATM/ATR kinase–mediated apoptosis of HeLa cells

Yim, Hyungshin; Hwang, In-Sul; Choi, Joon-Seok; Chun, Kukjin; Jin, Ying; Ham, Young-Mi; Lee, Kwang Youl; Lee, Seung Ki

doi:10.1083/jcb.200509141

Cited by 34 publications

(41 citation statements)

References 32 publications

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“…The increased nuclear retention of the truncated CDC6 may act as a dominant-negative inhibitor of DNA replication (Yim et al, 2006). These results suggest that in addition to causing apoptosis directly, caspases may also at the same time uncouple the regulation of the cell cycle.…”

Section: Discussionmentioning

confidence: 93%

Generation of an indestructible cyclin B1 by caspase-6-dependent cleavage during mitotic catastrophe

2008

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Overriding the G 2 DNA damage checkpoint permits precocious entry into mitosis that ultimately leads to mitotic catastrophe. Mitotic catastrophe is manifested by an unscheduled activation of CDK1, caspase activation and apoptotic cell death. We found that although cyclin B1 was required for mitotic catastrophe, it was cleaved into a B35 kDa protein by a caspase-dependent mechanism during the process. Cyclin B1 cleavage occurred after Asp123 in the motif ILVD 123 k, and mutation of this motif attenuated the cleavage. Cleavage was abolished by a pan-caspase inhibitor as well as by specific inhibitors for the effector caspase-6 and the initiator caspase-8. Cleavage created a truncated cyclin B1 lacking part of the NH 2 -terminal regulatory domain that included the destruction box sequence. Although cleavage of cyclin B1 itself was not absolutely required for mitotic catastrophe, expression of the truncated product enhanced cell death. In support of this, ectopic expression of this truncated cyclin B1 was not only sufficient to induce mitotic block and apoptosis but also enhanced mitotic catastrophe induced by ionizing radiation and caffeine. These data underscore a possible linkage between mitotic and apoptotic functions by caspase-dependent processing of mitotic activators.

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Section: Discussionmentioning

confidence: 93%

Generation of an indestructible cyclin B1 by caspase-6-dependent cleavage during mitotic catastrophe

2008

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“…Although benzonase has been extensively reported to digest both chromatinassociated and nonassociated DNA, we verified the effectiveness of benzonase to digest chromatin-associated nucleic acid, which is required for complete removal of viral DNA not protected by a capsid ( Fig. 2A and B) (1,24,29,49). Aliquots of 20-day CVPs were digested with benzonase for up to 3 h. The viral and cellular DNA was then extracted, followed by SYBR green-based qPCR.…”

Section: Resultsmentioning

confidence: 99%

Tissue-Spanning Redox Gradient-Dependent Assembly of Native Human Papillomavirus Type 16 Virions

Conway¹,

Alam²,

Ryndock³

et al. 2009

J Virol

189

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Papillomavirus capsids are composed of 72 pentamers reinforced through inter-and intrapentameric disulfide bonds. Recent research suggests that virus-like particles and pseudovirions (PsV) can undergo a redox-dependent conformational change involving disulfide interactions. We present here evidence that native virions exploit a tissue-spanning redox gradient that facilitates assembly events in the context of the complete papillomavirus life cycle. DNA encapsidation and infectivity titers are redox dependent in that they can be temporally modulated via treatment of organotypic cultures with oxidized glutathione. These data provide evidence that papillomavirus assembly and maturation is redox-dependent, utilizing multiple steps within both suprabasal and cornified layers.Human papillomaviruses (HPVs) exclusively infect cutaneous or mucosal epithelial tissues (14,15,30). HPV types that infect the mucosal epithelia can lead to the development of benign or malignant neoplasms, thus allowing for their categorization into low-risk or high-risk HPV types, respectively (14,15,30). A small subset of the more than 200 HPV types now identified are the causative agents of over 75% of all cervical cancers. HPV16 is the most prevalent type worldwide, found in ca. 50 to 62% of squamous cell carcinomas (14, 50).HPV16 virions contain a single, circular double-stranded DNA genome of ϳ8 kb which associates with histones to form a chromatin-like structure. This minichromosome is packaged within a nonenveloped, icosahedral capsid composed of the major capsid protein L1 and the minor capsid protein L2. Similar to polyomaviruses, 72 capsomeres of L1 are geometrically arranged on a Tϭ7 icosahedral lattice (2,9,17,19,36,42). Recent cryoelectron microscopy images of HPV16 pseudovirions (PsV) suggest that L2 is arranged near the inner conical hollow of each L1 pentamer, although it is not known whether each L1 pentamer is occupied with a single L2 protein (5, 42).Due to technical constraints in the production of native HPV virions in organotypic culture, assembly studies of HPV particles have largely been restricted to the utilization of in vitro-derived particles such as virus-like particles (VLPs), PsV, and quasivirions (QV) (6,12,25,40,43). Recent research suggests that HPV and bovine papillomavirus PsV can undergo a redox-dependent conformational change that takes place over the course of many hours. This conformational change is characterized by resistance to proteolysis and chemical reduction and the appearance of a more orderly capsid structure via transmission electron microscopy (TEM) (7,20).We present evidence that native virions, in the context of the complete papillomavirus life cycle, utilize a tissue-spanning redox gradient that facilitates multiple redox-dependent assembly and maturation events over the course of many days. We show that stability and specific infectivity of 20-day virions increases over 10-day virions, 20-day virions are more susceptible to neutralization than 10-day virions, and both viral DNA encapsidation...

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“…17 More recently, cleavage of the cdc6 protein by caspase-3 was described, leading to the accumulation of a nuclear pro-apoptotic fragment of this protein. 18,19 Interestingly, a major regulator of cdc6 is CDK2/ cyclinA: N-terminal phosphorylation of cdc6 by this complex induces its cytoplasmic translocation and inactivation. 20 The cleavage of p27 Kip1 by caspase is of special interest when considering CDC25A, as the target of these two proteins is the same.…”

Section: Discussionmentioning

confidence: 99%

A caspase-dependent cleavage of CDC25A generates an active fragment activating cyclin-dependent kinase 2 during apoptosis

et al. 2008

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The cellular level of the CDC25A phosphatase is tightly regulated during both the normal and genotoxic-perturbed cell cycle. Here, we describe a caspase-dependent cleavage of this protein at residue D223 in non-genotoxic apoptotic conditions. This specific proteolysis generates a catalytically active C-terminal fragment that localizes to the nuclear compartment. Accumulation of this active CDC25A fragment leads to reduced inhibitory phosphorylation of the CDC25A substrate cyclin-dependent kinase 2 (CDK2) on Tyr15. Moreover, CDK2 was found stably associated with this fragment, as well as with an ectopically expressed CDC25A224-525 truncation mutant that mimicks the cleavage product. Ectopic expression of this mutant induced CDK2 Tyr15 dephosphorylation, whereas its catalytically inactive version did not. Finally, this 224-525 mutant initiated apoptosis when transfected into HeLa cells, whereas its catalytic inactive form did not. Altogether, this study demonstrates for the first time that caspase-dependent cleavage of CDC25A is a central step linking CDK2 activation with non-genotoxic apoptotic induction. The dual-specificity phosphatase CDC25A is an effector of key molecular steps during G1/S and M phases of the cell cycle. The initial function described for CDC25A was to remove the phosphate groups from the Thr14 and Tyr15 residues of cyclin-dependent kinase 2 (CDK2), allowing complete activation of this kinase and further progression of cells to S phase. 1,2 A similar function of CDC25A was also described during mitosis, participating in the activation of the Cdc2 kinase. 3 CDC25A is one of the three members of the CDC25 phosphatase family, which all directly activate CDKs throughout the cell cycle (for a recent review, see Boutros et al. 4 ). The specificity of CDC25A, over the B and C isoforms, is its essential function as a G1 phase regulator, and therefore, its regulation by extracellular signaling events. In addition, regulation through ubiquitin-dependent proteasomal degradation of the protein has been extensively described during the last few years, and participates in the G1/S cell cycle arrest and checkpoint (for a review, see Busino et al. 5 ). The cellular level of CDC25A is also dependent on mitogenic extracellular signals such as growth factors 6 and IL-7. 7 We also recently described that adhesion of acute myeloid leukemia (AML) cells to an extracellular matrix upregulates the level of this protein, leading to increased proliferation. 8 Less characterized than its role during the cell cycle are the potential functions of CDC25A during the apoptotic process. A pro-apoptotic role has been attributed to this phosphatase downstream of Myc-induced apoptosis, 9 with CDC25A being a direct transcriptional target of Myc in this context. Conversely, an antiapoptotic function has also been ascribed to the protein in response to serum starvation-induced cell death. 10 Interestingly, a direct interaction of CDC25A with the apoptosis signal regulated kinase-1 (ASK-1) was reported, leading to the inhibition ...

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Cleavage of Cdc6 by caspase-3 promotes ATM/ATR kinase–mediated apoptosis of HeLa cells

Cited by 34 publications

References 32 publications

Generation of an indestructible cyclin B1 by caspase-6-dependent cleavage during mitotic catastrophe

Generation of an indestructible cyclin B1 by caspase-6-dependent cleavage during mitotic catastrophe

Tissue-Spanning Redox Gradient-Dependent Assembly of Native Human Papillomavirus Type 16 Virions

A caspase-dependent cleavage of CDC25A generates an active fragment activating cyclin-dependent kinase 2 during apoptosis

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