Critical requirement for cell cycle inhibitors in sustaining nonproliferative states

Pajalunga, Deborah; Mazzola, Alessia; Salzano, Anna Maria; Biferi, Maria Grazia; Luca, Gabriele De; Crescenzi, Marco

doi:10.1083/jcb.200608109

Cited by 77 publications

(107 citation statements)

References 57 publications

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“…It is of note that although the levels of this cyclin-dependent kinase inhibitors decrease, it is required for maintaining the post-mitotic state of myotubes. 19 The apoptosis protease-activating factor 1 (Apaf-1) is progressively downregulated in post-mitotic muscle cells. The decrease in Apaf-1 of myotubes has been associated with their resistance to apoptosis.…”

Section: Resultsmentioning

confidence: 99%

DNA damage response by single-strand breaks in terminally differentiated muscle cells and the control of muscle integrity

et al. 2012

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DNA single-strand breaks (SSB) formation coordinates the myogenic program, and defects in SSB repair in post-mitotic cells have been associated with human diseases. However, the DNA damage response by SSB in terminally differentiated cells has not been explored yet. Here we show that mouse post-mitotic muscle cells accumulate SSB after alkylation damage, but they are extraordinarily resistant to the killing effects of a variety of SSB-inducers. We demonstrate that, upon SSB induction, phosphorylation of H2AX occurs in myotubes and is largely ataxia telangiectasia mutated (ATM)-dependent. However, the DNA damage signaling cascade downstream of ATM is defective as shown by lack of p53 increase and phosphorylation at serine 18 (human serine 15). The stabilization of p53 by nutlin-3 was ineffective in activating the cell death pathway, indicating that the resistance to SSB inducers is due to defective p53 downstream signaling. The induction of specific types of damage is required to activate the cell death program in myotubes. Besides the topoisomerase inhibitor doxorubicin known for its cardiotoxicity, we show that the mitochondria-specific inhibitor menadione is able to activate p53 and to kill effectively myotubes. Cell killing is p53-dependent as demonstrated by full protection of myotubes lacking p53, but there is a restriction of p53-activated genes. This new information may have important therapeutic implications in the prevention of muscle cell toxicity. Cells respond to genotoxic stress by activating a signaling cascade known as the DNA damage response (DDR). The DDR is a complex interlaced network comprised of DNA damage repair factors and cell cycle regulators. 1 Our knowledge of the mechanisms of DDR mainly relies on studies conducted in proliferating cells, in which the cell cycle machinery is integrated with the DNA damage signaling. Much less is known in post-mitotic cells that undergo irreversible cell cycle withdrawal. DNA repair is strongly affected by the exit from the cell cycle as revealed by downregulation of the major DNA repair pathways. 2 This occurs during differentiation-associated gene reprogramming at transcriptional level as in the case of genes coding for proteins shared by DNA repair and replication (e.g., replicative DNA polymerases, Flap structure-specific endonuclease 1, proliferating cell nuclear antigen and DNA ligase 1) 3 or repair proteins that are cell-cycle related (e.g., XRCC1 (X-ray repair complementing defective repair in Chinese hamster cells 1), uracil-DNA glycosylase). 4,5 Alternatively, post-translational modifications may modify the efficiency of specific DNA repair components as in the case of transcription factor II H that, because of reduced ubiquitination, may lead to decreased global genomic nucleotide excision repair typical of differentiated cells. 6 Exposure of single-stranded (ss) DNA and/or the generation of double-strand breaks (DSB) are powerful activators of DDR by recruiting and activating two protein kinases, ataxia telangiectasia and Rad3-related (ATR)...

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Section: Resultsmentioning

confidence: 99%

DNA damage response by single-strand breaks in terminally differentiated muscle cells and the control of muscle integrity

et al. 2012

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“…However, we found that this state is readily and rapidly reversed by inactivating the oncogenic stimulus in combination with the transient knockdown of p16 and p21. Likewise, the cell-cycle exit of replicatively senescent human cells or terminally differentiated myotubes is readily reversed by CKI knockdown (Beausejour et al, 2003;Pajalunga et al, 2007). The senescent state thus appears to depend on a dynamic regulatory network that must be actively maintained in order to prevent reversion.…”

Section: Discussionmentioning

confidence: 99%

Parallel pathways in RAF-induced senescence and conditions for its reversion

et al. 2011

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We developed a clonal WI-38hTERT/GFP-RAF1-ER immortal cell line to study RAF-induced senescence of human fibroblasts. Activation of the GFP-RAF1-ER kinase by addition of 4-hydroxy-tamoxifen led to a robust induction of senescence within one population doubling, accompanied by the assembly of heterochromatic foci. At least two pathways contribute in parallel to this senescence leading to the accumulation of p15, p16, p21 and p27 inhibitors of cyclin-dependent kinases (CKIs). Cells that traversed S phase after RAF1 kinase activation experienced a replicative stress manifested by phosphorylation of H2AX and Chk2 and synthesis of p21. However, about half the cells in the population were blocked without passing through S phase and did not show activation of DNA-damage checkpoints. When the cells were cultivated in 5% oxygen, RAF1 activation generated minimal reactive oxygen species, but RAF-induced senescence occurred efficiently in these conditions even in the presence of anti-oxidants or inhibitors of DNA checkpoint pathways. Despite the presence of heterochromatic foci, simultaneous knockdown of p16 and p21 with inactivation of the GFP-RAF1-ER kinase led to rapid reversion of the senescent state with the majority of cells becoming competent for long-term proliferation. These results demonstrate that replicative and oxidative stresses are not required for RAF-induced senescence, and this senescence is readily reversed upon loss of CKIs.

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“…It is noteworthy that there are also reports in the literature indicating that different forms of senescence might be reversible. For instance, a previous study demonstrated that non-proliferating cells can be mitotically reactivated by the sole suppression of cyclin-dependent kinase inhibitors (CKIs) in the absence of exogenous mitogens (Pajalunga et al, 2007). RNA interference-mediated suppression of CKIs efficiently triggered mitosis in terminally differentiated skeletal muscle cells, quiescent fibroblasts and senescent embryo kidney cells.…”

Section: Downregulation Of Tacc3 Induces Cellular Senescence S Schmidmentioning

confidence: 99%

The centrosomal protein TACC3 controls paclitaxel sensitivity by modulating a premature senescence program

et al. 2010

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Microtubule-interfering cancer drugs such as paclitaxel (PTX) often cause chemoresistance and severe side effects, including neurotoxicity. To explore potentially novel antineoplastic molecular targets, we investigated the cellular response of breast carcinoma cells to short hairpin(sh)RNA-mediated depletion of the centrosomal protein transforming acidic coiled coil (TACC) 3, an Aurora A kinase target expressed during mitosis. Unlike PTX, knockdown of TACC3 did not trigger a cell death response, but instead resulted in a progressive loss of the pro-apoptotic Bcl-2 protein Bim that links microtubule integrity to spindle poison-induced cell death. Interestingly, TACC3-depleted cells arrested in G 1 through a cellular senescence program characterized by the upregulation of nuclear p21 WAF , downregulation of the retinoblastoma protein and extracellular signal-regulated kinase 1/2, formation of HP1c (phospho-Ser83)-positive senescence-associated heterochromatic foci and increased senescence-associated b-galactosidase activity. Remarkably, the onset of senescence following TACC3 knockdown was strongly accelerated in the presence of non-toxic PTX concentrations. Thus, we conclude that mitotic spindle stress is a major trigger of premature senescence and propose that the combined targeting of the centrosomal Aurora A-TACC3 axis together with drugs interfering with microtubule dynamics may efficiently improve the chemosensitivity of cancer cells.

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Critical requirement for cell cycle inhibitors in sustaining nonproliferative states

Cited by 77 publications

References 57 publications

DNA damage response by single-strand breaks in terminally differentiated muscle cells and the control of muscle integrity

DNA damage response by single-strand breaks in terminally differentiated muscle cells and the control of muscle integrity

Parallel pathways in RAF-induced senescence and conditions for its reversion

The centrosomal protein TACC3 controls paclitaxel sensitivity by modulating a premature senescence program

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