Table of Contents

Volume 9, Issue 5

  March 1, 2010

  Pages 851 - 1023

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 Issue Catalogue (MARC XML)

EDITORIALS: CELL CYCLE FEATURES

Open Access Article

Differences underlying EGFR and HER2 oncogene addiction

Anthony C. Faber, Kwok-Kin Wong and Jeffrey Engelman
Pages 851 - 852
http://dx.doi.org/10.4161/cc.9.5.11096
Abstract | PDF

Comment on: Faber AC, et al. Differential induction of apoptosis in HER2 and EGFR addicted cancers following PI3K inhibition. Proc Natl Acad Sci USA 2009; 106:19503-8.

Open Access Article

PERpetual motion of the circadian negative feedback loop

Choogon Lee, Rongmin Chen and Hyeong-min Lee
Pages 853 - 854
http://dx.doi.org/10.4161/cc.9.5.11098
Abstract | PDF

Comment on: Chen R, et al. Rhythmic PER Abundance Defines a Critical Nodal Point for Negative Feedback within the Circadian Clock Mechanism. Mol Cell 2009; 36:417-30.

Open Access Article

Deregulation of innate immune signaling in myelodysplastic syndromes is associated with deletion of chromosome arm 5q

Daniel T. Starczynowski and Aly Karsan
Pages 855 - 856
http://dx.doi.org/10.4161/cc.9.5.11156
Abstract | PDF

Comment on: Starczynowski DT, et al. Identification of miR-145 and miR-146a as mediators of the 5q- syndrome phenotype. Nat Med 2010; 16:49-58.

Open Access Article

Targeting the degradation of cyclin D1 will help to eliminate oncogene addiction

James T. Lee, Jing Shan and Wei Gu
Pages 857 - 858
http://dx.doi.org/10.4161/cc.9.5.11175
Abstract | PDF

Comment on: Shan J, et al. Mol Cell 2009;36:469-76.

Open Access Article

HIF-2α: Many cancers, one engine?

Aleksandra Franovic and Stephen Lee
Pages 859 - 860
http://dx.doi.org/10.4161/cc.9.5.11183
Abstract | PDF

Comment on: Franovic A, et al. Human cancers converge at the HIF-2α oncogenic axis. Proc Natl Acad Sci USA 2009; 106:21306-11.




CELL CYCLE NEWS & VIEWS

Open Access Article

Protein phosphatases and the dynamics of the DNA damage response

Dennis R. Harris and Fred Bunz
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.10862
Abstract | PDF

Comment on: Freeman AK, et al. Cell Cycle 2010; 9:736-47.

Open Access Article

Freezing Polo in its sleep: Targeting the inactive conformation of Polo-like kinase 1 in cancer cells

Frank Eckerdt
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.10868
Abstract | PDF

Comment on: Keppner S, et al. Cell Cycle 2010; 9:761-74.

Open Access Article

Fine-tuning the DNA damage response: Protein Phosphatase 2A checks on CHK2

Marcus B. Smolka
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.10906
Abstract | PDF

Comment on: Freeman AK, et al. Cell Cycle 2009; 9:In press.

Open Access Article

Cell cycle regulated genes in the three domains of life

Lori M. Kelman and Zvi Kelman
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.10907
Abstract | PDF

Comment on: Bernander R, et al. Cell Cycle 2010; 9:795-806.

Open Access Article

Mitochondrial targeting signals: Another barcode in p14ARF?

Koji Itahana and Yanping Zhang
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.11039
Abstract | PDF

Comment on: Irvine M, et al. Cell Cycle 2010; 9:829-39.

Open Access Article

p53 and Mdm2: An auld alliance

Arnaud Vigneron and Karen H. Vousden
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.11040
Abstract | PDF

Comment on: Lane DP, et al. Cell Cycle 2009; 9:540-7 and Lane DP, et al. Cell Cycle 2009; 9:748-54.

Open Access Article

Cryptic mitochondrial targeting signals in human ARF: Clues to controlling cancer?

Beric R. Henderson
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.11041
Abstract | PDF

Comment on: Irvine M, et al. Cell Cycle 2010; 9:829-39.

Open Access Article

The archaeal cell cycle: Clues from comparative genomics

Patrick Forterre
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.10997
Abstract | PDF

Comment on: Bernander R, et al. Cell Cycle 2010; 9:795-806.

Open Access Article

mTOR goes to the nucleus

Yu Jiang
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.11070
Abstract | PDF

Comment on: Tsang CK, et al. Cell Cycle 2010; 9:953-7.

Open Access Article

Skip the nucleus, AKT drives Skp2 and FOXO1 to the same place?

Hui Zhang
Pages 861 - 869
http://dx.doi.org/10.4161/cc.9.5.11153
Abstract | PDF

Comment on: Wang H, et al. Cell Cycle 2010; 9:1021-2.




REVIEWS

Open Access Article

The p53 response element and transcriptional repression

Bei Wang, Ziwei Xiao, Hui Ling Ko and Ee-Chee Ren
Pages 870 - 879
http://dx.doi.org/10.4161/cc.9.5.10825
Abstract | PDF

p53 tumor suppressor has been widely recognized as the "Guardian of the Genome", reflecting its importance in ensuring the proper functioning of the cell. It is well-known for its function as a transcription factor, capable of mediating both transcriptional activation and repression, which brings about many cellular outcomes such as cell-cycle arrest, apoptosis, cellular senescence and DNA repair. The canonical p53 response element (p53RE), which contains two repeats of a decamer motif "RRRCWWGYYY" separated by a spacer of 0 to 13 base-pairs, has been characterized as the regulatory region on the target genes that p53 binds for transcriptional activation. It was thought that p53 generally represses genes that lack this canonical p53RE, presumably through the sequestration of basal transcriptional machinery components or transcription activators. However, characterization of individual genes as well as genome-wide studies utilizing gene expression profiling and chromatin immunoprecipitation uncovered a large number of potential p53-repressed targets. Taken together, there appears to be multiple modes of gene repression by p53 with some being mediated through direct binding of p53 to DNA. The aim of this review is to assess the evidence of p53 mediated transcriptional repression and discuss its role in cellular function.

Open Access Article

Human iPS cell-based therapy: Considerations before clinical applications

Ning Sun, Michael T. Longaker and Joseph C. Wu
Pages 880 - 885
http://dx.doi.org/10.4161/cc.9.5.10827
Abstract | PDF

Generation of induced pluripotent stem (iPS) cells has revolutionized the field of regenerative medicine. With the exponential increase in iPS cell research in the past three years, human iPS cells have been derived with different technologies and from various cell types. From a translational perspective, however, a number of issues must be addressed before safe and high quality patient-specific iPS cells can be derived for clinical applications. In addition, iPS cell-based therapies also need to be thoroughly evaluated in pre-clinical animal models before they can be applied to human subjects.

Open Access Article

ZEB proteins link cell motility with cell cycle control and cell survival in cancer

Gareth Browne, A. Emre Sayan and Eugene Tulchinsky
Pages 886 - 891
http://dx.doi.org/10.4161/cc.9.5.10839
Abstract | PDF

Epithelial mesenchymal transitions (EMT), the generation of motile mesenchymal cells from epithelial sheets, are differentiation programs which take place at several critical steps of embryonic development and in metastatic cancer. Recent data have shown that the transcription factors which are master regulators of EMT also regulate cell cycle progression, apoptosis and senescence. In light of these new observations, the role of these factors in human cancer may be broader than previously anticipated. Here we review recent literature on non-EMT functions of EMT-controlling transcription factors. We will mainly focus on transcription factors belonging to the ZEB family, but some important results obtained by investigators studying other key EMT regulators, Snail and Twist are also discussed.

Open Access Article

Regulation of myogenic stem cell behaviour by vessel cells: The "ménage à trois" of satellite cells, periendothelial cells and endothelial cells

Rana Abou-Khalil, Rémi Mounier and Bénédicte Chazaud
Pages 892 - 896
http://dx.doi.org/10.4161/cc.9.5.10851
Abstract | PDF

In skeletal muscle, satellite cells, that are responsible of muscle repair, are localized close to capillaries. Although angiogenesis is known for a long time to be crucial for muscle repair and satellite cell survival, cellular interplays between vessel cells and satellite/myogenic cells have been poorly explored. We analyzed the interrelationships between myogenic cells, endothelial cells, and periendothelial cells that includes smooth muscle cells and endomysial fibroblasts. We found that endothelial cells strongly stimulate myogenic cell growth and, inversely, myogenic cells increase angiogenesis. VEGF plays a essential role in this bidirectional interaction. On the contrary, periendothelial cells promote the return to quiescence of a subset of muscle precursor cells to quiescence that ensures self-renewal of adult muscle stem cells. We have shown that Angiopoietin-1/Tie-2 signalling controls the entry into quiescence. We propose that during muscle regeneration, i.e. while vessels are not stabilized, endothelial cells and myogenic cells interact with each other to promote both myogenesis and angiogenesis, that have been shown to be concomitant processes in several models. On the other hand, once homeostasis of muscle is reached, the proximity of satellite cells and periendothelial cells allows the responsiveness of satellite cells, that bear Tie-2 receptor, to the secretion of Angiopoietin-1 by periendothelial cells, that, in the same time, stabilize vessels by promoting quiescence of endothelial cells.

Open Access Article

Tumor initiation via loss of cell contact inhibition versus Ras mutation: Do all roads lead to EMT?

Yongqing Liu and Douglas C. Dean
Pages 897 - 900
http://dx.doi.org/10.4161/cc.9.5.10933
Abstract | PDF

Many tumors have been found to contain a subset of cells referred to as cancer stem cells. As opposed to the remainder of the tumor, these cells are undifferentiated—they do not express markers of differentiation and they can re-express stem cell specification genes. The cells can divide asymmetrically to yield differentiated cells as well as cells comprising the original heterogeneous population of the tumor while maintaining their number. Because cancer stem cells display some properties of stem cells, it has been presumed that the presence of such cells in tumors reflects a stem cell origin for cancer. However, recent studies suggest that cancer can originate with outgrowth of differentiated somatic cells, and that cells with properties of cancer stem cells can be generated as tumors progress. Appearance of such cells may be linked to reprogramming of differentiated somatic cells to a stem cell-like phenotype by overexpression of transcription factors involved in epithelial-mesenchymal transition (EMT). We discuss recent studies from our group and other laboratories linking cell outgrowth in tumors to inhibition of the RB1 pathway, loss of cell contact inhibition and generation of undifferentiated cancer stem-like cells from somatic cells. And, we compare this pathway to that arising with introduction of mutant Ras in cells.




EXTRA VIEWS

Open Access Article

Epstein-Barr virus in Burkitt’s lymphoma: A role for latent membrane protein 2A

Kathryn T. Bieging, Michelle Swanson-Mungerson, Alexandra C. Amick and Richard Longnecker
Pages 901 - 908
http://dx.doi.org/10.4161/cc.9.5.10840
Abstract | PDF

Burkitt’s lymphoma (BL) is characterized by translocation of the MYC gene to an immunoglobulin locus. Transgenic mouse models have been used to study the molecular changes that are necessary to bypass tumor suppression in the presence of translocated MYC. Inactivation of the p53 pathway is a major step to tumor formation in mouse models that is also seen in human disease. Human BL is often highly associated with Epstein-Barr virus (EBV). The EBV latency protein latent membrane protein 2A (LMP2A) is known to promote B cell survival by affecting levels of pro-survival factors. Using LMP2A transgenic mouse models, we have identified a novel mechanism that permits lymphomagenesis in the presence of an intact p53 pathway. This work uncovers a contribution of EBV to molecular events that have documented importance in BL pathogenesis, and may underlie the poorly understood link between EBV and BL.

Open Access Article

Biologic and experimental variation of measured cancer stem cells

Michael Heuser and R. Keith Humphries
Pages 909 - 912
http://dx.doi.org/10.4161/cc.9.5.10852
Abstract | PDF

Whereas it has become clear that measured stem cell frequencies in tumors greatly depend on the assay system used, the research focus now shifts towards identification of the biologic variability of cancer stem cells in different disease subsets. In a recent study we quantified the frequency and in vitro expansion potential of leukemia initiating cells in a murine model of acute myeloid leukemia driven either by retroviral overexpression of MN1 and a control vector, or by MN1 and a HOX gene through limiting dilution transplantation assays in syngeneic mice. Both leukemia-initiating cell frequency and expansion potential were increased by over two orders of magnitude in the two-oncogene compared to the one-oncogene model, documenting the functional heterogeneity of leukemia-initiating cells. Loss-of-function studies showed that STAT5b and STAT1 are critical for the enhanced self-renewal activity. Here we discuss implications of our findings and potential sources of experimental variability of measured leukemia or cancer stem cell frequencies.

Open Access Article

Pathological angiogenesis facilitates tumor cell dissemination and metastasis

Pegah Rouhi, Samantha Lin Chiou Lee, Ziquan Cao, Eva-Maria Hedlund, Lasse Dahl Jensen and Yihai Cao
Pages 913 - 917
http://dx.doi.org/10.4161/cc.9.5.10853
Abstract | PDF

Clinically detectable metastases represent an ultimate consequence of the metastatic cascade that consists of distinct processes including tumor cell invasion, dissemination, metastatic niche formation, and re-growth into a detectable metastatic mass. Although angiogenesis is known to promote tumor growth, its role in facilitating early events of the metastatic cascade remain poorly understood. We have recently developed a zebrafish tumor model that enables us to study involvement of pathological angiogenesis in tumor invasion, dissemination and metastasis. This non-invasive in vivo model allows detection of single malignant cell dissemination under both normoxia and hypoxia. Further, hypoxia-induced VEGF significantly facilitates tumor cell invasion and dissemination. These findings demonstrate that VEGF-induced pathological angiogenesis is essential for tumor dissemination and further corroborates potentially beneficial effects of clinically ongoing anti-VEGF drugs for the treatment of various malignancies.

Open Access Article

Micro-orchestrating differentiation in cancer

Riccardo Taulli, Francesca Bersani and Carola Ponzetto
Pages 918 - 922
http://dx.doi.org/10.4161/cc.9.5.10864
Abstract | PDF

Differentiation involves repression of genes governing proliferation and self-renewal, and transcriptional activation of lineage-specific genes. The mechanisms underlying these changes are epigenetic. In cancer cells differentiation genes are locked into a transcriptionally inactive state. Recent results show that in spite of the diversity of the genetic lesions leading to a cancerous phenotype, it may still be possible to release this block and to force differentiation. The key may be microRNAs (miRNAs) which directly or indirectly target epigenetic modifiers. These miRNAs could allow the non-toxic differentiative approach currently adopted in some hematologic malignancies to apply to solid tumors.

Open Access Article

The transcriptional regulation of miR-21, its multiple transcripts and their implication in prostate cancer

Judit Ribas and Shawn E. Lupold
Pages 923 - 929
http://dx.doi.org/10.4161/cc.9.5.10930
Abstract | PDF

MicroRNAs (miRNAs) are a natural part of the most recently discovered and global regulatory pathway known as RNA interference. Functional studies have shown how specific miRNAs can function as tumor suppressors or oncogenes and, correspondingly, deregulated miRNA profiles have been observed in prostate and other cancers. However, the upstream pathways which regulate miRNA expression are only currently being uncovered. The Androgen Receptor (AR) is a nuclear hormone receptor and transcription factor which plays a paramount role in prostate cancer pathobiology. We performed high throughput miRNA microarray analysis on two AR-responsive cell lines to identified 16 candidate AR-regulated miRNAs. One of the most androgen-induced candidates was a known oncogenic miRNA, miR-21. In a small study of early grade Prostate cancer (PCa) samples we found that miR-21 levels were frequently elevated in comparison to adjacent normal tissue. This observation was supported in the literature and suggests clinical relevance. We found that the activated AR directly interacts with miR-21 regulatory regions, indicating direct transcriptional induction. Furthermore, we provide new reporter studies supporting AR-regulation. Importantly, in functional studies, we found that a modest over-expression of miR-21 enhanced tumor xenograft growth and was sufficient to support androgen-independent proliferation following surgical castration. Thus, our studies suggest a model where miR-21 contributes to androgen-dependent and androgen-independent PCa growth. However, the AR is only one of many reported transcriptional regulators of miR-21. Here we review our recent discoveries and further analyze the reported miR-21 regulatory regions, inhibitory and stimulatory signaling pathways, and primary transcripts.

Open Access Article

Tip60: Connecting chromatin to DNA damage signaling

Yingli Sun, Xiaofeng Jiang and Brendan D. Price
Pages 930 - 936
http://dx.doi.org/10.4161/cc.9.5.10931
Abstract | PDF

Cells are constantly exposed to genotoxic events that can damage DNA. To counter this, cells have evolved a series of highly conserved DNA repair pathways to maintain genomic integrity. The ATM protein kinase is a master regulator of the DNA double-strand break (DSB) repair pathway. DSBs activate ATM’s kinase activity, promoting the phosphorylation of proteins involved in both checkpoint activation and DNA repair. Recent work has revealed that two DNA damage response proteins, the Tip60 acetyltransferase and the mre11-rad50-nbs1 (MRN) complex, co-operate in the activation of ATM in response to DSBs. MRN functions to target ATM and the Tip60 acetyltransferase to DSBs. Tip60’s chromodomain then interacts with histone H3 trimethylated on lysine 9, activating Tip60’s acetyltransferase activity and stimulating the subsequent acetylation and activation of ATM’s kinase activity. These results underscore the importance of chromatin structure in regulating DNA damage signaling and emphasize how histone modifications co-ordinate DNA repair. In addition, human tumors frequently exhibit altered patterns of histone methylation. This rewriting of the histone methylation code in tumor cells may impact the efficiency of DSB repair, increasing genomic instability and contributing to the initiation and progression of cancer.




REPORTS

Open Access Article

Induction of pluripotency in human endothelial cells resets epigenetic profile on genome scale

Maria A. Lagarkova, Maria V. Shutova, Alexandra N. Bogomazova, Ekaterina M. Vassina, Evgeny A. Glazov, Ping Zhang, Albert A. Rizvanov, Ilya V. Chestkov and Sergey L. Kiselev
Pages 937 - 946
http://dx.doi.org/10.4161/cc.9.5.10869
PDF

Open Access Article

p53-mediated induction of Noxa and p53AIP1 requires NFκB

Jim O’Prey, Diane Crighton, Angel G. Martin, Karen H. Vousden, Howard O. Fearnhead and Kevin M. Ryan
Pages 947 - 952
http://dx.doi.org/10.4161/cc.9.5.10872
Abstract | PDF

The intricate regulation of cell survival and cell death is critical for the existence of both normal and transformed cells. Two factors central to these processes are p53 and NFκB, with both factors having ascribed roles in both promoting and repressing cell death. Not surprisingly, a number of studies have previously reported interplay between p53 and NFκB. The mechanistic basis behind these observations, however, is currently incomplete. We report here further insights into this interplay using a system where blockade of NFκB inhibits cell death from p53, but at the same time sensitizes cells to death by TNFα. We found in agreement with a recent report showing that NFκB is required for the efficient activation of the BH3-only protein Noxa by the p53 family member p73, that p53’s ability to induce Noxa is also impeded by inhibition of NFκB. In contrast to the regulation by p73, however, blockade of NFκB downstream of p53 decreases Noxa protein levels without effects on Noxa mRNA. Our further analysis of the effects of NFκB inhibition on p53 target gene expression revealed that while most target genes analysed where unaffected by blockade of NFκB, the p53-mediated induction of the pro-apoptotic gene p53AIP1 was significantly dependent on NFκB. These studies therefore add further insight into the complex relationship of p53 and NFκB and since both Noxa and p53AIP1 have been shown to be important components of p53-mediated cell death responses, these findings may also indicate critical points where NFκB plays a pro-apoptotic role downstream of p53.

Open Access Article

mTOR binds to the promoters of RNA polymerase I- and III-transcribed genes

Chi Kwan Tsang, Hui Liu and X.F. Steven Zheng
Pages 953 - 957
http://dx.doi.org/10.4161/cc.9.5.10876
Abstract | PDF

Target of rapamycin (TOR) is a conserved regulator of gene expression from yeast to humans. In budding yeast, TOR is associated with ribosomal DNA (rDNA) promoter, which is critical for ribosome biogenesis and transfer RNA (tRNA) synthesis. Whether mTOR behaves similarly in mammalian cells is unknown. Here, we report that mTOR is detected at several different promoters in human and murine cells, including that of rDNA and tRNA genes. The association of mTOR with these promoters is regulated by growth signals and sensitive to rapamycin. Together, our observations suggest that mTOR is closely involved in gene regulation at the promoters, which is a conserved mechanism to control RNA polymerase I- and III-dependent genes that are critical for protein synthesis and cell growth.

Open Access Article

Genomic analysis of pandemic (H1N1) 2009 reveals association of increasing disease severity with emergence of novel hemagglutinin mutations

Gennadi V. Glinsky
Pages 958 - 970
http://dx.doi.org/10.4161/cc.9.5.10913
Abstract | PDF

Experimental studies and epidemiological observations during the first wave of the pandemic (H1N1) 2009 suggest that a novel influenza A (H1N1) virus has significant pandemic potential based on high transmissibility of the virus. Substantial uncertainty remains regarding evolution of the clinical severity of this pandemic during the transition to the second wave which is currently underway in the Northern Hemisphere. We carried-out analysis of large volume of clinical, epidemiological and genomics data for assessment of evolution of the current pandemic in United States, Canada, United Kingdom, Australia and Japan based on official reports of public health agencies of corresponding countries. Analysis of reported sequences of virus strains isolated from postmortem samples indicates that 42.9% of individuals who died from laboratory-confirmed cases of the pandemic (H1N1) were infected with the hemagglutinin (HA) Q310H mutant virus. Overall, six of seven (86%) of virus isolates recovered from the necropsy samples have at least one mutation within the HA 301–316 or HA 219–240 regions. During the second wave of the pandemic (H1N1) 2009, there is an increased number of reported double mutant virus isolates with mutations within both of these HA regions. Mutations within HA 219-240 region at the position D239 (D239E/G/N) are reported with higher frequency. In addition, D239G mutants were detected more frequently in viruses isolated from patients with fatal outcomes and in isolates from lungs. Multiple viral isolates with the novel HA 301-316 mutations (I312V and P314S) have been documented. Statistically significant increase of detection of mutant viruses and H1N1-related death rates is documented in July-September reporting time periods. Our analysis seems to indicate that evolution of current pandemic is associated with notable changes in mortality rate among hospitalized patients and increasing number of reported cases of novel mutations of HA gene. Recently emerged HA mutants are: (1) detected in large proportion of virus isolates recovered from the postmortem samples; (2) documented in multiple independent reports around the world; (3) expanding within global viral population; (4) manifesting spatial and temporal patterns of association with increased mortality rate of hospitalized patients. Identification of candidate virus mutants with potential association to increasing disease severity should facilitate clinical and experimental testing of the validity of both “antigenic drift” and increase virulence hypotheses. The results of these follow-up experiments may have a significant impact on ultimate outcomes of current pandemic. Our analysis indicates the urgent need for international surveillance systems that track disease severity and individual patient influenza sequence data in a representative fashion. Information gained from this type of surveillance will direct experimental work that assesses influenza strain-specific features of virulence and transmissibility through carefully designed and timely executed laboratory studies. Practical implementation of these surveillance systems would facilitate the timely evidence-based resolution of critically important relationships between the antigenic drift of mutant strains and immunogenicity of existing vaccines which should be assessed in the laboratory setting during the course of the ongoing pandemic.

Open Access Article

Phosphorylation of Ser72 is dispensable for Skp2 assembly into an active SCF ubiquitin ligase and its subcellular localization

Tarig Bashir, Julia K. Pagan, Luca Busino and Michele Pagano
Pages 971 - 974
http://dx.doi.org/10.4161/cc.9.5.10914
Abstract | PDF

F-box proteins are the substrate recognition subunits of SCF (Skp1, Cul1, F-box protein) ubiquitin ligase complexes. Skp2 is a nuclear F-box protein that targets the CDK inhibitor p27 for ubiquitin- and proteasome-dependent degradation. In G0 and during the G1 phase of the cell cycle, Skp2 is degraded via the APC/CCdh1 ubiquitin ligase to allow stabilization of p27 and inhibition of CDKs, facilitating the maintenance of the G0/G1 state. APC/CCdh1 binds Skp2 through an N-terminal domain (amino acids 46-94 in human Skp2). It has been shown that phosphorylation of Ser69 and Ser72 in this domain dissociates Skp2 from APC/C. More recently, it has instead been proposed that phosphorylation of Skp2 on Ser72 by Akt/PKB allows Skp2 binding to Skp1, promoting the assembly of an active SCFSkp2 ubiquitin ligase, and Skp2 relocalization/retention into the cytoplasm, promoting cell migration via an unknown mechanism. According to these reports, a Skp2 mutant in which Ser72 is substituted with Ala is unable to promote cell proliferation and loses its oncogenic potential. Given the contrasting reports, we revisited these results and conclude that phosphorylation of Skp2 on Ser72 does not control Skp2 binding to Skp1 and Cul1, has no influence on SCFSkp2 ubiquitin ligase activity, and does not affect the subcellular localization of Skp2.

Open Access Article

Phosphorylation of Ser72 does not regulate the ubiquitin ligase activity and subcellular localization of Skp2

Christel Boutonnet, Pierre-Luc Tanguay, Catherine Julien, Geneviève Rodier, Philippe Coulombe and Sylvain Meloche
Pages 975 - 979
http://dx.doi.org/10.4161/cc.9.5.10915
Abstract | PDF

Skp2 is the substrate binding subunit of the SCFSkp2 ubiquitin ligase, which plays a key role in the regulation of cell cycle progression. The activity of Skp2 is regulated by the APCCdh1, which targets Skp2 for degradation in early G1 and prevent premature S phase entry. Overexpression of Skp2 leads to dysregulation of the cell cycle and is commonly observed in human cancers. We have previously shown that Skp2 is phosphorylated on Ser64 and Ser72 in vivo, and that these modifications regulate its stability. Recently, two studies have proposed a role for Ser72 phosphorylation in the cytosolic relocalization of Skp2 and in the assembly and activity of SCFSkp2 ubiquitin ligase complex. We have revisited this question and analyzed the impact of Ser72 phosphorylation site mutations on the biological activity and subcellular localization of Skp2. We show here that phosphorylation of Ser72 does not control Skp2 binding to Skp1 and Cul1, has no influence on SCFSkp2 ubiquitin ligase activity, and does not affect the subcellular localization of Skp2 in a panel of cell lines.

Open Access Article

Ubiquitin editing enzyme UCH L1 and microtubule dynamics: Implication in mitosis

Anjali Bheda, Anuradha Gullapalli, Michael Caplow, Joseph S. Pagano and Julia Shackelford
Pages 980 - 994
http://dx.doi.org/10.4161/cc.9.5.10934
Abstract | PDF

Microtubules are essential components of the cytoskeleton and are involved in many aspects of cell responses including cell division, migration, and intracellular signal transduction. Among other factors, post-translational modifications play a significant role in the regulation of microtubule dynamics. Here, we demonstrate that the ubiquitin-editing enzyme UCH L1, abundant expression of which is normally restricted to brain tissue, is also a part of the microtubule network in a variety of transformed cells. Moreover, during mitosis, endogenous UCH L1 is expressed and tightly associated with the mitotic spindle through all stages of M phase, suggesting that UCH L1 is involved in regulation of microtubule dynamics. Indeed, addition of recombinant UCH L1 to the reaction of tubulin polymerization in vitro had an inhibitory effect on microtubule formation. Unexpectedly, Western blot analysis of tubulin fractions after polymerization revealed the presence of a specific ~50 kDa band of UCH L1 (not the normal ~25 kDa) in association with microtubules, but not with free tubulin. In addition, we show that along with 25 kDa UCH L1, endogenous high molecular weight UCH L1 complexes exist in cells, and that levels of 50 kDa UCH L1 complexes are increasing in cells during mitosis. Finally, we provide evidence that ubiquitination is involved in tubulin polymerization: the presence of ubiquitin during polymerization in vitro by itself inhibited microtubule formation and enhanced the inhibitory effect of added UCH L1. The inhibitory effects of UCH L1 correlate with an increase in ubiquitination of microtubule components. Since besides being a deubiquitinating enzyme, UCH L1 as a dimer has also been shown to exhibit ubiquitin ligase activity, we discuss the possibility that the ~50 kDa UCH L1 observed is a dimer which prevents microtubule formation through ubiquitination of tubulins and/or microtubule-associated proteins.

Open Access Article

Chk1 inhibition after replicative stress activates a double strand break response mediated by ATM and DNA-dependent protein kinase

Samuel McNeely, Chiara Conti, Tahir Sheikh, Himali Patel, Sonya Zabludoff, Yves G. Pommier, Gary K. Schwartz and Archie Tse
Pages 995 - 1004
http://dx.doi.org/10.4161/cc.9.5.10935
Abstract | PDF

Checkpoint kinase 1 (Chk1) regulates cell cycle checkpoints and DNA damage repair in response to genotoxic stress. Inhibition of Chk1 is an emerging strategy for potentiating the cytotoxicity of chemotherapeutic drugs. Here, we demonstrate that AZD7762, an ATP-competitive Chk1/2 inhibitor induces γ-H2AX in gemcitabine-treated cells by altering both dynamics and stability of replication forks, allowing the firing of suppressed replication origins as measured by DNA fiber combing and causing a dramatic increase in DNA breaks as measured by comet assay. Furthermore, we identify ATM and DNA-PK, rather than ATR, as the kinases mediating γ-H2AX induction, suggesting AZD7762 converts stalled forks into double strand breaks (DSBs). Consistent with DSB formation upon fork collapse, cells deficient in DSB repair by lacking BRCA2, XRCC3, or DNA-PK were selectively more sensitive to combined AZD7762 and gemcitabine. Checkpoint abrogation by AZD7762 also caused premature mitosis in gemcitabine-treated cells arrested in G1/early S-phase. Prevention of premature mitotic entry via Cdk1 siRNA knockdown suppressed apoptosis. These results demonstrate that chemosensitization of gemcitabine by Chk1 inhibition results from at least three cellular events namely activation of origin firing, destabilization of stalled replication forks, and entry of cells with damaged DNA into lethal mitosis. Additionally, the current study indicates that the combination of Chk1 inhibitor and gemcitabine may be particularly effective in targeting tumors with specific DNA repair defects.

Open Access Article

Inhibition of focal adhesion kinase decreases tumor growth in human neuroblastoma

Elizabeth A. Beierle, Xiaojie Ma, Jerry Stewart, Carl Nyberg, Angelica Trujillo, William G. Cance and Vita M. Golubovskaya
Pages 1005 - 1015
http://dx.doi.org/10.4161/cc.9.5.10936
Abstract | PDF

Neuroblastoma is the most common extracranial solid tumor of childhood. Focal adhesion kinase (FAK) is an intracellular kinase that regulates both cellular adhesion and apoptosis. FAK is overexpressed in a number of human tumors including neuroblastoma. Previously, we have shown that the MYCN oncogene, the primary adverse prognostic indicator in neuroblastoma, regulates the expression of FAK in neuroblastoma. In this study, we have examined the effects of FAK inhibition upon neuroblastoma using a small molecule [1,2,4,5-benzenetetraamine tetrahydrochloride (Y15)] to inhibit FAK expression and the phosphorylation of FAK at the Y397 site. Utilizing both non-isogenic and isogenic MYCN+/MYCN- neuroblastoma cell lines, we found that Y15 effectively diminished phosphorylation of the Y397 site of FAK. Treatment with Y15 resulted in increased detachment, decreased cell viability and increased apoptosis in the neuroblastoma cell lines. We also found that the cell lines with higher MYCN are more sensitive to Y15 treatment than their MYCN negative counterparts. In addition, we have shown that treatment with Y15 in vivo leads to less tumor growth in nude mouse xenograft models, again with the greatest effects seen in MYCN+ tumor xenografts. The results of the current study suggest that FAK and phosphorylation at the Y397 site plays a role in neuroblastoma cell survival, and that the FAK Y397 phosphorylation site is a potential therapeutic target for this childhood tumor.

Open Access Article

Mutation of the DEAD-box helicase belle downregulates the cyclin-dependent kinase inhibitor Dacapo

Aaron M. Ambrus and Maxim V. Frolov
Pages 1016 - 1020
http://dx.doi.org/10.4161/cc.9.5.10953
Abstract | PDF

The retinoblastoma protein (pRB) negatively regulates cell proliferation by limiting the activity of the family of E2F transcription factors. In Drosophila, mutation of the DEAD-box helicase belle (bel) relieves an E2F/pRB induced G₁ cell cycle arrest; however, the mechanism of this rescue is unknown. Here, we show that the level of the cyclin-dependent kinase inhibitor Dacapo (Dap), homolog of mammalian p21/p27, is strongly reduced both in bel mutant cells in vivo and in tissue culture cells depleted of Bel by RNA interference. Interestingly, the loss of bel also partially alleviates an ectopically induced G₁ cell cycle arrest. Additionally, we show that Bel undergoes nucleocytoplasmic shuttling. Thus, inactivation of bel renders cells less sensitive to several anti-proliferative signals inducing G₁ arrest.




LETTER TO THE EDITOR

Open Access Article

A comparison between Skp2 and FOXO1 for their cytoplasmic localization by Akt1

Hongbo Wang, Jinhua Cui, Frederick Bauzon and Liang Zhu
Pages 1021 - 1022
http://dx.doi.org/10.4161/cc.9.5.10916
PDF




ERRATUM

Open Access Article

Spermidinyl-CoA-based HAT inhibitors block DNA repair and provide cancer-specific chemo- and radiosensitization

Keya Bandyopadhyay, Jean-Louis Banères, Aimée Martin, Casimir Blonski, Joseph Parello and Ruth Gjerset
Page 1023
http://dx.doi.org/10.4161/cc.9.5.11373
Abstract | PDF

In the article by Bandyopadhyay et al. entitled “Spermidinyl-CoA-based HAT inhibitors block DNA repair and provide cancer-specific chemo- and radiosensitization,” which appeared in the September 1, 2009 issue of Cell Cycle (pp.2779-88), the first sentence in part (D) of the legend to Figure 3 appeared incorrectly as “(D) Inhibition of g-H2A.X acetylation by 1a: Top row – SDS-PAGE/Western of g-H2A.X in H358 cells treated with 10 µM CPT (1 hour) followed by incubation for an additional hour in the presence (right) or absence (left) of 50 µM 1a (60 µg/lane).” The correct sentence reads as follows: “(D) Inhibition of g-H2A.X acetylation by 1a: Top row – SDS-PAGE/Western of g-H2A.X in H358 cells treated with 5nM CPT (18 hours) in the presence (right lane) or absence (left lane) of 50 µM 1a (60 µg/lane).”