Table of Contents

Volume 9, Issue 16

  August 15, 2010

  Pages 3131 - 3382

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EDITORIALS: CELL CYCLE FEATURES

Open Access Article

NFκB anti-apoptotic or pro-apoptotic, maybe both

Angel G. Martin
Pages 3131 - 3132
http://dx.doi.org/10.4161/cc.9.16.12780
Abstract | PDF

Comment on: Martin AG, et al. Aging 2009;1:335-49.

And Jim O’Prey, et al. Cell Cycle 2010; 9:947-52.

Open Access Article

Stem cell pluripotency: Alternative modes of transcription regulation

Nathan Salomonis and Bruce R. Conklin
Pages 3133 - 3134
http://dx.doi.org/10.4161/cc.9.16.12888
Abstract | PDF

Comment on: Salomonis N, et al. Proc Natl Acad Sci USA 2010;107:10514-9.

Open Access Article

Tristetraprolin and E6-AP: Killing the messenger in cervical cancer

Sandhya Sanduja and Dan A. Dixon
Pages 3135 - 3136
http://dx.doi.org/10.4161/cc.9.16.12951
Abstract | PDF

Comment on: Sanduja S, et al. Aging 2009;1:803-17.

Open Access Article

Processing of human telomeres by the Werner syndrome protein

Sita Reddy, Baomin Li and Lucio Comai
Pages 3137 - 3138
http://dx.doi.org/10.4161/cc.9.16.12952
Abstract | PDF

Comment on: Li B, et al. Aging 2009; 1:289-302.

Open Access Article

Genetic mutants illuminate the roles of RecQ helicases in recombinational repair or response to replicational stress

Monika Aggarwal and Robert M. Brosh
Pages 3139 - 3141
http://dx.doi.org/10.4161/cc.9.16.12953
Abstract | PDF

Comment on: Aggarwal M, et al. Aging; 1:219-33.

Open Access Article

Cdk5: Links to DNA damage

En Huang, Dianbo Qu and David S. Park
Pages 3142 - 3143
http://dx.doi.org/10.4161/cc.9.16.12955
Abstract | PDF

Comment on: Huang E, et al. Nat Cell Biol 2010; 12:563-71.


Open Access Article

Genes essential for cell viability that are linked to tumor suppressor genes play a role in cancer susceptibility

Yuxun Wang, Bin Huang, Lisa Edelmann, Richard D. Kolodner and Winfried Edelmann
Pages 3144 - 3145
http://dx.doi.org/10.4161/cc.9.16.12956
Abstract | PDF

Comment on: Wang Y, et al. Proc Natl Acad Sci USA 2010; 107:5511-5.





NEWS & VIEWS

Open Access Article

tRNA trafficking along the TOR pathway

Thomas P. Neufeld and Andrew M. Arsham
Pages 3146 - 3150
http://dx.doi.org/10.4161/cc.9.16.12781
Abstract | PDF

Comment on: Huynh LN, et al. Cell Cycle 2010; 9:3112-18.

Open Access Article

A role for ATM kinase activity and Mre11 in microhomology-mediated end-joining

Anthony J. Davis and David J. Chen

http://dx.doi.org/10.4161/cc.9.16.12814
Abstract | PDF

Comment on: Rahal EA, et al. Cell Cycle 2010; 9:2866-77.

Open Access Article

The flap about ATM and MRE11

Bevin Engelward

http://dx.doi.org/10.4161/cc.9.16.12815
Abstract | PDF

Comment on:Rahal EA, et al. Cell Cycle 2010; 9:2866-77.

Open Access Article

Fat: Love it to death!

Cristina Mazzoni

http://dx.doi.org/10.4161/cc.9.16.12834
Abstract | PDF

Comment on: Rockenfeller P, et al. Cell Cycle 2010; 9:2836-42.

Open Access Article

p16Ink4A, not only a G₁ inhibitor?

Vjekoslav Dulić

http://dx.doi.org/10.4161/cc.9.16.12887
Abstract | PDF

Comment on:  Chien WW, et al. Cell Cycle 2010; 9:3286-96.




REVIEW OF CONCEPT

Open Access Article

Revisiting the antagonistic pleiotropy theory of aging: TOR-driven program and quasi-program

Mikhail V. Blagosklonny
Pages 3151 - 3156
http://dx.doi.org/10.4161/cc.9.16.13120
Abstract | PDF

A half century ago, antagonistic pleiotropy (AP) theory had solved a mystery of aging, by postulating genes beneficial early in life at the cost of aging. Recently it was argued however that there are very few clear-cut examples of antagonistically pleiotropic (AP) genes other than p53. In contrast, here I discuss that p53 is not a clear-cut example of AP genes but is rather an aging-suppressor (gerosuppressor). In contrast, clear-cut examples of AP genes are genes that encode the TOR (target of rapamycin) pathway. TOR itself is the ultimate example of AP gene because its deletion is lethal in embryogenesis. Early in life the TOR pathway drives developmental program, which persists later in life as an aimless quasi-program of aging and age-related diseases.




EXTRA VIEWS

Open Access Article

Evolution of CST function in telomere maintenance

Carolyn Price, Kara A. Boltz, Mary F. Chaiken, Jason A. Stewart, Mark A. Beilstein and Dorothy E. Shippen
Pages 3157 - 3165
http://dx.doi.org/10.4161/cc.9.16.12547
Abstract | PDF

Telomeres consist of an elaborate, higher-order DNA architecture, and a suite of proteins that provide protection for the chromosome terminus by blocking inappropriate recombination and nucleolytic attack, and facilitate telomeric DNA replication by physical interactions with telomerase and the lagging strand replication machinery. The prevailing view has been that two distinct telomere capping complexes evolved, shelterin in vertebrates and a trimeric complex comprised of Cdc13, Stn1 and Ten1 (CST) in yeast.  The recent discovery of a CST-like complex in plants and humans raises new questions about the composition of telomeres and their regulatory mechanisms in multicellular eukaryotes. In this review we discuss the evolving functions and interactions of CST components and their contributions to chromosome end protection and DNA replication.

Open Access Article

Histone H2AX: The missing link in AIF-mediated caspase-independent programmed necrosis

Mathieu Baritaud, Hanan Boujrad, Hans K. Lorenzo, Slavica Krantic and Santos A. Susin
Pages 3166 - 3173
http://dx.doi.org/10.4161/cc.9.16.12552
Abstract | PDF

Caspase-independent programmed necrosis is a highly regulated cellular demise that displays morphological and biochemical necrotic hallmarks, such as an earlier permeability of the plasma membrane and lactate dehydrogenase (LDH) leakiness. This form of programmed cell death (PCD) is regulated by AIF, a FAD-dependent oxidoreductase, which is released from the mitochondria to the nucleus where it induces chromatin condensation and DNA fragmentation. Some years ago, it has been established that the sequential activation of poly(ADP-ribose) polymerase-1 (PARP-1), calpains, and Bax regulate the mitochondrial AIF release associated to programmed necrosis. But, what happens when AIF is in the nucleus? How does this protein induce chromatinolysis and programmed necrosis? Recently, we have unraveled some of the mechanisms underlying the nuclear action of AIF in this type of caspase-independent cell death. Indeed, AIF plays a key role in programmed necrosis by its ability to organize a DNA-degrading complex with H2AX and Cyclophiline A (CypA). The AIF/H2AX link is indeed a critical event and explains the nuclear AIF apoptogenic action. In the present article, we outline the current knowledge on cell death by programmed necrosis and discuss the relevance of the AIF/H2AX/CypA DNA-degrading complex in the regulation of this original form of cell death.

Open Access Article

Roles of the ubiquitin-proteosome system in neurogenesis

Tran Cong Tuoc and Anastassia Stoykova
Pages 3174 - 3180
http://dx.doi.org/10.4161/cc.9.16.12551
Abstract | PDF

The ubiquitin-proteosome system (UPS) is a non-lysosomal proteolysis system involved in the degradation of irrelevant/misfolded intracellular proteins. The protein substrates of this system are tagged by ubiquitin in sequential reactions that target them for proteasome-dependent destruction. In the developing central nervous system, ubiquitin-mediated proteolysis has recently emerged as an important player in the regulation of neural progenitor proliferation, cell specification, neuronal differentiation, maturation, and migration. E3 ubiquitin ligases are crucial components in the UPS because they provide the specificity that determines which substrates are targeted for ubiquitin-dependent proteolysis. In this review, we discuss the molecular mechanisms of the UPS, focusing primarily on the roles of E3 ligases and their substrates in sequential steps of neurogenesis.


Open Access Article

Life without geminin

Dimitris Karamitros, Panorea Kotantaki, Zoi Lygerou, Henrique Veiga-Fernandes, Vassilis Pachnis, Dimitris Kioussis and Stavros Taraviras
Pages 3181 - 3185
http://dx.doi.org/10.4161/cc.9.16.12554
Abstract | PDF

The interplay of proliferation and differentiation is essential for normal development and organogenesis. Geminin is a cell cycle regulator which controls licensing of origins for DNA replication, safeguarding genomic stability. Geminin has also been shown to regulate cellular decisions of self-renewal versus commitment of neuronal progenitor cells. We discuss here our recent analysis of mice with conditional inactivation of the Geminin gene in the immune system. Our data indicate that Geminin is not indispensable for every cell division: in the absence of Geminin, development of progenitor T cells appears largely unaffected. In contrast, rapid cell divisions, taking place in vitro upon TCR receptor activation or in vivo during homeostatic proliferation, are defective.

Open Access Article

Nutrient sensing and insulin signaling in neuropeptide-expressing immortalized, hypothalamic neurons: A cellular model of insulin resistance

Laura J. Fick and Denise D. Belsham
Pages 3186 - 3193
http://dx.doi.org/10.4161/cc.9.16.12601
Abstract | PDF

Obesity and type 2 diabetes mellitus represent a significant global health crisis.  These two interrelated diseases are typified by perturbed insulin signaling in the hypothalamus.  Using novel hypothalamic cell lines, we have begun to elucidate the molecular and intracellular mechanisms involved in the hypothalamic control of energy homeostasis and insulin resistance.  In this review, we present evidence of insulin and glucose signaling pathways that lead to changes in neuropeptide gene expression.  We have identified some of the molecular mechanisms involved in the control of de novo hypothalamic insulin mRNA expression.  And finally, we have defined key mechanisms involved in the etiology of cellular insulin resistance in hypothalamic neurons that may play a fundamental role in cases of high levels of insulin or saturated fatty acids, often linked to the exacerbation of obesity and diabetes.

Open Access Article

The fate of metaphase kinetochores is weighed in the balance of SUMOylation during S phase

Debaditya Mukhopadhyay and Mary Dasso
Pages 3194 - 3201
http://dx.doi.org/10.4161/cc.9.16.12619
Abstract | PDF

Genetic evidence suggests that conjugation of Small Ubiquitin-like Modifier proteins (SUMOs) plays an important role in kinetochore function, although the mechanism underlying these observations are poorly defined. We found that depletion of the SUMO protease SENP6 from HeLa cells causes chromosome misalignment, prolonged mitotic arrest and chromosome missegregation. Many inner kinetochore proteins (IKPs) were mis-localized in SENP6-depleted cells. This gross mislocalization of IKPs is due to proteolytic degradation of CENP-I and CENP-H via the SUMO targeted Ubiquitin Ligase (STUbL) pathway. Our findings show that SENP6 is a key regulator of inner kinetochore assembly that antagonizes the cellular STUbL pathway to protect IKPs from degradation during S phase. Here, we will briefly review the implications of our findings and present new data on how SUMOylation during S phase can control chromosome alignment in the subsequent metaphase.

Open Access Article

Lgl, the SWH pathway and tumorigenesis: It’s a matter of context and competition!

Nicola A Grzeschik, Linda M Parsons and Helena Richardson
Pages 3202 - 3212
http://dx.doi.org/10.4161/cc.9.16.12633
Abstract | PDF

Loss of function of the neoplastic tumors suppressors, lgl, scrib and dlg, or overexpression of the apical polarity components, Crumbs and atypical protein kinase C (aPKC), are associated with polarity loss and tissue overgrowth, however, the mechanism behind these effects is poorly understood. In our recent study, we showed that Lgl, aPKC and Crumbs mediate their effects on proliferation and survival via the Salvador/Warts/Hippo (SWH) tumor suppressor pathway to mediate their effects on tissue growth. Loss of lgl can lead to substantial overgrowth, however the lgl mutant phenotype can be quite variable and the amount of overgrowth of the mutant tissue, its survival and ultimate fate is strongly determined by context and competition. In this extra-view we present a more detailed description of the lgl mutant phenotype and highlight the phenotypic differences between lgl and SWH pathway mutant phenotypes. In addition, we explore the role for the Jun kinase (JNK) pathway in the development of the lgl mutant phenotype.

Open Access Article

AKT-ing via microRNA

Danish Sayed and Maha Abdellatif
Pages 3213 - 3217
http://dx.doi.org/10.4161/cc.9.16.12634
Abstract | PDF

MicroRNAs are involved in almost every aspect of a mammalian cell's functionality, from stem cell differentiation to aging and pathogenesis; however, their role in immediate cell signaling is less defined.  This has been recently demonstrated by the rapid increase or decrease of miR-21's abundance within minutes of activation or inhibition of the AKT pathway, respectively, which mediates its regulation of Fas ligand (FasL) and phosphatase and tensin homologue deleted on chromosome 10 (PTEN) expression, among other targets.  Conversely, AKT induces rapid downregulation of miR-199a-5p to effect upregulation of hypoxia-inducible factor 1a (Hif-1a) and sirtuin 1 (Sirt1).  This suggests that posttranscriptional mechanisms regulate miRNAs' processing and/or stability to induce the rapid fluctuation in their levels.  In support, a growing number of studies are showing specific posttranscriptional regulation of miRNAs.  The data potentially explain how AKT, and plausibly other signaling pathways, can specifically and promptly modulate a gene's translation while circumventing the need for transcription during transient signaling events.  In this article we present our views regarding cell signaling via miRNAs.

Open Access Article

Clusters, factories and domains: The complex structure of S phase comes into focus

Peter J. Gillespie and Julian Blow
Pages 3218 - 3226
http://dx.doi.org/10.4161/cc.9.16.12644
Abstract | PDF

During S-phase of the cell cycle, chromosomal DNA is replicated according to a complex replication timing program, with megabase-sized domains replicating at different times. DNA fibre analysis reveals that clusters of adjacent replication origins fire near-synchronously. Analysis of replicating cells by light microscopy shows that DNA synthesis occurs in discrete foci or factories. The relationship between timing domains, origin clusters and replication foci is currently unclear. Recent work, using a hybrid Xenopus/hamster replication system, has shown that when CDK levels are manipulated during S-phase the activation of replication factories can be uncoupled from progression through the replication timing program. Here, we use data from this hybrid system to investigate potential relationships between timing domains, origin clusters and replication foci. We suggest that each timing domain typically comprises several replicon clusters, which are usually processed sequentially by replication factories. We discuss how replication might be regulated at different levels to create this complex organisation and the potential involvement of CDKs in this process.




PERSPECTIVE

Open Access Article

Nucleostemin: Another nucleolar "Twister" of the p53-MDM2 loop

Dorothy Lo and Hua Lu
Pages 3227 - 3232
http://dx.doi.org/10.4161/cc.9.16.12605
Abstract

Several nucleolar proteins, such as ARF, ribosomal protein (RP) L5, L11, L23 and S7, have been shown to induce p53 activation by inhibiting MDM2 E3 ligase activity and consequently to trigger cell cycle arrest and/or apoptosis. Our recent study revealed another nucleolar protein called nucleostemin (NS), a nucleolar GTP binding protein, as a novel regulator of the p53-MDM2 feedback loop. However, unlike other known nucleolar regulators of this loop, NS surprisingly plays a dual role, as both up and down regulations of its levels could turn on p53 activity. Here, we try to offer some prospective views for this unusual phenomenon by reconciling previously and recently published studies in the field in hoping to better depict the role of NS in linking the p53 pathway with ribosomal biogenesis during cell growth and proliferation as well as to propose NS as another potential molecular target for anti-cancer drug development.




REVIEWS

Open Access Article

Regulation of yeast forkhead transcription factors and FoxM1 by cyclin-dependent and polo-like kinases

Hiroshi Murakami, Hirofumi Aiba, Makoto Nakanishi and Yuko Murakami-Tonami
Pages 3233 - 3242
http://dx.doi.org/10.4161/cc.9.16.12599
Abstract | PDF

Members of the forkhead-box (Fox) family of transcription factors are present in many eukaryotes. More than 100 such proteins that share homology in the winged-helix DNA-binding domain have been identified in higher eukaryotes. This family of transcription factors is implicated in the regulation of a variety of cellular processes, including the cell cycle, apoptosis, DNA repair, stress resistance, and metabolism. A subfamily of Fox proteins are required to activate expression of the genes encoding B-type cyclins, Cdc25 and Polo-like kinase (Plk) during the mitotic cell cycle and meiosis in organisms from yeast to mammals. These proteins are activators of cyclin-dependent kinase 1 (Cdk1). Cdk1 and Plk phosphorylate Fox and its associated proteins at different sites, resulting in activation or repression of Fox transcriptional activity, depending on the target genes. In addition to their documented transcriptional functions, Fox proteins are involved in the regulation of pre-mRNA processing, at least in yeast. In this review, we will focus on the role of Fox proteins in the fission yeast Schizosaccharomyces pombe and budding yeast Saccharomyces cerevisiae, in addition to the role of FoxM1 in mammals in the cell cycle and in pre-mRNA processing, as revealed in recent studies.

Open Access Article

Three-step model for condensin activation during mitotic chromosome condensation

Franck Bazile, Julie St-Pierre and Damien D'Amours
Pages 3243 - 3255
http://dx.doi.org/10.4161/cc.9.16.12620
Abstract | PDF

Chromosomes undergo a major structural reorganization during mitosis. The first step in this reorganization is the compaction of interphase chromatin into highly condensed mitotic chromosomes. An evolutionarily conserved multi-subunit ATPase, the condensin complex, plays a critical role in establishing chromosome architecture and promoting chromosome condensation in mitosis. How does condensin promote chromosome condensation and how, in turn, is the cell cycle machinery activating or restraining condensin activity during the cell cycle are fundamental questions for cell biology. In this review, we examine the role of post-translational modifications, and in particular multi-site phosphorylation, in the regulation of condensin activity during the cell cycle. Remarkably, inspection of phosphorylation sites identified through multiple proteome-wide mass spectrometry analyses reveals that the phosphorylation landscape of condensin is highly conserved evolutionarily and that several kinases regulate condensin in vivo. This analysis leads us to propose the ultrasensitive-kinase switch model, whereby the phosphorylation of condensin by multiple kinases allows the process of chromosome condensation to be maintained and even increased under fluctuating levels of cyclin-CDK activity during mitosis. Our model reconciles how chromosome condensation might be highly sensitive to low levels of CDK activity in early mitosis and subsequently insensitive to the declining levels CDK activity in late mitosis.




REPORTS

Open Access Article

Oxidative stress in cancer associated fibroblasts drives tumor-stroma co-evolution: A new paradigm for understanding tumor metabolism, the field effect and genomic instability in cancer cells

Ubaldo E. Martinez-Outschoorn, Renee M. Balliet, Dayana Rivadeneira, Barbara Chiavarina, Stephanos Pavlides, Chenguang Wang, Diana Whitaker-Menezes, Kristin Daumer, Zhao Lin, Agnieszka Witkiewicz, Neal Flomenberg, Anthony Howell, Richard Pestell, Erik Knudsen, Federica Sotgia and Michael P. Lisanti
Pages 3256 - 3276
http://dx.doi.org/10.4161/cc.9.16.12553
Abstract | PDF

Loss of stromal fibroblast caveolin-1 (Cav-1) is a powerful single independent predictor of poor prognosis in human breast cancer patients, and is associated with early tumor recurrence, lymph node metastasis, and tamoxifen-resistance. We developed a novel co-culture system to understand the mechanism(s) by which a loss of stromal fibroblast Cav-1 induces a "lethal tumor micro-environment". Here, we propose a new paradigm to explain the powerful prognostic value of stromal Cav-1. In this model, cancer cells induce oxidative stress in cancer associated fibroblasts, which then acts as a "metabolic" and "mutagenic" motor to drive tumor-stroma co-evolution, DNA damage, and aneuploidy in cancer cells. More specifically, we show that an acute loss of Cav-1 expression leads to mitochondrial dysfunction, oxidative stress, and aerobic glycolysis in cancer associated fibroblasts. Also, we propose that defective mitochondria are removed from cancer-associated fibroblasts by autophagy/mitophagy that is induced by oxidative stress. As a consequence, cancer associated fibroblasts provide nutrients (such as lactate) to stimulate mitochondrial biogenesis and oxidative metabolism in adjacent cancer cells (the "Reverse Warburg Effect"). We provide evidence that oxidative stress in cancer associated fibroblasts is sufficient to induce genomic instability in adjacent cancer cells, via a bystander effect, potentially increasing their aggressive behavior. Finally, we directly demonstrate that nitric oxide (NO) over-production, secondary to Cav-1 loss, is the root cause for mitochondrial dysfunction in cancer associated fibroblasts. In support of this notion, treatment with anti-oxidants (such as N-acetyl-cysteine, metformin, and quercetin), or NO inhibitors (L-NAME) was sufficient to reverse many of the cancer-associated fibroblast phenotypes that we describe. Thus, cancer cells use "oxidative stress" in adjacent fibroblasts i) as an "engine" to fuel their own survival via the stromal production of nutrients, and ii) to drive their own mutagenic evolution towards a more aggressive phenotype, by promoting genomic instability. We also present evidence that the "field effect" in cancer biology could also be related to the stromal production of ROS and NO species. eNOS-expressing fibroblasts have the ability to down-regulate Cav-1 and induce mitochondrial dysfunction in adjacent fibroblasts that do not express eNOS. As such, the effects of stromal oxidative stress can be laterally propagated, amplified, and are effectively "contagious"-spread from cell-to-cell like a virus-creating an "oncogenic/mutagenic" field promoting widespread DNA damage.

Open Access Article

miR-33-mediated downregulation of p53 controls hematopoietic stem cell self-renewal

Antonio Herrera-Merchan, Celia Cerrato, Guadalupe Luengo, Orlando Dominguez, Miguel A. Piris, Manuel Serrano and Susana Gonzalez
Pages 3277 - 3285
http://dx.doi.org/10.4161/cc.9.16.12598
Abstract | PDF

Hematopoietic stem cells (HSCs) are defined by their exclusive capacity to both self-renew and to give rise to multipotent progenitors (MPPs) that in turn differentiate into the mature blood cell lineages. The tumor suppressor p53, in addition to its role in the regulation of the cell cycle, plays an importatn role in HSC self-renewal, although it has not fully resolved. Here we report that in super-p53 mice (sp53), which carry one extra gene dose of p53, the miR-33 is down-regulated in HSCs and highly expressed in MPPs. Transplantation assays of miR-33-transduced sp53 HSC results in a significant acquisition of repopulating capacity and a decrease of recipients survival. Moreover, high levels of miR-33 represses the endogenous level of p53 protein in murine embryonic fibroblasts (MEFs), leads both to neoplastic transformation and anchorage independent growth of MEFs, and displays a decrease of apoptotic response using tumor-derived cell lines. Accordingly, we demonstrate that miR-33-mediated down-regulation of p53 is dependent on the binding of miR-33 to two conserved motifs in the 3′UTR of p53. Together, these data show that the miR-33 modifies HSC repopulating efficiency of sp53 mice by impairing the p53 function. Defining the role of miR-33 in controlling the HSC self-renewal through p53 may lead to the prevention and treatment of hematopoietic disorders.

Open Access Article

S-phase lengthening induced by p16^INK4a overexpression in malignant cells with wild-type pRb and p53

Wei Wen Chien, Carine Domenech, Régine Catallo, Gilles Salles and Martine Ffrench
Pages 3286 - 3296
http://dx.doi.org/10.4161/cc.9.16.12600
PDF

Open Access Article

RAF associates with phosphorylated nuclear BubR1 during endoreduplication induced by JAK inhibition

Gudrun Reiterer, Lihua Chen, Chang-Yan Chen and Andrew Yen
Pages 3297 - 3304
http://dx.doi.org/10.4161/cc.9.16.12631
Abstract | PDF

The role of JAK signaling in cell cycle transit and maintenance of genomic stability was determined in HL-60 human myeloblastic leukemia cells. We have previously reported that a pan-JAK inhibitor caused ERK-dependent endoreduplication. In the current study we find that JAK inhibition caused nuclear re-localization of RAF-1 which could be inhibited by RAF inhibitor GW5074. GW5074 also inhibited JAK inhibitor-induced appearance of nuclear phosphorylated RAF-1(pS621RAF) and MEK; and it inhibited the JAK inhibitor-induced co-immunoprecipitation of nuclear RAF-1 and MEK. JAK inhibition also increased nuclear BubR1 phosphorylation, which was diminished by RAF inhibitor GW5074. RAF-1 and BubR1 in the nucleus co-immunoprecipitated; and GW5074 eliminated this. Furthermore, inhibiting RAF with GW5074 blocked the pan-JAK inhibitor-induced endoreduplication. These data thus show that JAK inhibition causes nuclear re-localization and phosphorylation of RAF and MEK where RAF binds BubR1 with ensuing nuclear RAF-dependent BubR1 phosphorylation. Inhibiting RAF inhibited this and endoreduplication. The results suggest that there is a JAK/RAF/MEK/BubR1 axis that can regulate genomic stability. In this hypothetical model JAK suppresses RAF/MEK phosphorylation and nuclear re-localization, but JAK inhibition induces the phosphorylations and relocalization with association of RAF and phosphorylated BubR1 in the nucleus leading to endoreduplication.

Open Access Article

Cooperativity of Cdk4R24C and Ras in melanoma development

Rachna Chawla, Judith A Procknow, Ramana V Tantravahi, Jasbir Khurana, Judith Litvin and E. Premkumar Reddy
Pages 3305 - 3314
http://dx.doi.org/10.4161/cc.9.16.12632
Abstract | PDF

The importance of the CDK4 protein in human cancer first became evident following the identification of a germ line mutation in the Cdk4 locus that predisposes humans to melanoma.  This mutation results in substitution of arginine with cysteine at position 24 (R24C).  In an earlier study, we introduced the R24C mutation into the Cdk4 locus of mice using Cre-loxP-mediated "knock-in" technology and observed a very low incidence of spontaneous melanomas in Cdk4^R24C/R24C mice.  This suggested that additional oncogenic mutations might be required for development of melanomas.  Here we report an increased incidence of spontaneous cutaneous melanoma in mice expressing the oncogene HRAS(G12V) in melanocytes on a Cdk4^R24C background.  Treatment of Tyr-HRas:Cdk4^R24C/R24C mice with the carcinogen, DMBA/TPA resulted in a further increase in the number of nevi and melanomas developed when compared with Tyr-HRas:Cdk4^+/+ mice.  In summary, in Tyr-HRas:Cdk4^R24C/R24C mice, we observed that activated CDK4 cooperates with the oncogenic HRAS(G12V) protein to increase the susceptibility of melanoma development in vivo.

Open Access Article

Eco1 is important for DNA damage repair in S. cerevisiae

Shuai Lu, Matthew Goering, Scarlett Gard, Bo Xiong, Adrian J. McNairn, Sue L. Jaspersen and Jennifer L. Gerton
Pages 3315 - 3327
http://dx.doi.org/10.4161/cc.9.16.12673
Abstract | PDF

The cohesin network has an essential role in chromosome segregation, but also plays a role in DNA damage repair. Eco1 is an acetyltransferase that targets subunits of the cohesin complex and is involved in both the chromosome segregation and DNA damage repair roles of the network. Using budding yeast as a model system, we find that mutations in Eco1, including a genocopy of a human Roberts syndrome allele, do not cause gross defects in chromosome cohesion. We examined how mitotic and meiotic DNA damage repair is affected by mutations in Eco1. Strains containing mutations in Eco1 are sensitive to DNA damaging agents that cause double-strand breaks, such as Xrays and bleomycin. While meiotic crossing over is relatively unaffected in strains containing the Roberts mutation, reciprocal mitotic crossovers occur with extremely low frequency in this mutant background. Our results suggest that Eco1 promotes the reciprocal exchange of chromosome arms and maintenance of heterozygosity during mitosis.

Open Access Article

Absence of p53-dependent apoptosis leads to UV radiation hypersensitivity, enhanced immunosuppression and cellular senescence

Omid Tavana, Cara L Benjamin, Nahum Puebla-Osorio, Mei Sang, Stephen E Ullrich, Honnavara Ananthaswamy and Chengming Zhu
Pages 3328 - 3336
http://dx.doi.org/10.4161/cc.9.16.12688
Abstract | PDF

Genotoxic stress triggers the p53 tumor suppressor network to activate cellular responses that lead to cell cycle arrest, DNA repair, apoptosis or senescence. This network functions mainly through transactivation of different downstream targets, including cell cycle inhibitor p21, which is required for short-term cell cycle arrest or long-term cellular senescence, or proapoptotic genes such as p53 upregulated modulator of apoptosis (PUMA) and Noxa. However, the mechanism that switches from cell cycle arrest to apoptosis is still unknown. In this study, we found that mice harboring a hypomorphic mutant p53, R172P, a mutation that abrogates p53-mediated apoptosis while keeping cell cycle control mostly intact, are more susceptible to ultraviolet-B (UVB)-induced skin damage, inflammation, and immunosuppression than wild-type mice. p53^R172P embryonic fibroblasts (MEFs) are hypersensitive to UVB and prematurely senesce after UVB exposure, in stark contrast to wild-type MEFs, which undergo apoptosis.  However, these mutant cells are able to repair UV-induced DNA lesions, indicating that the UV hypersensitive phenotype results from the subsequent damage response. Mutant MEFs show an induction of p53 and p21 after UVR, while wild-type MEFs additionally induce PUMA and Noxa. Importantly, p53^R172P MEFs failed to downregulate anti-apoptotic protein Bcl-2, which has been shown to play an important role in p53-dependent apoptosis. Taken together, these data demonstrate that in the absence of p53-mediated apoptosis, cells undergo cellular senescence to prevent genomic instability. Our results also indicate that p53-dependent apoptosis may play an active role in balancing cellular growth.

Open Access Article

Overexpression of the RNA binding protein HuR impairs tumor growth in triple negative breast cancer associated with deficient angiogenesis

Matthew M Gubin, Robert Calaluce, Justin Wade Davis, Joseph D Magee, Connie S. Strouse, Daniel P. Shaw, Lixin Ma, Ashley Brown, Timothy Hoffman, Tammy L. Rold and Ulus Atasoy
Pages 3337 - 3346
http://dx.doi.org/10.4161/cc.9.16.12711
Abstract | PDF

Interactions between RNA binding proteins (RBPs) and genes are not well understood, especially in regulation of angiogenesis. The RBP HuR binds to the AU-rich (ARE) regions of labile mRNAs, facilitating their translation into protein and has been hypothesized to be a tumor-maintenance gene. Elevated levels of cytoplasmic HuR directly correlate with increased invasiveness and poor prognosis for many cancers, including those of the breast. HuR controls the expression of multiple genes involved in angiogenesis including VEGFa, HIF1a, and thrombospondin 1 (TSP1). We investigated the role of HuR in estrogen receptor negative (ER-) breast cancer. MDA-MB-231 cells with higher levels of HuR have alterations in cell cycle kinetics and faster growth. Unexpectedly, HuR overexpression significantly interfered with tumor growth in orthotopic mouse models. The putative mechanism seems to be an anti-angiogenetic effect by increasing expression of TSP1 but also surprisingly, down-regulation of VEGF, a target of HuR which it normally increases. Our findings reveal that HuR may be regulating a cluster of genes involved in blood vessel formation which controls tumor angiogenesis. An approach of modulating HuR levels may overcome limitations associated with monotherapies targeting tumor vessel formation.

Open Access Article

The expression of p53-target genes in the hypoxia-tolerant subterranean mole-rat is hypoxia-dependent and similar to expression patterns in solid tumors

Mark Band, Osnat Ashur-Fabian and Aaron Avivi
Pages 3347 - 3352
http://dx.doi.org/10.4161/cc.9.16.12712
Abstract | PDF

The tumor suppressor gene p53, in response to DNA damage/hypoxia, induces growth arrest and/or apoptosis. Inactivation of p53, by mutations and/or overexpression of the mdm2 gene, confers a selective advantage to tumor cells under hypoxic microenvironment during tumor progression. The mole rat, Spalax, spends its life underground at low-oxygen tensions and hence has developed a wide range of respiratory/molecular adaptations to hypoxic stress. We previously reported that the highly conserved p53 Arg(R)-174 is substituted by lysine (K) in Spalax, identical to a tumor- associated mutation. Functionality assays revealed that Spalax p53 and human R174K-mutated p53 were unable to induce human/Spalax apaf1, an apoptotic target gene, while over-activating the mdm2 gene. Moreover, cells transfected with human p53 underwent more extensive apoptosis (44.8%) as compared to Spalax p53 (23.2%) transfected cells. To support our hypothesis that the pattern of activity in Spalax is related to hypoxia-tolerance, we quantified apaf1 and mdm2 mRNA levels under normoxia (21% O₂), short-acute hypoxic stress (5 h at 6% O₂), and long-mild hypoxic insult (44 h at 10% O₂). Results were compared to those of rats under similar conditions.  Following hypoxia, Spalax apaf1 mRNA levels decreased significantly, but increased in rats. apip mRNA levels, a negative regulator of apaf1, increased in Spalax and decreased in rats. mdm2 mRNA levels under hypoxia were significantly higher in Spalax. We conclude that, similar to our previous in-vitro work, two parallel hypoxia-adaptive mechanisms evolved in Spalax: mutated p53 and p53 response element leading to a bias against apoptosis and increased mdm2, which are analogous to observations in tumor development.

Open Access Article

Deregulated Ras signaling compromises DNA damage checkpoint recovery in S. cerevisiae

Matthew D. Wood and Yolanda Sanchez
Pages 3353 - 3363
http://dx.doi.org/10.4161/cc.9.16.12713
Abstract | PDF

The DNA damage checkpoint maintains genome stability by arresting the cell cycle and promoting DNA repair under genotoxic stress.  Cells must downregulate the checkpoint signaling pathways in order to resume cell division after completing DNA repair.  While the mechanisms of checkpoint activation have been well-characterized, the process of checkpoint recovery, and the signals regulating it, has only recently been investigated.  We have identified a new role for the Ras signaling pathway as a regulator of DNA damage checkpoint recovery.  Here we report that in budding yeast, deletion of the IRA1 and IRA2 genes encoding negative regulators of Ras prevents cellular recovery from a DNA damage induced arrest.  The checkpoint kinase Rad53 is dephosphorylated in an IRA-deficient strain, indicating that recovery failure is not caused by constitutive checkpoint pathway activation.  The ira1D ira2D recovery defect requires the checkpoint kinase Chk1 and the cAMP-dependent protein kinase (PKA) catalytic subunit Tpk2.  Furthermore, PKA phosphorylation sites on the anaphase promoting complex specificity factor Cdc20 are required for the recovery defect, indicating a link between the recovery defect and PKA regulation of mitosis.  This work identifies a new signaling pathway that can regulate DNA damage checkpoint recovery, and implicates the Ras signaling pathway as an important regulator of mitotic events.

Open Access Article

Small-molecule inducer of cancer cell polyploidy promotes apoptosis or senescence: Implications for therapy

Christian Tovar, Brian Higgins, Dayanand Deo, Kenneth Kolinsky, Jin-Jun Liu, David C Heimbrook and Lyubomir T Vassilev
Pages 3364 - 3375
http://dx.doi.org/10.4161/cc.9.16.12732
Abstract | PDF

Polyploidy results from deregulated cell division and has been considered an undesirable event leading to increased mutation rate and cancer development.  However, polyploidy may also render cancer cells more vulnerable to chemotherapy.  Here, we identify a small-molecule inducer of polyploidy, R1530,  which interferes with tubulin polymerization and mitotic checkpoint function in cancer cells, leading to abortive mitosis, endoreduplication and polyploidy.  In the presence of R1530, polyploid cancer cells underwent apoptosis or became senescent which translated into potent in vitro and in vivo efficacy.  Normal proliferating cells were resistant to R1530-induced polyploidy thus supporting the rationale for cancer therapy by induced polyploidy.  Mitotic checkpoint kinase BubR1 was found downregulated during R1530-induced exit from mitosis, a likely consequence of PLK4 inhibition.  BubR1 knockdown in the presence of nocodazole induced an R1530-like phenotype, suggesting that BubR1 plays a key role in polyploidy induction by R1530 and could be exploited as a target for designing more specific polyploidy inducers.

Open Access Article

HdmX overexpression inhibits oncogene induced cellular senescence

Kelly R. Miller, Kevin Kelley, Rebecca Tuttle and Steven Berberich
Pages 3376 - 3382
http://dx.doi.org/10.4161/cc.9.16.12779
Abstract | PDF

Cellular senescence is an irreversible state of terminal growth arrest that requires functional p53.  Acting to block tumor formation, induction of senescence has also been demonstrated to contribute to tumor clearance via the immune system following p53 reactivation.^{1, 2} The Hdm2-antagonist, Nutlin-3a, has been shown to reactivate p53 and induce a quiescent state in various cancer cell lines,^{3, 4} similar to the G1 arrest observed upon RNAi targeting of Hdm2 in MCF7 breast cancer.⁵ In the present study we show that HdmX, a negative regulator of p53, impacts the senescence pathway.  Specifically, overexpression of HdmX blocks Ras mediated senescence in primary human fibroblasts.  The interaction of HdmX with p53 and the re-localization of HdmX to the nucleus through Hdm2 association appear to be required for this activity.  Furthermore, inhibiting HdmX in prostate adenocarcinoma cells expressing wild-type p53, mutant Ras and high levels of HdmX induced cellular senescence as measured by an increase in irreversible b-galactosidase staining.  Together these results suggest that HdmX overexpression may contribute to tumor formation by blocking senescence and that targeting HdmX may represent an attractive anti-cancer therapeutic approach.