Table of Contents

Volume 9, Issue 4

  February 15, 2010

  Pages 623 - 849

View covers archive

 Issue Catalogue (MARC XML)

EDITORIALS: CELL CYCLE FEATURES

Open Access Article

The TGFβ-Par6 polarity pathway: Linking the Par complex to EMT and breast cancer progression

Alicia M. Viloria-Petit and Jeffrey L. Wrana
Pages 623 - 624
http://dx.doi.org/10.4161/cc.9.4.10779
Abstract | PDF

Comment on: Viloria-Petit AM, et al. Proc Natl Acad Sci USA 2009;106:14028-33.

Open Access Article

Relationship of GAD₆₇regulation to cell cycle and DNA repair in GABA neurons in the adult hippocampus: Bipolar disorder versus schizophrenia

Francine M. Benes
Pages 625 - 627
http://dx.doi.org/10.4161/cc.9.4.10820
Abstract | PDF

Benes FM, et al. Proc Natl Acad Sci USA 2009;106:11731-6.

Open Access Article

The “stem” of chemoresistance

Giulia Bertolini, Laura Gatti and Luca Roz
Pages 628 - 629
http://dx.doi.org/10.4161/cc.9.4.10821
Abstract | PDF

Comment on: Bertolini G, et al. Proc Natl Acad Sci USA 2009; 106:16281-6.

Open Access Article

RNA polymerase II stalling mediates cytokine gene expression

Karen Adelman and Inez Rogatsky
Pages 630 - 631
http://dx.doi.org/10.4161/cc.9.4.10841
Abstract | PDF

Comment on: Adelman K, et al. Proc Natl Acad Sci USA 2009; 106:18207-12.

Open Access Article

Changing pace: Viral mimicry of an anaphase promoting complex subunit

Min Mo, Stephen B. Fleming and Andrew A. Mercer
Pages 632 - 633
http://dx.doi.org/10.4161/cc.9.4.10842
Abstract | PDF

Comment on: Mo M, et al. Proc Natl Acad Sci USA 2009; 106:19527-32.

Open Access Article

Converting intestine into esophagus: Anterior-posterior patterning of the vertebrate gut requires Cdx2

Klaus H. Kaestner
Pages 634 - 635
http://dx.doi.org/10.4161/cc.9.4.10929
Abstract | PDF

Comment on: Gao N, et al. Dev Cell 2009; 16:588-99.

Open Access Article

Adult neurogenesis as a cellular model to study schizophrenia

Kimberly Christian, Hongjun Song and Guo-li Ming
Pages 636 - 637
http://dx.doi.org/10.4161/cc.9.4.10932
Abstract | PDF

Comment on: Kim JY, et al. Neuron 2009; 63:761-73.

Open Access Article

A new function for old drugs

Riku Das and Edward F. Plow
Pages 638 - 639
http://dx.doi.org/10.4161/cc.9.4.11016
Abstract | PDF

Das R, et al. Circ Res 2009; 105:167-75.

Open Access Article

Uncovering a unique role for DNA methylation in hematopoietic and leukemic stem cells

Lena Vockentanz, Ann-Marie Bröske and Frank Rosenbauer
Pages 640 - 641
http://dx.doi.org/10.4161/cc.9.4.11037
Abstract | PDF

Comment on: Bröske AM, et al. Nat Genet 2009; In press.




NEWS & VIEWS

Open Access Article

Will helicases contribute to the fight against malaria?

Mohamed Abdelhaleem
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10823
Abstract | PDF

Comment on: Tuteja R. Cell cycle 2010; 9: In press.

Open Access Article

The NF-Y angle to the CCAAT's tale

Ernest Martinez
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10828
Abstract | PDF

Comment on: Dolfini D, et al. Cell Cycle 2009; 8: 4127-37.

Open Access Article

Cycling mouse oocytes through meiosis

Ian R. Adams
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10830
Abstract | PDF

Comment on: Miles DC, et al. Cell Cycle 2010; 9:In press.

Open Access Article

Spindle assembly checkpoint inactivation: A new role for phosphatases

Jeffrey R. Skaar and Michele Pagano
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10831
Abstract | PDF

Comment on: Visconti R, et al. Cell Cycle 2010; 9:In press.

Open Access Article

New messages in the nuclear envelope

William T. Dauer and Howard Worman
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10844
Abstract | PDF

Comment on: Malhas A, et al. Cell Cycle 2010; 9:In press.

Open Access Article

Hedgehog takes a new “RHOad” to angiogenesis

Martin E. Fernandez-Zapico
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10854
Abstract | PDF

Comment on: Chinchilla P, et al. Cell Cycle 2010: 9:In press.

Open Access Article

A direct, non-canonical pathway for Hedgehog proteins in the endothelium

Qingshi Zhao, Amanda Beck and Diego Fraidenraich
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10855
Abstract | PDF

Comment on: Chinchilla P, et al. Cell Cycle 2010: 9:In press.

Open Access Article

p63 and canonical Wnt signaling

Flaminia Talos, Ramona Schulz and Ute Moll
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10858
Abstract | PDF

Comment on: Drewelus I, et al. Cell Cycle 2010; 9(3); In press.

Open Access Article

New insights into KLF8-mediated transactivation

Paul M. Evans and Chunming Liu
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10859
Abstract | PDF

Comment on: Uvalek AM, et al. Cell Cycle 2010; 9(3): In press.

Open Access Article

KLF8 sets the pace for the cell cycle through interactions with p300 and PCAF

Joyce A. Lloyd
Pages 642 - 651
http://dx.doi.org/10.4161/cc.9.4.10860
Abstract | PDF

Comment on: Urvalek et al. Cell Cycle 2010; 9(3): In press.




EXTRA VIEWS

Open Access Article

Activation of epithelial STAT3 regulates intestinal homeostasis

Clemens Neufert, Geethanjali Pickert, Yan Zheng, Nadine Wittkopf, Moritz Warntjen, Alexei Nikolae, Wenjun Ouyang, Markus F. Neurath and Christoph Becker
Pages 652 - 655
http://dx.doi.org/10.4161/cc.9.4.10615
Abstract | PDF

The intestinal epithelium that lines the mucosal surface along the GI-tract is a key player for the intestinal homeostasis of the healthy individual. In case of a mucosal damage or a barrier defect as seen in patients with inflammatory bowel disease, the balance is disturbed, and translocation of intestinal microbes to the submucosa is facilitated. We recently demonstrated a pivotal role of STAT3 activation in intestinal epithelial cells (IEC) for the restoration of the balance at the mucosal surface of the gut in an experimental colitis model. STAT3 was rapidly induced in intestinal epithelial cells upon challenge of mice in both experimental colitis and intestinal wound healing models. STAT3 activation was found to be dispensable in the steady-state conditions but was important for efficient regeneration of the epithelium in response to injury. Here, we extend our previous findings by showing epithelial STAT3 activation in human patients suffering from IBD and provide additional insights how the activation of epithelial STAT3 by IL-22 regulates intestinal homeostasis and mucosal wound healing. We also demonstrate that antibody-mediated neutralization of IL-22 has little impact on the development of experimental colitis in mice, but significantly delays recovery from colitis. Thus, our data suggest that targeting the STAT3 signalling pathway in IEC is a promising therapeutic approach in situations when the intestinal homeostasis is disturbed, e.g. as seen in Crohn´s disease or Ulcerative colitis.

Open Access Article

Role of thrombin as a tumor growth factor

David Green and Simon Karpatkin
Pages 656 - 661
http://dx.doi.org/10.4161/cc.9.4.10729
Abstract | PDF

The clinical observation that thrombosis in some patients heralds the onset of malignancy has been recognized for over a century. Thrombin the key terminal enzyme of coagulation also promotes angiogenesis and stimulates tumor-platelet adhesion, adhesion to endothelium, tumor implantation, tumor cell growth and metastasis. The thrombin receptor, a member of the protease-activated receptor family, is expressed on many tumor cell lines and on breast tumor biopsy specimens. In addition to mitogenic effects on fibroblast, smooth muscle cells and endothelial cells, thrombin also exerts direct effects on cancer cells by activation of the cell cycle through down-regulation of p27Kip1and induction of Skp2, and cyclins D and A. MicroRNA 222, which inhibits p27Kip1 , is upregulated by thrombin. In the transgenic TRAMP mouse model of prostate cancer inhibition of endogenous thrombin by hirudin retards spontaneous tumor growth. Inhibition of thrombin may lead to tumor dormancy and could explain inhibition of tumor growth and metastasis by anticoagulants observed in animal models and a beneficial effect on survival observed in some clinical trials of anticoagulants in cancer patients.

Open Access Article

γH2AX foci analysis for monitoring DNA double-strand break repair: Strengths, limitations and optimization

Markus Löbrich, Atsushi Shibata, Andrea Beucher, Anna Fisher, Michael Ensminger, Aaron A. Goodarzi, Olivia Barton and Penny A. Jeggo
Pages 662 - 669
http://dx.doi.org/10.4161/cc.9.4.10764
Abstract | PDF

DNA double-strand breaks (DSBs) represent an important radiation-induced lesion and impaired DSB repair provides the best available correlation with radiosensitivity. Physical techniques for monitoring DSB repair require high, non-physiological doses and cannot reliably detect subtle defects. One outcome from extensive research into the DNA damage response is the observation that H2AX, a variant form of the histone H2A, undergoes extensive phosphorylation at the DSB, creating γH2AX foci that can be visualised by immunofluorescence. There is a close correlation between γH2AX foci and DSB numbers and between the rate of foci loss and DSB repair, providing a sensitive assay to monitor DSB repair in individual cells using physiological doses. However, γH2AX formation can occur at single-stranded DNA regions which arise during replication or repair and thus does not solely correlate with DSB formation. Here, we present and discuss evidence that following exposure to ionising radiation, γH2AX foci analysis can provide a sensitive monitor of DSB formation and repair and describe techniques to optimise the analysis. We discuss the limitations and benefits of the technique, enabling the procedure to be optimally exploited but not misused.

Open Access Article

Mutations in a gene encoding a midbody protein in binucleated Reed-Sternberg cells of Hodgkin lymphoma

Maxwell M. Krem, Stephen J. Salipante and Marshall S. Horwitz
Pages 670 - 675
http://dx.doi.org/10.4161/cc.9.4.10780
PDF

Open Access Article

Unexpected deeds of familiar proteins: Interplay of Hsp70, PGRP S / Tag7, and S100A4 / Mts1 in host vs. cancer combat

Elena A. Dukhanina, Denis V. Yashin, Alexander V. Galkin and Lidia P. Sashchenko
Pages 676 - 682
http://dx.doi.org/10.4161/cc.9.4.10782
Abstract | PDF

We consider the novel means of attack and defense in the host versus cancer combat that involve interactions between widespread multifunctional proteins, focusing on the aspects that may seem paradoxical in the framework of established notions. Particularly, we show that a protein broadly known for its protective functions such as Hsp70 can make a tumoricidal “binary weapon” with another nontoxic protein Tag7 (PGRP-S); that the same Hsp70, a ubiquitous intracellular chaperone, when expressed on the MHC-negative tumor cell surface, can itself be the hallmark of immune evasion rather than a primordial MHC substitute; that a device functionally equivalent to the T-cell receptor (Tag7-Centered Recognizer) can be assembled of components in no way related to the classical pathways of T-cell-mediated immunity, and operate where the orthodox immunosurveillance fails; and that one and the same protein Mts1 (S100A4) under different circumstances may work as “reactive armor” of a tumor cell against humoral agents and as a vital part of the T-cell machinery aimed against immunoevasive cells, i.e., perform both prometastatic and antimetastatic functions.




PERSPECTIVES

Open Access Article

Calorie restriction: Decelerating mTOR-driven aging from cells to organisms (including humans)

Mikhail V. Blagosklonny
Pages 683 - 688
http://dx.doi.org/10.4161/cc.9.4.10766
Abstract | PDF

Although it has been known since 1917 that calorie restriction (CR) decelerates aging, the topic remains highly controversial.  What might be the reason? Here I discuss that the anti-aging effect of CR rules out accumulation of DNA damage and failure of maintenance as a cause of aging. Instead, it suggests that aging is driven in part by the nutrient-sensing TOR (target of rapamycin) network. CR deactivates the TOR pathway, thus slowing aging and delaying diseases of aging. Humans are not an exception and CR must increase both maximal and healthy lifespan in humans to the same degree as it does in other mammals. Unlike mice, however, humans benefit from medical care, which prolongs lifespan despite accelerated aging in non-restricted individuals. Therefore in humans the effect of CR may be somewhat blunted. Still how much does CR extend human lifespan? And could this extension be surpassed by gerosuppressants such as rapamycin?

Open Access Article

Rb inactivation in cell cycle and cancer: The puzzle of highly regulated activating phosphorylation of CDK4 versus constitutively active CDK-activating kinase

Sabine Paternot, Laurence Bockstaele, Xavier Bisteau, Hugues Kooken, Katia Coulonval and Pierre Roger
Pages 689 - 699
http://dx.doi.org/10.4161/cc.9.4.10611
Abstract | PDF

Cyclin-dependent kinase (CDK) 4 is a master integrator that couples mitogenic/oncogenic signalling cascades with the inactivation of the central oncosuppressor Rb and the cell cycle. Its activation requires binding to a D-type cyclin and then T-loop phosphorylation at T172 by the only identified CDK-activating kinase in animal cells, cyclin H-CDK7. In contrast with the observed constitutive activity of cyclin H-CDK7, we have recently identified the T172-phosphorylation of cyclin D-bound CDK4 as a crucial cell cycle regulatory target. Intriguingly, the homologous T177-phosphorylation of CDK6 is weak in several systems and does not present this regulation. In this Perspective, we review the recent advances and debates on the multistep mechanism leading to activation of D-type cyclin–CDK4 complexes. This involves a re-evaluation of the implication of Cip/Kip CDK “inhibitors” and CDK7 in this process.

Open Access Article

The many facets of the Tim-Tipin protein families’ roles in chromosome biology

Ramsay J. McFarlane, Saira Mian and Jacob Z. Dalgaard
Pages 700 - 705
http://dx.doi.org/10.4161/cc.9.4.10676
Abstract | PDF

Failures in DNA replication are a potent force for driving genome instability. The proteins which form the replisome, the DNA replication machinery, play a fundamental role in preventing replicative catastrophes. The Tim (TIMELESS/TIMEOUT) and Tipin proteins are two conserved replisome associated proteins which have functions in preventing replication fork collapse and replicative checkpoint signalling in response to factors which slow the progression of the replisome. Intriguingly, TIMELESS family members have been implicated in the regulation of the biological clock, giving a tantalising pointer to a possible link between DNA replication and circadian rhythm control. Here we report on our current understanding of the many facets of these protein families in maintaining genome stability and replication checkpoint control.

Open Access Article

Putting one step before the other: Distinct activation pathways for Cdk1 and Cdk2 bring order to the mammalian cell cycle

Karl A. Merrick and Robert P. Fisher
Pages 706 - 714
http://dx.doi.org/10.4161/cc.9.4.10732
Abstract | PDF

Eukaryotic cell division is controlled by the activity of cyclin-dependent kinases (CDKs). Cdk1 and Cdk2, which function at different stages of the mammalian cell cycle, both require cyclin-binding and phosphorylation of the activation (T-) loop for full activity, but differ with respect to the order in which the two steps occur in vivo. To form stable complexes with either of its partners-cyclins A and B-Cdk1 must be phosphorylated on its T-loop, but that phosphorylation in turn depends on the presence of cyclin. Cdk2 can follow a kinetically distinct path to activation in which T-loop phosphorylation precedes cyclin-binding, and thereby out-compete the more abundant Cdk1 for limiting amounts of cyclin A. Mathematical modeling suggests this could be a principal basis for the temporal ordering of CDK activation during S phase, which may dictate the sequence in which replication origins fire. Still to be determined are how: (1) the activation machinery discriminates between closely related CDKs, and (2) coordination of the cell cycle is affected when this mechanism of pathway insulation breaks down.

Open Access Article

Emerging functions of force-producing kinetochore motors

Yinghui Mao, Dileep Varma and Richard B Vallee
Pages 715 - 719
http://dx.doi.org/10.4161/cc.9.4.10763
Abstract | PDF

More than two decades of research has resulted in the identification of some 60 microtubule motor proteins, several of which have been implicated in mitosis.  Although some kinesin superfamily proteins function as microtubule depolymerases at kinetochores, such as Kinesin-8 and -13, it is now appreciated that there are only two force-producing kinetochore associated motors, the plus end-directed microtubule motor CENP-E and the minus end-directed microtubule motor cytoplasmic dynein.  Defining their roles at kinetochores has been hampered by the complexity of mitosis itself, and a multiplicity of mitotic roles, at least for cytoplasmic dynein.  Nonetheless, recent advances have served to define the primary roles of the two kinetochore motors in detail.




REVIEWS

Open Access Article

p73 tumor suppressor protein: A close relative of p53 not only in structure but also in anti-cancer approach?

Joanna Zawacka-Pankau, Anna Kostecka, Alicja Sznarkowska, Elisabeth Hedström and Anna Kawiak
Pages 720 - 728
http://dx.doi.org/10.4161/cc.9.4.10668
Abstract | PDF

The discovery of the p53 tumor suppressor protein in 1979 shed new light on cancer cell biology and introduced a trend in cancer research focusing on p53-like proteins. This in turn led to the discovery of two homologous proteins of p53 - p63 in 1998 and p73 in 1997.

The p53 family members are mainly involved in apoptosis induction under cellular stress, but also in early embryonic developmental processes. The p63 and p73 proteins activate the transcription of a number of p53 target genes. The precise role of p63 in cancer cells is not fully revealed yet, unlike that of p53 and p73.

The p53 tumor suppressor protein is found inactive in approximately 50% of human cancers. However, p73 is not as often inactivated in tumors. Of importance, transcriptionally active forms of p73 induce apoptosis in cancer cells independent of p53 status. Moreover, the regulatory mechanisms governing p73 stability in cells are well described. These features promoted the research concerning p73-targeted anti-cancer treatment.
The p73 protein is subject to sophisticated activatory and inhibitory regulatory mechanisms. The up-to-date anti-cancer compounds targeting p73 protein in vitro inhibit its negative regulators, which leads to the activation of p73 pro-apoptotic function in cancer cells. In the current review we present the recent scientific findings on p73 regulation in cells and the newest anti-cancer strategies concerning its tumor suppressor function.

Open Access Article

Multiple two-polymerase mechanisms in mammalian translesion DNA synthesis

Zvi Livneh, Omer Z and Sigal Shachar
Pages 729 - 735
http://dx.doi.org/10.4161/cc.9.4.10727
Abstract | PDF

The encounter of replication forks with DNA lesions may lead to fork arrest and/or the formation of single-stranded gaps. A major strategy to cope with these replication irregularities is translesion DNA replication (TLS), in which specialized error-prone DNA polymerases bypass the blocking lesions. Recent studies suggest that TLS across a particular DNA lesion may involve as many as four different TLS polymerases, acting in two-polymerase reactions in which insertion by a particular polymerase is followed by extension by another polymerase. Insertion determines the accuracy and mutagenic specificity of the TLS reaction, and is carried out by one of several polymerases such as polη, polκ or polι. In contrast, extension is carried out primarily by polζ. In cells from XPV patients, which are deficient in TLS across cyclobutane pyrimidine dimers (CPD) due to a deficiency in polη, TLS is carried out by at least two backup reactions each involving two polymerases: One reaction involves polκ and polζ, and the other polι and polζ. These mechanisms may also assist polη in normal cells under an excessive amount of UV lesions.




REPORTS

Open Access Article

Negative regulation of CHK2 activity by protein phosphatase 2A is modulated by DNA damage

Alyson K. Freeman, Virna Dapic and Alvaro N.A. Monteiro
Pages 736 - 747
http://dx.doi.org/10.4161/cc.9.4.10613
Abstract | PDF

Checkpoint kinase 2 (CHK2) is a major effector of the DNA damage response pathway and although its mechanism of activation has been well studied, the attenuation of its activity following DNA damage has not been explored. Here, we identify the B'α subunit of protein phosphatase 2A (PP2A) as a CHK2 binding partner and show that their interaction is modulated by DNA damage. B'α binds to the SQ/TQ repeat region of CHK2, which is a target of ATM phosphorylation. The induction of DNA double-strand breaks by gamma irradiation as well as treatment with doxorubicin causes dissociation of the B'α and CHK2 proteins. This dissociation correlates with an increase in the ATM-dependent phosphorylation of CHK2 at serines 33 and 35 in the SQ/TQ region. Indeed, mutating these sites to mimic phosphorylation increases the dissociation after irradiation. PP2A negatively regulates CHK2 phosphorylation at multiple sites, as well as its kinase activity. These data reveal a novel mechanism for PP2A to keep CHK2 inactive under normal conditions while also allowing for a rapid release from this regulation immediately following DNA damage. This is followed by a subsequent reconstitution of the PP2A/CHK2 complex in later time points after damage, which may help to attenuate the signal.

Open Access Article

The Mdm2 and p53 genes are conserved in the Arachnids

David P. Lane, Chit Fang Cheok, Christopher J. Brown, Arumugam Madhumalar, Farid J. Ghadessy and Chandra Verma
Pages 748 - 754
http://dx.doi.org/10.4161/cc.9.4.10616
Abstract | PDF

The p53 protein and its negative regulator the ubiquitin E3 ligase Mdm2 have been shown to be conserved from the Placazoan to man. In common with D.melanogaster and C.elegans, there is a single copy of the p53 gene in T.adhaerens, while in the vertebrates three p53-like genes can be found: p53 , p63 and p73. The Mdm2 gene is not present within the fully sequenced and highly annotated genomes of C.elegans and D.melanogaster. However, it is present in the Placazoan and the presence of multiple distinct p53 genes in the Sea anemone N.vectensis led us to examine the genomes of other phyla for p53 and Mdm2-like genes. We report here the discovery of an Mdm2-like gene and two distinct p53 like genes in the Arachnid Ioxodes scapularis (Northern Deer Tick). The two predicted Deer Tick p53 proteins are much more highly related to the human p53 protein in sequence than are the fruit fly and nematode proteins. One of the Deer tick genes encodes a p53 protein that is initiated within the DNA binding domain of p53 and shows remarkable homology to the newly described N-terminally truncated delta isoforms of human and zebrafish p53.

Open Access Article

DNA damage response links calpain to cellular senescence

Francesca Demarchi, Francesca Cataldo, Cosetta Bertoli and Claudio Schneider
Pages 755 - 760
http://dx.doi.org/10.4161/cc.9.4.10637
Abstract | PDF

Senescence represents an important barrier against cellular transformation. Here we show that CAPNS1 depletion impairs senescence induction both in BJ-ET H-Ras^v12 inducible human fibroblasts upon Ras induction and in HT1080 targeted to senescence by treatment with low doses of doxorubicin. We further show that CAPNS1 depletion is coupled to reduced levels of H2AX phosphorylation, not only in Ras^v12 induced BJ-ET fibroblasts, but also in a number of cellular systems upon genotoxic stress. In particular CAPNS1 depletion affects γ-H2AX appearance or persistence in U2OS osteosarcoma cells 24 hours after MMC addition or UV light exposure; HT1080 upon camptothecin treatment for 4 hours and 48 hours after addition of MMC; MDA-MB-231, 24 hours after UV light exposure and 2 hours after bleomycin addition. Overall this study unveils a novel link between calpain, cellular senescence and DNA damage response.

Open Access Article

Biological impact of freezing Plk1 in its inactive conformation in cancer cells

Sarah Keppner, Ewgenij Proschak, Manfred Kaufmann, Klaus Strebhardt, Gisbert Schneider and Birgit Spänkuch
Pages 761 - 774
http://dx.doi.org/10.4161/cc.9.4.10644
Abstract | PDF

Human polo-like kinase 1 (Plk1), a key regulator of mitosis, is over-expressed in various human tumors. It is a negative prognostic factor for cancer patients and a measure for the aggressiveness of a tumor. Thus, targeting Plk1 might be a promising approach for cancer therapy. Kinase inhibitors are divided in type I inhibitors, targeting the highly conserved active conformation, and the more selective type II inhibitors, targeting the inactive conformation of kinases. We analyzed our previously identified type II Plk1 inhibitor SBE13 which is able to inhibit Plk1 activity. To determine its ability to induce cell death in cancer cells, we applied kinase assays, Western blot analyses, FACScan analyses, Caspase assays and immunofluorescence studies. We detected decreased cell proliferation, delayed progression through the cell cycle in lower SBE13 concentrations, a G2/M arrest using higher SBE13 concentrations followed by apoptosis, and abnormal mitotic figures. Notably, SBE13 did not influence activity of other kinases (Plk2, Plk3, Aurora A), indicating the selectivity of this type II Plk1 inhibitor. This study suggests that Plk1 kinase inhibitors targeting the inactive conformation of Plk1 may be considered for the development of cancer therapeutics.

Open Access Article

CEP215 is involved in the dynein-dependent accumulation of pericentriolar matrix proteins for spindle pole formation

Seongju Lee and Kunsoo Rhee
Pages 775 - 784
http://dx.doi.org/10.4161/cc.9.4.10667
Abstract | PDF

CEP215 is a human orthologue of Drosophila centrosomin which is a core centrosome component for the pericentriolar matrix protein recruitment. Recent investigations revealed that CEP215 is required for centrosome cohesion, centrosomal attachment of the g-TuRC, and microtubule dynamics. However, it remains to be obscure how CEP215 functions for recruitment of the centrosomal proteins during the centrosome cycle. Here, we investigated a role of CEP215 during mitosis. Knockdown of CEP215 resulted in characteristic mitotic phenotypes, including monopolar spindle formation, a decrease in distance between the spindle pole pair, and detachment of the centrosomes from the spindle poles. We noticed that CEP215 is critical for centrosomal localization of dynein throughout the cell cycle. As a consequence, the selective centrosomal proteins were not recruited to the centrosome properly. Finally, the centrosomal localization of CEP215 also depends on the dynein-dynactin complex. Based on the results, we propose that CEP215 regulates a dynein-dependent transport of the pericentriolar matrix proteins during the centrosome maturation.

Open Access Article

Role of endogenous Annexin-A1 in the regulation of thymocyte positive and negative selection

Nikolaos Paschalidis, Antony Huggins, Nicola J. Rowbotham, Anna L. Furmanski, Tessa Crompton, Roderick J. Flower, Mauro Perretti and Fulvio D’Acquisto
Pages 785 - 794
http://dx.doi.org/10.4161/cc.9.4.10673
Abstract | PDF

We have recently shown that endogenous Annexin-A1 (AnxA1) plays a homeostatic regulatory role in mature T cells by modulating the strength of TCR signaling. In this study we investigated the role of endogenous AnxA1 in thymocyte maturation. Analysis of AnxA1^-/- thymocyte populations at the immature CD4-CD8- double negative (DN) stage showed a proportional decrease in the DN1 and an increase in the DN3 subsets compared to control littermates. There were no significant differences in thymocyte numbers or proportions of CD4⁺ and CD8⁺ single positive (SP) populations between Anx1^-/- and AnxA1^+/+ mice. However, when we crossed AnxA1^-/- mice onto HY-TCR transgenic mice, we observed an increase in CD4⁺CD8⁺ double positive (DP) and CD4 SP cells in male AnxA1-/-/HY-TCR compared to AnxA1^+/+/HY-TCR. Conversely, female AnxA1^-/-/HY-TCR mice showed an increase in DP and a decrease in CD8 (SP) cells compared to female AnxA1^+/+/HY-TCR. Biochemical analysis of the signaling pathways responsible for these effects showed a decrease in anti-CD3-induced Erk phosphorylation and NF-κB activation in AnxA1^-/- thymocytes compared to control littermates. Together these findings demonstrate a role for endogenous AnxA1 in regulating both positive and negative selection of the TCR repertoire. These results suggest that targeting AnxA1 expression or function in T cells could represent a useful approach for the development of novel therapies for the treatment of autoimmune diseases.

Open Access Article

Comparative and functional analysis of the archaeal cell cycle

Rolf Bernander, Magnus Lundgren and Thijs J. G. Ettema
Pages 795 - 806
http://dx.doi.org/10.4161/cc.9.4.10674
Abstract | PDF

The temporal and spatial organization of the chromosome replication, genome segregation and cell division processes is less well understood in species belonging to the Archaea, than in those from the Bacteria and Eukarya domains. Novel insights into the regulation and key components of the Sulfolobus acidocaldarius cell cycle have been obtained through genome-wide analysis of cell-cycle-specific gene expression, followed by cloning and characterization of gene products expressed at different cell cycle stages. Here, the results of the transcript profiling are further explored, and potential key players in archaeal cell cycle progression are highlighted in an evolutionary context, by comparing gene expression patterns and gene conservation between three selected microbial species from different domains of life. We draw attention to novel putative nucleases and helicases implicated in DNA replication, recombination and repair, as well as to potential genome segregation factors. Focus is also placed upon regulatory features, including transcription factors and protein kinases inferred to be involved in the execution of specific cell cycle stages, and regulation through metabolic coupling is discussed.

Open Access Article

Transient increases in p53-responsible gene expression at early stages of Epstein-Barr virus productive replication

Yoshitaka Sato, Noriko Shirata, Takayuki Murata, Sho Nakasu, Ayumi Kudoh, Satoko Iwahori, Sanae Nakayama, Shigeki Chiba, Hiroki Isomura, Teru Kanda and Tatsuya Tsurumi
Pages 807 - 814
http://dx.doi.org/10.4161/cc.9.4.10675
Abstract | PDF

Expression of Epstein-Barr Virus BZLF1, a key protein initiating the switch from latent to lytic infection, is known to cause cell growth arrest by accumulating p53 and p21^WAF1/CIP1 in epithelial cells, but its molecular mechanism remains elusive. We found here that the BZLF1 protein stimulates p53 binding to its recognition sequence. The BZLF1 accelerated the rate of p53-DNA complex formation through the interaction with p53 protein and also enhanced p53-specific transcription in vitro. Furthermore, p53 protein was found to bind to its target promoter regions specifically in the early stages of lytic replication. Overexpression of p53 at the early stages of lytic replication enhanced viral genome replication, supporting the idea that p53 plays an important role in the initiation steps of EBV replication. Taking the independent role of BZLF1 on p53 degradation into consideration, we propose that the BZLF1 protein regulates p53 and its target gene products in two distinctive manners; transient induction of p53 at the early stages for the initiation of viral productive replication and p53 degradation at the later stages for S-phase like environment preferable for viral replication.

Open Access Article

Study of DNA replication in Drosophila using cell free in vitro system

Anton Svitin and Igor Chesnokov
Pages 815 - 819
http://dx.doi.org/10.4161/cc.9.4.10730
Abstract | PDF

Using Drosophila early egg extracts we have developed an optimized cell free system to study DNA replication. The efficiency of replication depends on a cold treatment of Drosophila embryos before the extract preparation and a formation of nuclei facilitated by the addition of membrane fractions to the extracts. In vitro DNA replication is ORC and CDC6 dependent, as a removal of these proteins from the extracts abolishes DNA replication. The N-terminal part of Orc1 protein, which is important for non-replicative functions of ORC, is dispensable for the replication in vitro. We also show that the conserved ATPase motif of CDC6 is crucial for the replication. Our studies indicate that a Drosophila cell free system proves to be an extremely useful tool for a functional dissection of the processes and factors involved in DNA replication in metazoans.

Open Access Article

Cyclin D3 is dispensable for human diffuse large B-cell lymphoma survival and growth: evidence for redundancy with cyclin E

Maria R. Gumina, Chenjia Xu and Thomas C. Chiles
Pages 820 - 828
http://dx.doi.org/10.4161/cc.9.4.10783
Abstract | PDF

Genomic changes disrupting the expression of cyclin D3 are associated with aberrant growth of several human B-lymphoid malignancies. We demonstrate that the human diffuse large B-cell lymphoma (DLBCL), OCI-LY18 (LY18) expresses cyclin D3 but not cyclins D1 and D2. RNA interference was used to deplete cyclin D3 from LY18 cells. Surprisingly, knockdown of cyclin D3 did not inhibit pRb phosphorylation on cdk4/6- and cdk2-specific residues or measurably affect viability and proliferation. These results suggest that cyclin D3 is dispensable in LY18 cell proliferation and survival. Similar results were obtained following depletion of cyclin E. By contrast, combined knockdown of cyclins D3 and E had substantial consequences leading to G1-phase arrest and inhibition of proliferation. Whereas cell cycle distribution was not affected following individual depletion of cdk4, cdk6, or cdk2, the combined knockdown of cdk4 and cdk6 led to accumulation of LY18 cells in G1-phase of the cell cycle and inhibition of proliferation. Likewise treatment of LY18 cells with 2-Bromo-12,13-dihydro-5H-indolo[2,3-a]pyrrolo[3,4-c]carbazole-5,7(6H)-dione, a selective inhibitor of cdk4/6, led to inhibition of proliferation. Taken together, these results uncover a built-in redundancy with cyclins D3 and E for G1-S progression. Moreover these findings highlight the rationale for simultaneous disruption of cdk4/6 as a potential therapeutic cancer strategy.

Open Access Article

Amino terminal hydrophobic import signals target the p14ARF tumor suppressor to the mitochondria

Mal Irvine, Suzanah Philipsz, Monika Frausto, Branka Mijato, Stuart J. Gallagher, Carina Fung, Therese M. Becker, Richard F. Kefford and Helen Rizos
Pages 829 - 839
http://dx.doi.org/10.4161/cc.9.4.10785
Abstract | PDF

The p14ARF tumour suppressor is frequently targeted for inactivation in many human cancers and in individuals predisposed to cutaneous melanoma. The functions of p14ARF are closely linked with its subcellular distribution. Nucleolar p14ARF dampens ribosome biosynthesis and nucleoplasmic forms of p14ARF activate the p53 pathway and induce cell cycle arrest. p14ARF can also be recruited to mitochondria where it interacts with many mitochondrial proteins, including Bcl-xL and p32 to induce cell death. It has been suggested that the movement of p14ARF to mitochondria requires its interaction with p32, but we now show that the ARF-p32 interaction is not necessary for the accumulation of p14ARF in mitochondria. Instead, highly hydrophobic domains within the amino-terminal half of p14ARF act as mitochondrial import sequences. We suggest that once this hydrophobic pocket is exposed, possibly in a stimulus-dependent manner, it accelerates the mitochondrial import of p14ARF. This allows the interaction of p14ARF with mitochondrial proteins, including p32 and enables p53-independent cell death.

Open Access Article

26S proteasome exhibits endoribonuclease activity controlled by extra-cellular stimuli

Valentina A. Kulichkova, Olga A. Fedorova, Anna S. Tsimokha, Tatiana N. Moiseeva, Andrew Bottril, Larissa Lezina, Larissa N. Gauze, Irina M. Konstantinova, Alexey G. Mittenberg and Nickolai A. Barlev
Pages 840 - 849
http://dx.doi.org/10.4161/cc.9.4.10829
Abstract | PDF

26S proteasome is a large multi-subunit protein complex involved in proteolytic degradation of proteins. In addition to its canonical proteolytic activity, the proteasome is also associated with recently characterized endoribonuclease (endo-RNAse) activity. However, neither functional significance, nor the mechanisms of its regulation are currently known. In this report, we show that 26S proteasome is able to hydrolyze various cellular RNAs, including AU-rich mRNA of c-myc and c-fos. The endonucleolytic degradation of these mRNAs is exerted by one of the 26S proteasome subunits, PSMA5 (α5). The RNAse activity of 26S proteasome is differentially affected by various extra-cellular signals. Moreover, this activity contributes to the process of degradation of c-myc mRNA during induced differentiation of K562 cells, and may be controlled by phosphorylation of the adjacent subunits, PSMA1 (α6) and PSMA3 (α7). Collectively, the data presented in this report suggest a causal link between cell signalling pathways, endo-RNAse activity of the 26S proteasome complex and metabolism of cellular RNAs.