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Sample records for checkpoint kinase-dependent rad26

  1. The effect of ataxia-telangiectasia mutated kinase-dependent hyperphosphorylation of checkpoint kinase-2 on oligodeoxynucleotide 7909 containing CpG motifs-enhanced sensitivity to X-rays in human lung adenocarcinoma A549 cells

    Directory of Open Access Journals (Sweden)

    Liu XQ

    2015-06-01

    Full Text Available Xiaoqun Liu,1,* Xiangdong Liu,2,* Tiankui Qiao,1 Wei Chen,1 Sujuan Yuan1 1Department of Oncology, 2Department of Ophthalmology, Affiliated Jinshan Hospital, Fudan University, Shanghai, People’s Republic of China *These authors contributed equally to this work Objective: The aim of the study reported here was to further investigate the potential effect of ataxia-telangiectasia mutated (ATM kinase-dependent hyperphosphorylation of checkpoint kinase-2 (Chk2 on radiosensitivity enhanced by oligodeoxynucleotide 7909 containing CpG motifs (CpG ODN7909 in human lung adenocarcinoma A549 cells. Methods: In vitro A549 cells were randomly separated into control, CpG, X-ray, CpG+X-ray, ATM kinase-small interfering RNA (siRNA+CpG+X-ray (ATM-siRNA, and Chk2-siRNA+CpG+X-ray (Chk2-siRNA groups. siRNAs were adopted to silence the ATM and Chk2 genes. Expression and phosphorylation of ATM kinase and Chk2 were detected by Western blot assay. Cell colonies were observed under inverted phase-contrast microscopy. Cellular survival curves were fitted using a multi-target single-hitting model. Cell cycle and apoptosis were analyzed by flow cytometry. Results: Expression of ATM kinase and Chk2 was similar among the control, CpG, X-ray, and CpG+X-ray groups. Phosphorylated ATM kinase and Chk2 were significantly increased in the CpG+X-ray group compared with in the X-ray group (t=6.00, P<0.01 and t=3.13, P<0.05, respectively, though these were hardly detected in the control and CpG groups. However, expression of ATM kinase and Chk2 was clearly downregulated in the ATM-siRNA and Chk2-siRNA groups, respectively. Similarly, their phosphorylation levels were also significantly decreased in the ATM-siRNA group (t=14.35, P<0.01 and t=8.46, P<0.01, respectively and a significant decrease in phosphorylated Chk2 was observed in the Chk2-siRNA group (t=7.28, P<0.01 when compared with the CpG+X-ray group. Further, the number of A549 cells at Gap 2/mitotic phase and the apoptosis

  2. Tfb5 Is Partially Dispensable for Rad26 Mediated Transcription Coupled Nucleotide Excision Repair in Yeast

    OpenAIRE

    Ding, Baojin; Ruggiero, Christine; Chen, Xuefeng; Li, Shisheng

    2007-01-01

    Nucleotide excision repair (NER) is a conserved DNA repair mechanism capable of removing a variety of helix-distorting DNA lesions. A specialized NER pathway, called transcription coupled NER (TC-NER), refers to preferential repair in the transcribed strand of an actively transcribed gene. To be distinguished from TCR-NER, the genome-wide NER process is termed as global genomic NER (GG-NER). In Saccharomyces cerevisiae, GG-NER is dependent on Rad7, whereas TC-NER is mediated by Rad26, the hom...

  3. Immune checkpoints

    OpenAIRE

    Chawla, Akhil; Philips, Anne V; Alatrash, Gheath; Mittendorf, Elizabeth,

    2014-01-01

    Early clinical trials investigating monoclonal antibodies targeting the T-cell inhibitory receptor programmed cell death 1 (PD-1) and its ligand PD-L1 have shown efficacy in melanoma, non-small cell lung cancer and renal cell carcinoma. We recently demonstrated PD-L1 expression in 20% of triple negative breast cancers suggesting that targeting the PD-1/PD-L1 immune checkpoint may be an effective treatment modality in patients with this disease.

  4. Checkpoint control and cancer

    Czech Academy of Sciences Publication Activity Database

    Medema, R.H.; Macůrek, Libor

    2012-01-01

    Roč. 31, č. 21 (2012), s. 2601-2613. ISSN 0950-9232 R&D Projects: GA ČR GAP301/10/1525; GA ČR GPP305/10/P420 Institutional support: RVO:68378050 Keywords : DNA damage * checkpoint control * anticancer strategies Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.357, year: 2012

  5. Checkpoint Kinases Regulate a Global Network of Transcription Factors in Response to DNA Damage

    Directory of Open Access Journals (Sweden)

    Eric J. Jaehnig

    2013-07-01

    Full Text Available DNA damage activates checkpoint kinases that induce several downstream events, including widespread changes in transcription. However, the specific connections between the checkpoint kinases and downstream transcription factors (TFs are not well understood. Here, we integrate kinase mutant expression profiles, transcriptional regulatory interactions, and phosphoproteomics to map kinases and downstream TFs to transcriptional regulatory networks. Specifically, we investigate the role of the Saccharomyces cerevisiae checkpoint kinases (Mec1, Tel1, Chk1, Rad53, and Dun1 in the transcriptional response to DNA damage caused by methyl methanesulfonate. The result is a global kinase-TF regulatory network in which Mec1 and Tel1 signal through Rad53 to synergistically regulate the expression of more than 600 genes. This network involves at least nine TFs, many of which have Rad53-dependent phosphorylation sites, as regulators of checkpoint-kinase-dependent genes. We also identify a major DNA damage-induced transcriptional network that regulates stress response genes independently of the checkpoint kinases.

  6. Checkpoint inhibition in meningiomas.

    Science.gov (United States)

    Bi, Wenya Linda; Wu, Winona W; Santagata, Sandro; Reardon, David A; Dunn, Ian F

    2016-06-01

    Meningiomas are increasingly appreciated to share similar features with other intra-axial central nervous system neoplasms as well as systemic cancers. Immune checkpoint inhibition has emerged as a promising therapy in a number of cancers, with durable responses of years in a subset of patients. Several lines of evidence support a role for immune-based therapeutic strategies in the management of meningiomas, especially high-grade subtypes. Meningiomas frequently originate juxtaposed to venous sinuses, where an anatomic conduit for lymphatic drainage resides. Multiple populations of immune cells have been observed in meningiomas. PD-1/PD-L1 mediated immunosuppression has been implicated in high-grade meningiomas, with association between PD-L1 expression with negative prognostic outcome. These data point to the promise of future combinatorial therapeutic strategies in meningioma. PMID:27197540

  7. Toward an Optimal Online Checkpoint Solution under a Two-Level HPC Checkpoint Model

    OpenAIRE

    Di, Sheng; Robert, Yves; Vivien, Frédéric; Cappello, Franck

    2016-01-01

    The traditional single-level checkpointing method suffers from significantoverhead on large-scale platforms. Hence, multilevel checkpointing protocols have been studied extensively in recent years. The multilevel checkpoint approach allows different levels of checkpoints to be set (each with different checkpoint overheads and recovery abilities), in order to further improve the fault tolerance performance of extreme-scale HPC applications. How to optimize the checkpoint intervals for each lev...

  8. Efficient Incremental Checkpointing of Java Programs

    DEFF Research Database (Denmark)

    Lawall, Julia Laetitia; Muller, Gilles

    2000-01-01

    This paper investigates the optimization of language-level checkpointing of Java programs. First, we describe how to systematically associate incremental checkpoints with Java classes. While being safe, the genericness of this solution induces substantial execution overhead. Second, to solve...

  9. Cambridge checkpoint English workbook 1

    CERN Document Server

    Reynolds, John

    2013-01-01

    This Workbook supports our bestselling Checkpoint English series, with exercises specifically matched to the Cambridge Progression tests and the Checkpoint English tests. - Offers plenty of additional questions for use in class or as homework. - Includes clearly identified questions on grammar and punctuation, comprehension, use of language and essay planning. - Follows the structure of the relevant textbook to ensure a thorough understanding of all aspects of the course. - Provides a space for Students to write their answers. This Workbook is matched to the Cambridge Secondary 1 Curriculum Fr

  10. Network support for system initiated checkpoints

    Science.gov (United States)

    Chen, Dong; Heidelberger, Philip

    2013-01-29

    A system, method and computer program product for supporting system initiated checkpoints in parallel computing systems. The system and method generates selective control signals to perform checkpointing of system related data in presence of messaging activity associated with a user application running at the node. The checkpointing is initiated by the system such that checkpoint data of a plurality of network nodes may be obtained even in the presence of user applications running on highly parallel computers that include ongoing user messaging activity.

  11. Checkpointing for a hybrid computing node

    Science.gov (United States)

    Cher, Chen-Yong

    2016-03-08

    According to an aspect, a method for checkpointing in a hybrid computing node includes executing a task in a processing accelerator of the hybrid computing node. A checkpoint is created in a local memory of the processing accelerator. The checkpoint includes state data to restart execution of the task in the processing accelerator upon a restart operation. Execution of the task is resumed in the processing accelerator after creating the checkpoint. The state data of the checkpoint are transferred from the processing accelerator to a main processor of the hybrid computing node while the processing accelerator is executing the task.

  12. FAK dimerization controls its kinase-dependent functions at focal adhesions

    KAUST Repository

    Brami-Cherrier, Karen

    2014-01-30

    Focal adhesion kinase (FAK) controls adhesion-dependent cell motility, survival, and proliferation. FAK has kinase-dependent and kinase-independent functions, both of which play major roles in embryogenesis and tumor invasiveness. The precise mechanisms of FAK activation are not known. Using x-ray crystallography, small angle x-ray scattering, and biochemical and functional analyses, we show that the key step for activation of FAK\\'s kinase-dependent functions-autophosphorylation of tyrosine-397-requires site-specific dimerization of FAK. The dimers form via the association of the N-terminal FERM domain of FAK and are stabilized by an interaction between FERM and the C-terminal FAT domain. FAT binds to a basic motif on FERM that regulates co-activation and nuclear localization. FAK dimerization requires local enrichment, which occurs specifically at focal adhesions. Paxillin plays a dual role, by recruiting FAK to focal adhesions and by reinforcing the FAT:FERM interaction. Our results provide a structural and mechanistic framework to explain how FAK combines multiple stimuli into a site-specific function. The dimer interfaces we describe are promising targets for blocking FAK activation. © 2014 The Authors.

  13. DNA damage checkpoint recovery and cancer development

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Haiyong [First affiliated hospital, Zhejiang University, School of medicine, Cancer Center, 79 Qingchun Road, Hangzhou 310003 (China); Zhang, Xiaoshan [Department of Genetics, University of Texas M.D. Anderson Cancer Center, Department of Genetics Unit 1010, 1515 Holcombe Blvd. Houston, TX 77030 (United States); Teng, Lisong, E-mail: lsteng@zju.edu.cn [First affiliated hospital, Zhejiang University, School of medicine, Cancer Center, 79 Qingchun Road, Hangzhou 310003 (China); Legerski, Randy J., E-mail: rlegersk@mdanderson.org [Department of Genetics, University of Texas M.D. Anderson Cancer Center, Department of Genetics Unit 1010, 1515 Holcombe Blvd. Houston, TX 77030 (United States)

    2015-06-10

    Cell cycle checkpoints were initially presumed to function as a regulator of cell cycle machinery in response to different genotoxic stresses, and later found to play an important role in the process of tumorigenesis by acting as a guard against DNA over-replication. As a counterpart of checkpoint activation, the checkpoint recovery machinery is working in opposition, aiming to reverse the checkpoint activation and resume the normal cell cycle. The DNA damage response (DDR) and oncogene induced senescence (OIS) are frequently found in precancerous lesions, and believed to constitute a barrier to tumorigenesis, however, the DDR and OIS have been observed to be diminished in advanced cancers of most tissue origins. These findings suggest that when progressing from pre-neoplastic lesions to cancer, DNA damage checkpoint barriers are overridden. How the DDR checkpoint is bypassed in this process remains largely unknown. Activated cytokine and growth factor-signaling pathways were very recently shown to suppress the DDR and to promote uncontrolled cell proliferation in the context of oncovirus infection. In recent decades, data from cell line and tumor models showed that a group of checkpoint recovery proteins function in promoting tumor progression; data from patient samples also showed overexpression of checkpoint recovery proteins in human cancer tissues and a correlation with patients' poor prognosis. In this review, the known cell cycle checkpoint recovery proteins and their roles in DNA damage checkpoint recovery are reviewed, as well as their implications in cancer development. This review also provides insight into the mechanism by which the DDR suppresses oncogene-driven tumorigenesis and tumor progression. - Highlights: • DNA damage checkpoint works as a barrier to cancer initiation. • DDR machinary response to genotoxic and oncogenic stress in similar way. • Checkpoint recovery pathways provide active signaling in cell cycle control. • Checkpoint

  14. DNA damage checkpoint recovery and cancer development

    International Nuclear Information System (INIS)

    Cell cycle checkpoints were initially presumed to function as a regulator of cell cycle machinery in response to different genotoxic stresses, and later found to play an important role in the process of tumorigenesis by acting as a guard against DNA over-replication. As a counterpart of checkpoint activation, the checkpoint recovery machinery is working in opposition, aiming to reverse the checkpoint activation and resume the normal cell cycle. The DNA damage response (DDR) and oncogene induced senescence (OIS) are frequently found in precancerous lesions, and believed to constitute a barrier to tumorigenesis, however, the DDR and OIS have been observed to be diminished in advanced cancers of most tissue origins. These findings suggest that when progressing from pre-neoplastic lesions to cancer, DNA damage checkpoint barriers are overridden. How the DDR checkpoint is bypassed in this process remains largely unknown. Activated cytokine and growth factor-signaling pathways were very recently shown to suppress the DDR and to promote uncontrolled cell proliferation in the context of oncovirus infection. In recent decades, data from cell line and tumor models showed that a group of checkpoint recovery proteins function in promoting tumor progression; data from patient samples also showed overexpression of checkpoint recovery proteins in human cancer tissues and a correlation with patients' poor prognosis. In this review, the known cell cycle checkpoint recovery proteins and their roles in DNA damage checkpoint recovery are reviewed, as well as their implications in cancer development. This review also provides insight into the mechanism by which the DDR suppresses oncogene-driven tumorigenesis and tumor progression. - Highlights: • DNA damage checkpoint works as a barrier to cancer initiation. • DDR machinary response to genotoxic and oncogenic stress in similar way. • Checkpoint recovery pathways provide active signaling in cell cycle control. • Checkpoint

  15. Visualizing the spindle checkpoint in Drosophila spermatocytes

    OpenAIRE

    Rebollo, Elena; González, Cayetano

    2000-01-01

    The spindle assembly checkpoint detects defects in spindle structure or in the alignment of the chromosomes on the metaphase plate and delays the onset of anaphase until defects are corrected. Thus far, the evidence regarding the presence of a spindle checkpoint during meiosis in male Drosophila has been indirect and contradictory. On the one hand, chromosomes without pairing partners do not prevent meiosis progression. On the other hand, some conserved components of the spindle checkpoint ma...

  16. GRID COMPUTING AND CHECKPOINT APPROACH

    Directory of Open Access Journals (Sweden)

    Pankaj gupta

    2011-05-01

    Full Text Available Grid computing is a means of allocating the computational power of alarge number of computers to complex difficult computation or problem. Grid computing is a distributed computing paradigm thatdiffers from traditional distributed computing in that it is aimed toward large scale systems that even span organizational boundaries. In this paper we investigate the different techniques of fault tolerance which are used in many real time distributed systems. The main focus is on types of fault occurring in the system, fault detection techniques and the recovery techniques used. A fault can occur due to link failure, resource failure or by any other reason is to be tolerated for working the system smoothly and accurately. These faults can be detected and recovered by many techniques used accordingly. An appropriate fault detector can avoid loss due to system crash and reliable fault tolerance technique can save from system failure. This paper provides how these methods are applied to detect and tolerate faults from various Real Time Distributed Systems. The advantages of utilizing the check pointing functionality are obvious; however so far the Grid community has notdeveloped a widely accepted standard that would allow the Gridenvironment to consciously utilize low level check pointing packages.Therefore, such a standard named Grid Check pointing Architecture isbeing designed. The fault tolerance mechanism used here sets the jobcheckpoints based on the resource failure rate. If resource failureoccurs, the job is restarted from its last successful state using acheckpoint file from another grid resource. A critical aspect for anautomatic recovery is the availability of checkpoint files. A strategy to increase the availability of checkpoints is replication. Grid is a form distributed computing mainly to virtualizes and utilize geographically distributed idle resources. A grid is a distributed computational and storage environment often composed of

  17. Overlapped checkpointing with hardware assist

    Energy Technology Data Exchange (ETDEWEB)

    Mitchell, Christopher J [Los Alamos National Laboratory; Nunez, James A [Los Alamos National Laboratory; Wang, Jun [U. OF CENTRAL FLORIDA (UCF)

    2009-01-01

    We present a new approach to handling the demanding I/O workload incurred during checkpoint writes encountered in High Performance Computing. Prior efforts to improve performance have been primarily bound by mechanical limitations of the hard drive. Our research surpasses this limitation by providing a method to: (1) write checkpoint data to a high-speed, non-volatile buffer, and (2) asynchronously write this data to permanent storage while resuming computation. This removes the hard drive from the critical data path because our I/O node based buffers isolate the compute nodes from the storage servers. This solution is feasible because of industry declines in cost for high-capacity, non-volatile storage technologies. Testing was conducted on a small-scale cluster to prove the design, and then scaled at Los Alamos National Laboratory. Results show a definitive speedup factor for select workloads over writing directly to a typical global parallel file system; the Panasas ActiveScale File System.

  18. Kinase signaling in the spindle checkpoint.

    Science.gov (United States)

    Kang, Jungseog; Yu, Hongtao

    2009-06-01

    The spindle checkpoint is a cell cycle surveillance system that ensures the fidelity of chromosome segregation. In mitosis, it elicits the "wait anaphase" signal to inhibit the anaphase-promoting complex or cyclosome until all chromosomes achieve bipolar microtubule attachment and align at the metaphase plate. Because a single kinetochore unattached to microtubules activates the checkpoint, the wait anaphase signal is thought to be generated by this kinetochore and is then amplified and distributed throughout the cell to inhibit the anaphase-promoting complex/cyclosome. Several spindle checkpoint kinases participate in the generation and amplification of this signal. Recent studies have begun to reveal the activation mechanisms of these checkpoint kinases. Increasing evidence also indicates that the checkpoint kinases not only help to generate the wait anaphase signal but also actively correct kinetochore-microtubule attachment defects. PMID:19228686

  19. Visualizing the spindle checkpoint in Drosophila spermatocytes

    Science.gov (United States)

    Rebollo, Elena; González, Cayetano

    2000-01-01

    The spindle assembly checkpoint detects defects in spindle structure or in the alignment of the chromosomes on the metaphase plate and delays the onset of anaphase until defects are corrected. Thus far, the evidence regarding the presence of a spindle checkpoint during meiosis in male Drosophila has been indirect and contradictory. On the one hand, chromosomes without pairing partners do not prevent meiosis progression. On the other hand, some conserved components of the spindle checkpoint machinery are expressed in these cells and behave as their homologue proteins do in systems with an active spindle checkpoint. To establish whether the spindle checkpoint is active in Drosophila spermatocytes we have followed meiosis progression by time-lapse microscopy under conditions where the checkpoint is likely to be activated. We have found that the presence of a relatively high number of misaligned chromosomes or a severe disruption of the meiotic spindle results in a significant delay in the time of entry into anaphase. These observations provide the first direct evidence substantiating the activity of a meiotic spindle checkpoint in male Drosophila. PMID:11256627

  20. REVIEW OF CHECKPOINTING ALGORITHMS IN DISTRIBUTED SYSTEMS

    Directory of Open Access Journals (Sweden)

    Poonam Gahlan

    2010-06-01

    Full Text Available Checkpointing is the process of saving the status information. Checkpoint is defined as a designated place in a program at which normal processing is interrupted specifically to preserve the status information necessary to allow resumption of processing at a later time. Mobile computing raises many new issues such as lack of stablestorage, low bandwidth of wireless channel, high mobility, and limited battery life. Coordinated checkpointing is an attractive approach for transparently adding fault tolerance to distributed applications since it avoids domino effects and minimizes the stable storage requirement. This paper presents the review of the algorithms,which have been reported in the literature for checkpointing. This paper also covers backward error recovery techniques for distributed systems specially the distributed mobile systems.

  1. A Nonblocking Coordinated Checkpointing Algorithm for Mobile Computing Systems

    Directory of Open Access Journals (Sweden)

    Rachit Garg

    2010-05-01

    Full Text Available A checkpoint algorithm for mobile computing systems needs to handle many new issues like: mobility, low bandwidth of wireless channels, lack of stable storage on mobile nodes, disconnections, limited battery power and high failure rate of mobile nodes. These issues make traditional checkpointing techniques unsuitable for such environments. Minimum-process coordinated checkpointing is an attractive approach to introduce fault tolerance in mobile distributed systems transparently. This approach is domino-free, requires at most two checkpoints of a process on stable storage, and forces only a minimum number of processes to checkpoint. But, it requires extra synchronization messages, blocking of the underlying computation or taking some useless checkpoints. In this paper, we propose a nonblocking coordinated checkpointing algorithm for mobile computing systems, which requires only a minimum number of processes to take permanent checkpoints. We reduce the message complexity as compared to the Cao-Singhal algorithm [4], while keeping the number of useless checkpoints unchanged. We also address the related issues like: failures during checkpointing, disconnections, concurrent initiations of the algorithm and maintaining exact dependencies among processes. Finally, the paper presents an optimization technique, which significantly reduces the number of useless checkpoints at the cost of minor increase in the message complexity. In coordinated checkpointing, if a single process fails to take its tentative checkpoint; all the checkpoint effort is aborted. We try to reduce this effort by taking soft checkpoints in the first phase at Mobile Hosts.

  2. Regulation of p73 by Hck through kinase-dependent and independent mechanisms

    Directory of Open Access Journals (Sweden)

    Radha Vegesna

    2007-05-01

    Full Text Available Abstract Background p73, a p53 family member is a transcription factor that plays a role in cell cycle, differentiation and apoptosis. p73 is regulated through post translational modifications and protein interactions. c-Abl is the only known tyrosine kinase that phosphorylates and activates p73. Here we have analyzed the role of Src family kinases, which are involved in diverse signaling pathways, in regulating p73. Results Exogenously expressed as well as cellular Hck and p73 interact in vivo. In vitro binding assays show that SH3 domain of Hck interacts with p73. Co-expression of p73 with Hck or c-Src in mammalian cells resulted in tyrosine phosphorylation of p73. Using site directed mutational analysis, we determined that Tyr-28 was the major site of phosphorylation by Hck and c-Src, unlike c-Abl which phosphorylates Tyr-99. In a kinase dependent manner, Hck co-expression resulted in stabilization of p73 protein in the cytoplasm. Activation of Hck in HL-60 cells resulted in tyrosine phosphorylation of endogenous p73. Both exogenous and endogenous Hck localize to the nuclear as well as cytoplasmic compartment, just as does p73. Ectopically expressed Hck repressed the transcriptional activity of p73 as determined by promoter assays and semi-quantitative RT-PCR analysis of the p73 target, Ipaf and MDM2. SH3 domain- dependent function of Hck was required for its effect on p73 activity, which was also reflected in its ability to inhibit p73-mediated apoptosis. We also show that Hck interacts with Yes associated protein (YAP, a transcriptional co-activator of p73, and shRNA mediated knockdown of YAP protein reduces p73 induced Ipaf promoter activation. Conclusion We have identified p73 as a novel substrate and interacting partner of Hck and show that it regulates p73 through mechanisms that are dependent on either catalytic activity or protein interaction domains. Hck-SH3 domain-mediated interactions play an important role in the inhibition of p73

  3. EGFR kinase-dependent and kinase-independent roles in clear cell renal cell carcinoma.

    Science.gov (United States)

    Cossu-Rocca, Paolo; Muroni, Maria R; Sanges, Francesca; Sotgiu, Giovanni; Asunis, Anna; Tanca, Luciana; Onnis, Daniela; Pira, Giovanna; Manca, Alessandra; Dore, Simone; Uras, Maria G; Ena, Sara; De Miglio, Maria R

    2016-01-01

    Epidermal growth factor receptor (EGFR) is associated with progression of many epithelial malignancies and represents a significant therapeutic target. Although clear cell renal cell carcinoma (CCRCC) has been widely investigated for EGFR molecular alterations, genetic evidences of EGFR gene activating mutations and/or gene amplification have been rarely confirmed in the literature. Therefore, until now EGFR-targeted therapies in clinical trials have been demonstrated unsuccessful. New evidence has been given about the interactions between EGFR and the sodium glucose co-transporter-1 (SGLT1) in maintaining the glucose basal intracellular level to favour cancer cell growth and survival; thus a new functional role may be attributed to EGFR, regardless of its kinase activity. To define the role of EGFR in CCRCC an extensive investigation of genetic changes and functional kinase activities was performed in a series of tumors by analyzing the EGFR mutational status and expression profile, together with the protein expression of downstream signaling pathways members. Furthermore, we investigated the co-expression of EGFR and SGLT1 proteins and their relationships with clinic-pathological features in CCRCC. EGFR protein expression was identified in 98.4% of CCRCC. Furthermore, it was described for the first time that SGLT1 is overexpressed in CCRCC (80.9%), and that co-expression with EGFR is appreciable in 79.4% of the tumours. Moreover, the activation of downstream EGFR pathways was found in about 79.4% of SGLT1-positive CCRCCs. The mutational status analysis of EGFR failed to demonstrate mutations on exons 18 to 24 and the presence of EGFR-variantIII (EGFRvIII) in all CCRCCs analyzed. FISH analysis revealed absence of EGFR amplification, and high polysomy of chromosome 7. Finally, the EGFR gene expression profile showed gene overexpression in 38.2% of CCRCCs. Our study contributes to define the complexity of EGFR role in CCRCC, identifying its bivalent kinase-dependent

  4. Asynchronous Checkpoint Migration with MRNet in the Scalable Checkpoint / Restart Library

    Energy Technology Data Exchange (ETDEWEB)

    Mohror, K; Moody, A; de Supinski, B R

    2012-03-20

    Applications running on today's supercomputers tolerate failures by periodically saving their state in checkpoint files on stable storage, such as a parallel file system. Although this approach is simple, the overhead of writing the checkpoints can be prohibitive, especially for large-scale jobs. In this paper, we present initial results of an enhancement to our Scalable Checkpoint/Restart Library (SCR). We employ MRNet, a tree-based overlay network library, to transfer checkpoints from the compute nodes to the parallel file system asynchronously. This enhancement increases application efficiency by removing the need for an application to block while checkpoints are transferred to the parallel file system. We show that the integration of SCR with MRNet can reduce the time spent in I/O operations by as much as 15x. However, our experiments exposed new scalability issues with our initial implementation. We discuss the sources of the scalability problems and our plans to address them.

  5. Checkpoint triggering in a computer system

    Energy Technology Data Exchange (ETDEWEB)

    Cher, Chen-Yong

    2016-09-06

    According to an aspect, a method for triggering creation of a checkpoint in a computer system includes executing a task in a processing node of the computer system and determining whether it is time to read a monitor associated with a metric of the task. The monitor is read to determine a value of the metric based on determining that it is time to read the monitor. A threshold for triggering creation of the checkpoint is determined based on the value of the metric. Based on determining that the value of the metric has crossed the threshold, the checkpoint including state data of the task is created to enable restarting execution of the task upon a restart operation.

  6. Garbage Collection in Uncoordinated Checkpointing Algorithms

    Institute of Scientific and Technical Information of China (English)

    LIU Yunlong; CHEN Junliang

    1999-01-01

    In this paper, the hard problem of thethorough garbage collection in uncoordinated checkpointing algorithms isstudied. After introduction of the traditional garbage collectingscheme, with which only obsolete checkpoints can be discarded, it isshown that this kind of traditional method may fail to discard anycheckpoint in some special cases, and it is necessary and urgent to finda thorough garbage collecting method, with which all the checkpointsuseless for any future rollback-recovery including the obsolete ones canbe discarded. Then, the Thorough Garbage Collection Theorem is proposedand proved, which ensures the feasibility of the thorough garbagecollection, and gives the method to calculate the set of the usefulcheckpoints as well.

  7. Checkpointing and Recovery in Distributed and Database Systems

    Science.gov (United States)

    Wu, Jiang

    2011-01-01

    A transaction-consistent global checkpoint of a database records a state of the database which reflects the effect of only completed transactions and not the results of any partially executed transactions. This thesis establishes the necessary and sufficient conditions for a checkpoint of a data item (or the checkpoints of a set of data items) to…

  8. Targeting the Checkpoint to Kill Cancer Cells

    Czech Academy of Sciences Publication Activity Database

    Benada, Jan; Macůrek, Libor

    2015-01-01

    Roč. 6, č. 3 (2015), s. 1912-1937. ISSN 2218-273X R&D Projects: GA ČR(CZ) GA14-34264S Institutional support: RVO:68378050 Keywords : checkpoint * DNA damage response * cancer Subject RIV: EB - Genetics ; Molecular Biology

  9. VDAC electronics: 3. VDAC-Creatine kinase-dependent generation of the outer membrane potential in respiring mitochondria.

    Science.gov (United States)

    Lemeshko, Victor V

    2016-07-01

    Mitochondrial energy in cardiac cells has been reported to be channeled into the cytosol through the intermembrane contact sites formed by the adenine nucleotide translocator, creatine kinase and VDAC. Computational analysis performed in this study showed a high probability of the outer membrane potential (OMP) generation coupled to such a mechanism of energy channeling in respiring mitochondria. OMPs, positive inside, calculated at elevated concentrations of creatine are high enough to restrict ATP release from mitochondria, to significantly decrease the apparent Km,ADP for state 3 respiration and to maintain low concentrations of Ca(2+) in the mitochondrial intermembrane space. An inhibition by creatine of Ca(2+)-induced swelling of isolated mitochondria and other protective effects of creatine reported in the literature might be explained by generated positive OMP. We suggest that VDAC-creatine kinase-dependent generation of OMP represents a novel physiological factor controlling metabolic state of mitochondria, cell energy channeling and resistance to death. PMID:27085978

  10. Replication licensing and the DNA damage checkpoint

    OpenAIRE

    Cook, Jeanette Gowen

    2009-01-01

    Accurate and timely duplication of chromosomal DNA requires that replication be coordinated with processes that ensure genome integrity. Significant advances in determining how the earliest steps in DNA replication are affected by DNA damage have highlighted some of the mechanisms to establish that coordination. Recent insights have expanded the relationship between the ATM and ATR-dependent checkpoint pathways and the proteins that bind and function at replication origins. These findings sug...

  11. Immune Checkpoint Inhibitors in Older Adults.

    Science.gov (United States)

    Elias, Rawad; Morales, Joshua; Rehman, Yasser; Khurshid, Humera

    2016-08-01

    Cancer is primarily a disease of older adults. The treatment of advanced stage tumors usually involves the use of systemic agents that may be associated with significant risk of toxicity, especially in older patients. Immune checkpoint inhibitors are newcomers to the oncology world with improved efficacy and better safety profiles when compared to traditional cytotoxic drugs. This makes them an attractive treatment option. While there are no elderly specific trials, this review attempts to look at the current available data from a geriatric oncology perspective. We reviewed data from phase III studies that led to newly approved indications of checkpoint inhibitors in non-small cell lung cancer, melanoma, and renal cell cancer. Data were reviewed with respect to response, survival, and toxicity according to three groups: 75 years. Current literature does not allow one to draw definitive conclusions regarding the role of immune checkpoint inhibitors in older adults. However, they may offer a potentially less toxic but equally efficacious treatment option for the senior adult oncology patient. PMID:27287329

  12. An Anti-Checkpoint Activity for Rif1

    OpenAIRE

    Xue, Yuan; Rushton, Michael D.; Maringele, Laura

    2011-01-01

    Cells accumulate single-stranded DNA (ssDNA) when telomere capping, DNA replication, or DNA repair is impeded. This accumulation leads to cell cycle arrest through activating the DNA–damage checkpoints involved in cancer protection. Hence, ssDNA accumulation could be an anti-cancer mechanism. However, ssDNA has to accumulate above a certain threshold to activate checkpoints. What determines this checkpoint-activation threshold is an important, yet unanswered question. Here we identify Rif1 (R...

  13. A Monitor for Bud Emergence in the Yeast Morphogenesis Checkpoint

    OpenAIRE

    Theesfeld, Chandra L.; Zyla, Trevin R.; Bardes, Elaine G.S.; Lew, Daniel J.

    2003-01-01

    Cell cycle transitions are subject to regulation by both external signals and internal checkpoints that monitor satisfactory progression of key cell cycle events. In budding yeast, the morphogenesis checkpoint arrests the cell cycle in response to perturbations that affect the actin cytoskeleton and bud formation. Herein, we identify a step in this checkpoint pathway that seems to be directly responsive to bud emergence. Activation of the kinase Hsl1p is dependent upon...

  14. Anaphase onset before complete DNA replication with intact checkpoint responses

    DEFF Research Database (Denmark)

    Torres-Rosell, Jordi; De Piccoli, Giacomo; Cordon-Preciado, Violeta;

    2007-01-01

    Cellular checkpoints prevent mitosis in the presence of stalled replication forks. Whether checkpoints also ensure the completion of DNA replication before mitosis is unknown. Here, we show that in yeast smc5-smc6 mutants, which are related to cohesin and condensin, replication is delayed, most...

  15. Immune checkpoint inhibition in lymphoid disease.

    Science.gov (United States)

    Eyre, Toby A; Collins, Graham P

    2015-08-01

    It has long been understood that the immune system has intrinsic anti-tumour activity in humans, and that a key mechanism of tumour progression is the ability of a tumour to escape this immune surveillance. A number of attempts have been made to harness this anti-tumour immunity in both solid tumour oncology and haematological malignancies with variable success. Examples include the use of allogeneic stem cell transplantation and donor lymphocyte infusion in haematological cancer and vaccine studies in solid tumours. Enhanced signalling of the Programmed cell death-1 (PDCD1, PD-1)/cytotoxic T-lymphocyte-associated protein 4 (CTLA4) 'immune checkpoint' pathway has emerged recently as a critical mechanism by which tumours can escape the natural anti-tumour immune response. As such, novel therapies have been developed to help enhance this natural immunity by switching off the PDCD1/CTLA4 immune checkpoint pathway. The following review will discuss the pathobiology of these pathways and the exciting new data now available in lymphoid malignancies. PMID:25824455

  16. Developmental checkpoints guarded by regulated necrosis.

    Science.gov (United States)

    Dillon, Christopher P; Tummers, Bart; Baran, Katherine; Green, Douglas R

    2016-06-01

    The process of embryonic development is highly regulated through the symbiotic control of differentiation and programmed cell death pathways, which together sculpt tissues and organs. The importance of programmed necrotic (RIPK-dependent necroptosis) cell death during development has recently been recognized as important and has largely been characterized using genetically engineered animals. Suppression of necroptosis appears to be essential for murine development and occurs at three distinct checkpoints, E10.5, E16.5, and P1. These distinct time points have helped delineate the molecular pathways and regulation of necroptosis. The embryonic lethality at E10.5 seen in knockouts of caspase-8, FADD, or FLIP (cflar), components of the extrinsic apoptosis pathway, resulted in pallid embryos that did not exhibit the expected cellular expansions. This was the first suggestion that these factors play an important role in the inhibition of necroptotic cell death. The embryonic lethality at E16.5 highlighted the importance of TNF engaging necroptosis in vivo, since elimination of TNFR1 from casp8 (-/-), fadd (-/-), or cflar (-/-), ripk3 (-/-) embryos delayed embryonic lethality from E10.5 until E16.5. The P1 checkpoint demonstrates the dual role of RIPK1 in both the induction and inhibition of necroptosis, depending on the upstream signal. This review summarizes the role of necroptosis in development and the genetic evidence that helped detail the molecular mechanisms of this novel pathway of programmed cell death. PMID:27056574

  17. Immune checkpoints in cancer clinical trials

    Institute of Scientific and Technical Information of China (English)

    Elad Sharon; Howard Streicher; Priscila Goncalves; Helen XChen

    2014-01-01

    Immunology-based therapy is rapidly developing into an effective treatment option for a surprising range of cancers. We have learned over the last decade that powerful immunologic effector cells may be blocked by inhibitory regulatory pathways controlled by specific molecules often called“immune checkpoints.” These checkpoints serve to control or turn off the immune response when it is no longer needed to prevent tissue injury and autoimmunity. Cancer cells have learned or evolved to use these mechanisms to evade immune control and elimination. The development of a new therapeutic class of drugs that inhibit these inhibitory pathways has recently emerged as a potent strategy in oncology. Three sets of agents have emerged in clinical trials exploiting this strategy. These agents are antibody-based therapies targeting cytotoxic T-lymphocyte antigen4 (CTLA4), programmed cell death1 (PD-1), and programmed cell death ligand 1 (PD-L1). These inhibitors of immune inhibition have demonstrated extensive activity as single agents and in combinations. Clinical responses have been seen in melanoma, renal cellcarcinoma, non-smal celllung cancer, and several other tumor types. Despite the autoimmune or inflammatory immune-mediated adverse effects which have been seen, the responses and overall survival benefits exhibited thus far warrant further clinical development.

  18. Checkpoint kinase 1 negatively regulates somatic hypermutation.

    Science.gov (United States)

    Frankenberger, Samantha; Davari, Kathrin; Fischer-Burkart, Sabine; Böttcher, Katrin; Tomi, Nils-Sebastian; Zimber-Strobl, Ursula; Jungnickel, Berit

    2014-04-01

    Immunoglobulin (Ig) diversification by somatic hypermutation in germinal center B cells is instrumental for maturation of the humoral immune response, but also bears the risk of excessive or aberrant genetic changes. Thus, introduction of DNA damage by activation-induced cytidine deaminase as well as DNA repair by multiple pathways need to be tightly regulated during the germinal center response to prevent lymphomagenesis. In the present study, we show that DNA damage checkpoint signaling via checkpoint kinase 1 (Chk1) negatively regulates somatic hypermutation. Chk1 inhibition in human B cell lymphoma lines as well as inactivation of Chk1 alleles by gene targeting in DT40 B cells leads to increased somatic hypermutation. This is apparently due to changes in DNA repair pathways regulated by Chk1, such as a decreased homologous recombination efficiency that also leads to decreased Ig gene conversion in DT40. Our data show that Chk1 signaling plays a crucial role in regulation of Ig diversification and sheds unexpected light on potential origins of aberrant somatic hypermutation in B cell lymphomagenesis. PMID:24423870

  19. DMTCP: Scalable User-Level Transparent Checkpointing for Cluster Computations

    CERN Document Server

    Ansel, Jason; Arya, Kapil

    2008-01-01

    As the size of clusters increases, failures are becoming increasingly frequent. Applications must become fault tolerant if they are to run for extended periods of time. We present DMTCP (Distributed MultiThreaded CheckPointing), the first user-level distributed checkpointing package not dependent on a specific message passing library. This contrasts with existing approaches either specific to libraries such as MPI or requiring kernel modification. DMTCP provides fault tolerance through checkpointing. DMTCP transparently checkpoints general cluster computations consisting of many nodes, processes, and threads. DMTCP automatically accounts for TCP/IP sockets, UNIX domain sockets, pipes, ptys (pseudo-terminals), signal handlers, ordinary file descriptors, shared file descriptors, and other operating system artifacts. We demonstrate checkpointing and restart of applications communicating through MPICH2, OpenMPI, and sockets directly. These applications were written with a variety of languages including Fortran, C...

  20. Epidermal growth factor induces Ca(2+) sensitization through Rho-kinase-dependent phosphorylation of myosin phosphatase target subunit 1 in vascular smooth muscle.

    Science.gov (United States)

    Sasahara, Tomoya; Okamoto, Hiroshi; Ohkura, Natsumi; Kobe, Asami; Yayama, Katsutoshi

    2015-09-01

    We previously found that the protein tyrosine phosphatase inhibitor orthovanadate evoked a vasoconstrictor effect in rat aortas via Rho-kinase-dependent inactivation of myosin light chain phosphatase (MLCP) downstream of epidermal growth factor (EGF) receptor signaling. To determine whether the direct activation of EGF receptor by EGF also induces Rho-kinase-dependent vasoconstriction, isometric tension changes were measured in rat aortic rings without endothelium. Although EGF did not produce a contractile effect, the Ca(2+)-induced force in Ca(2+)-depleted rings significantly increased after treatment with 100nM EGF, suggesting that EGF induces Ca(2+) sensitization by MLCP inactivation. In addition, EGF induced the activation of Rho-kinase and phosphorylation of myosin phosphatase target subunit 1 (MYPT1) in rat aortic smooth muscle cells (VSMCs). The effects of EGF on Ca(2+) sensitivity in aortas and MYPT1 phosphorylation in VSMCs were blocked by inhibitors of EGF receptor (AG1478), Rho-kinase (Y27632), extracellular signal-regulated kinase 1/2 (Erk1/2; FR180204), and mitogen/extracellular signal-regulated kinase (MEK; PD98059), but not by inhibitors of p38 kinase (SB203580) and c-Jun amino-terminal kinase (AS601245). EGF-induced Erk1/2 phosphorylation was not abrogated by the Rho-kinase inhibitor, suggesting that Rho-kinase-dependent phosphorylation of MYPT1 is downstream of EGF receptor/MEK/Erk1/2 signaling. These results suggest that EGF induces Ca(2+) sensitization in vascular smooth muscle by Rho-kinase-dependent inactivation of MLCP mediated by the EGF receptor/MEK/Erk1/2 pathway. PMID:26004531

  1. FORMAL VERIFICATION OF DISTRIBUTED CHECKPOINTING USING EVENT-B

    Directory of Open Access Journals (Sweden)

    Girish Chandra

    2015-10-01

    Full Text Available The development of complex system makes challenging task for correct software development. Due to faulty specification, software may involve errors. The traditional testing methods are not sufficient to verify the correctness of such complex system. In order to capture correct system requirements and rigorous reasoning about the problems, formal methods are required. Formal methods are mathematical techniques that provide precise specification of problems with their solutions and proof of correctness. In this paper, we have done formal verification of check pointing process in a distributed database system using Event B. Event-B is an event driven formal method which is used to develop formal models of distributed database systems. In a distributed database system, the database is stored at different sites that are connected together through the network. Checkpoint is a recovery point which contains the state information about the site. In order to do recovery of a distributed transaction a global checkpoint number (GCPN is required. A global checkpoint number decides which transaction will be included for recovery purpose. All transactions whose timestamp are less than global checkpoint number will be marked as before checkpoint transaction (BCPT and will be considered for recovery purpose. The transactions whose timestamp are greater than GCPN will be marked as after checkpoint transaction (ACPT and will be part of next global checkpoint number.

  2. Keeping checkpoint/restart viable for exascale systems.

    Energy Technology Data Exchange (ETDEWEB)

    Riesen, Rolf E.; Bridges, Patrick G. (IBM Research, Ireland, Mulhuddart, Dublin); Stearley, Jon R.; Laros, James H., III; Oldfield, Ron A.; Arnold, Dorian (University of New Mexico, Albuquerque, NM); Pedretti, Kevin Thomas Tauke; Ferreira, Kurt Brian; Brightwell, Ronald Brian

    2011-09-01

    Next-generation exascale systems, those capable of performing a quintillion (10{sup 18}) operations per second, are expected to be delivered in the next 8-10 years. These systems, which will be 1,000 times faster than current systems, will be of unprecedented scale. As these systems continue to grow in size, faults will become increasingly common, even over the course of small calculations. Therefore, issues such as fault tolerance and reliability will limit application scalability. Current techniques to ensure progress across faults like checkpoint/restart, the dominant fault tolerance mechanism for the last 25 years, are increasingly problematic at the scales of future systems due to their excessive overheads. In this work, we evaluate a number of techniques to decrease the overhead of checkpoint/restart and keep this method viable for future exascale systems. More specifically, this work evaluates state-machine replication to dramatically increase the checkpoint interval (the time between successive checkpoint) and hash-based, probabilistic incremental checkpointing using graphics processing units to decrease the checkpoint commit time (the time to save one checkpoint). Using a combination of empirical analysis, modeling, and simulation, we study the costs and benefits of these approaches on a wide range of parameters. These results, which cover of number of high-performance computing capability workloads, different failure distributions, hardware mean time to failures, and I/O bandwidths, show the potential benefits of these techniques for meeting the reliability demands of future exascale platforms.

  3. Protein phosphatases acting on the replication checkpoint

    OpenAIRE

    Conde, Rui Miguel Esteves Antunes Seabra

    2010-01-01

    A génese de um cancro está dependente da acumulação de mutações genéticas que dão origem a instabilidade genómica, que por sua vez resulta na proliferação descontrolada. Para prevenir a acumulação destas mutações, as células têm mecanismos de controlo (checkpoints) que suspendem o ciclo celular e accionam as vias de reparação do ADN. Estes eventos são muitas vezes regulados por dinâmicas de (des)fosforilação de proteínas. As proteínas fosfatases (PPs), enzimas responsáveis p...

  4. Managing Adverse Events With Immune Checkpoint Agents.

    Science.gov (United States)

    Dadu, Ramona; Zobniw, Chrystia; Diab, Adi

    2016-01-01

    Immune checkpoint inhibitors (anti-cytotoxic T-lymphocyte antigen 4 and anti programmed cell death 1/programmed cell death 1 ligand antibodies) have shown impressive clinical activity in multiple cancer types. Despite achieving great clinical success, challenges and limitations of these drugs as monotherapy or various combinational strategies include the development of a unique set of immune-related adverse events (irAEs) that can be severe and even fatal. Therefore, identification of patients at risk, prevention, consistent communication between patients and medical team, rapid recognition, and treatment of irAEs are critical in optimizing treatment outcomes. This review focuses on the description of more common irAEs and provides a suggested approach for management of specific irAEs. PMID:27111908

  5. Targeting lung cancer through inhibition of checkpoint kinases

    OpenAIRE

    Syljuåsen, Randi G; Hasvold, Grete; Hauge, Sissel; Helland, Åslaug

    2015-01-01

    Inhibitors of checkpoint kinases ATR, Chk1, and Wee1 are currently being tested in preclinical and clinical trials. Here, we review the basic principles behind the use of such inhibitors as anticancer agents, and particularly discuss their potential for treatment of lung cancer. As lung cancer is one of the most deadly cancers, new treatment strategies are highly needed. We discuss how checkpoint kinase inhibition in principle can lead to selective killing of lung cancer cells while sparing t...

  6. A New Adaptive Checkpointing Strategy for Mobile Computing

    Institute of Scientific and Technical Information of China (English)

    MENChaoguang; ZUODecheng; YANGXiaozong

    2005-01-01

    Adaptive checkpointing strategy is an efficient recovery scheme, which is suitable for mobile computing system. However, all existing adaptive checkpointing schemes are not correct to recover system when failure occurs in some special period. In this paper, the issues that will lead to system inconsistency are first discussed and then a new adaptive strategy that can recover system to correct consistent state is proposed. Our algorithm improves system recovery performance because only failure process needs rollback through logging.

  7. A Hierarchical Checkpointing Protocol for Parallel Applications in Cluster Federations

    OpenAIRE

    Monnet, Sébastien; Morin, Christine; Badrinath, Ramamurthy

    2004-01-01

    Code coupling applications can be divided into communicating modules, that may be executed on different clusters in a cluster federation. As a cluster federation comprises of a large number of nodes, there is a high probability of a node failure. We propose a hierarchical checkpointing protocol that combines a synchronized checkpointing technique inside clusters and a communication-induced technique between clusters. This protocol fits to the characteristics of a cluster federation (large num...

  8. Cyclin-dependent kinase CDK1/CDC28 and checkpoints

    International Nuclear Information System (INIS)

    The genetic instability induced by defects in the cell cycle progression contributes to different human diseases, particularly neoplastic transformation. The control mechanisms of correct cell cycle progression are the most studied in the yeast Saccharomyces cerevisiae, in which checkpoint was first discovered. Many components of these processes have been identified by now. Here, the role of the central kinase of cell cycle CDK1/CDC28 is considered in checkpoint in different phases.

  9. DMTCP: bringing interactive checkpoint-restart to Python

    Science.gov (United States)

    Arya, Kapil; Cooperman, Gene

    2015-01-01

    DMTCP (Distributed MultiThreaded CheckPointing) is a mature checkpoint-restart package. It operates in user space without kernel privilege, and adapts to application-specific requirements through plugins. While DMTCP has been able to checkpoint Python and IPython ‘from the outside’ for many years, a Python module has recently been created to support DMTCP. IPython support is included through a new DMTCP plugin. A checkpoint can be requested interactively within a Python session or under the control of a specific Python program. Further, the Python program can execute specific Python code prior to checkpoint, upon resuming (within the original process) and upon restarting (from a checkpoint image). Applications of DMTCP are demonstrated for: (i) Python-based graphics using virtual network client, (ii) a fast/slow technique to use multiple hosts or cores to check one (Cython Behnel S et al 2011 Comput. Sci. Eng. 13 31-39) computation in parallel, and (iii) a reversible debugger, FReD, with a novel reverse-expression watchpoint feature for locating the cause of a bug.

  10. A survey of checkpointing algorithms for parallel and distributed computers

    Indian Academy of Sciences (India)

    S Kalaiselvi; V Rajaraman

    2000-10-01

    Checkpoint is defined as a designated place in a program at which normal processing is interrupted specifically to preserve the status information necessary to allow resumption of processing at a later time. Checkpointing is the process of saving the status information. This paper surveysthe algorithms which have been reported in the literature for checkpointing parallel/distributed systems. It has been observed that most of the algorithms published for checkpointing in message passing systems are based on the seminal article by Chandy and Lamport. A large number of articles have been published in this area by relaxing the assumptions made in this paper and by extending it to minimise the overheads of coordination and context saving. Checkpointing for sharedmemory systems primarily extend cache coherence protocolstomaintain a consistent memory. All of them assume that the main memory is safe for storing the context. Recently algorithms have been published for distributed shared memory systems, which extend the cache coherence protocols used in shared memory systems. They however also include methods for storing the status of distributed memory in stable storage. Most of the algorithms assume that there is no knowledge about the programs being executed.It is howeverfelt that in development of parallel programs the user has to do a fair amount of work in distributing tasks and this information can be effectively used to simplify checkpointing and rollback recovery.

  11. An Analysis of Checkpointing Algorithms for Distributed Mobile Systems

    Directory of Open Access Journals (Sweden)

    Ajay Khunteta

    2010-07-01

    Full Text Available Distributed snapshots are an important building block for distributed systems, and are useful for constructing efficient checkpointing protocols, among other uses. Direct application of these algorithms to mobile systems is not easible, however, due to differences in the environment in which mobile systems operate, relative to general distributed systems. The mobile computing environment introduces newchallenges in the area of fault-tolerant computing. Compared to traditional distributed environments, wireless networks are typically slower, providing lower throughput and latency, comparing to wireline networks. In addition, the mobile hosts have limited computation esources, are often exposed to harsh operating environment that makes them more likely to fail, and can roam while operating. Over the past two decades, intensive research work has been carried out on providing efficient checkpointing protocols in traditional distributed computing. Recently, more attention has been paid to providing checkpointing protocols for mobile systems. Some of these protocols have been adapted from the traditional distributed environment; others have been created from scratch for mobile systems. Checkpoint is defined as a designated place in a program at which normal processing is interrupted specifically to preserve the status information necessary to allow resumption of processing at alater time. Checkpointing is the process of saving the status information. This paper surveys the algorithms which have been reported in the literature for checkpointing in Mobile Distributed systems.

  12. A Tunable Checkpointing Algorithm for Distributed Mobile Applications

    Directory of Open Access Journals (Sweden)

    Sungchae Lim

    2011-11-01

    Full Text Available The aim of a distributed checkpointing algorithm is to efficiently restore the execution state of distributed applications in face of hardware or software failures. Originally, such algorithms were devised for fixed networking systems, of which computing components communicate with each other via wired networks. Therefore, those algorithms usually suffer from heavy networking costs coming from frequent data transits over wireless networks, if they are used in the wireless computing environment. In this paper, to reduce usage of wireless communications, our checkpointing algorithm allows the distributed mobile application to tune the level of its checkpointing strictness. The strictness is defined by the maximum rollback distance (MRD that says how many recent local checkpoints can be rolled back in the worst case. Since our algorithm have more flexibility in checkpointing schedule due to the use of MRD, it is possible to reduce the number of enforced local checkpointing. In particular, the amount of data transited on wirelesses networks becomes smaller than in earlier methods; thus, our algorithm provides less communication cost and shortened blocking time.

  13. Review of Some Checkpointing Schemes for Distributed and Mobile Computing Environments

    Directory of Open Access Journals (Sweden)

    Mr Raman Kumar

    2015-05-01

    Full Text Available Fault Tolerance Techniques facilitate systems to carry out tasks in the incidence of faults. A checkpoint is a local state of a process saved on stable storage. In a distributed system, since the processes in the system do not share memory; a global state of the system is defined as a combination of local states, one from each process. In case of a fault in distributed systems, checkpointing enables the execution of a program to be resumed from a previous consistent global state rather than resuming the execution from the commencement. In this way, the sum of constructive processing vanished because of the fault is appreciably reduced. In this paper, we talk about various issues related to the checkpointing for distributed systems and mobile computing environments. We also confer various types of checkpointing: coordinated checkpointing, asynchronous checkpointing, communication induced checkpointing and message logging based checkpointing. We also present a survey of some checkpointing algorithms for distributed systems.

  14. Endogenous ephrinB2 mediates colon-urethra cross-organ sensitization via Src kinase-dependent tyrosine phosphorylation of NR2B.

    Science.gov (United States)

    Peng, Hsien-Yu; Chen, Gin-Den; Lai, Cheng-Hung; Tung, Kwong-Chung; Chang, Junn-Liang; Lin, Tzer-Bin

    2010-01-01

    Recently, the role of EphB receptor (EphBR) tyrosine kinase and their ephrinB ligands in spinal pain-related neural plasticity has been identified. To test whether Src-family non-receptor tyrosine kinase-dependent glutamatergic N-methyl-d-aspartate receptor (NMDAR) NR2B subunit phosphorylation underlies lumbosacral spinal EphBR activation to mediate cross-organ sensitization between the colon and the urethra, external urethra sphincter electromyogram activity evoked by pelvic nerve stimulation and protein expression in the lumbosacral (L6-S2) dorsal horn were studied before and after intracolonic mustard oil (MO) instillation. We found MO instillation produced colon-urethra reflex sensitization along with an upregulation of endogenous ephrinB2 expression as well as phosphorylation of EphB 1/2, Src-family kinase, and NR2B tyrosine residues. Intrathecal immunoglobulin fusion protein of EphB1 and EphB2 as well as PP2 reversed the reflex sensitization and NR2B phosphorylation caused by MO. All these results suggest that EphBR-ephrinB interactions, which provoke Src-family kinase-dependent NMDAR NR2B phosphorylation at the lumbosacral spinal cord level, are involved in cross-organ sensitization, contributing to the development of viscero-visceral referred pain between the bowel and the urethra. PMID:19864302

  15. Berkeley lab checkpoint/restart (BLCR) for Linux clusters

    Science.gov (United States)

    Hargrove, Paul H.; Duell, Jason C.

    2006-09-01

    This article describes the motivation, design and implementation of Berkeley Lab Checkpoint/Restart (BLCR), a system-level checkpoint/restart implementation for Linux clusters that targets the space of typical High Performance Computing applications, including MPI. Application-level solutions, including both checkpointing and fault-tolerant algorithms, are recognized as more time and space efficient than system-level checkpoints, which cannot make use of any application-specific knowledge. However, system-level checkpointing allows for preemption, making it suitable for responding to ''fault precursors'' (for instance, elevated error rates from ECC memory or network CRCs, or elevated temperature from sensors). Preemption can also increase the efficiency of batch scheduling; for instance reducing idle cycles (by allowing for shutdown without any queue draining period or reallocation of resources to eliminate idle nodes when better fitting jobs are queued), and reducing the average queued time (by limiting large jobs to running during off-peak hours, without the need to limit the length of such jobs). Each of these potential uses makes BLCR a valuable tool for efficient resource management in Linux clusters.

  16. Message Efficient Checkpointing and Rollback Recovery in Heterogeneous Mobile Networks

    Science.gov (United States)

    Jaggi, Parmeet Kaur; Singh, Awadhesh Kumar

    2016-06-01

    Heterogeneous networks provide an appealing way of expanding the computing capability of mobile networks by combining infrastructure-less mobile ad-hoc networks with the infrastructure-based cellular mobile networks. The nodes in such a network range from low-power nodes to macro base stations and thus, vary greatly in their capabilities such as computation power and battery power. The nodes are susceptible to different types of transient and permanent failures and therefore, the algorithms designed for such networks need to be fault-tolerant. The article presents a checkpointing algorithm for the rollback recovery of mobile hosts in a heterogeneous mobile network. Checkpointing is a well established approach to provide fault tolerance in static and cellular mobile distributed systems. However, the use of checkpointing for fault tolerance in a heterogeneous environment remains to be explored. The proposed protocol is based on the results of zigzag paths and zigzag cycles by Netzer-Xu. Considering the heterogeneity prevalent in the network, an uncoordinated checkpointing technique is employed. Yet, useless checkpoints are avoided without causing a high message overhead.

  17. Berkeley lab checkpoint/restart (BLCR) for Linux clusters

    International Nuclear Information System (INIS)

    This article describes the motivation, design and implementation of Berkeley Lab Checkpoint/Restart (BLCR), a system-level checkpoint/restart implementation for Linux clusters that targets the space of typical High Performance Computing applications, including MPI. Application-level solutions, including both checkpointing and fault-tolerant algorithms, are recognized as more time and space efficient than system-level checkpoints, which cannot make use of any application-specific knowledge. However, system-level checkpointing allows for preemption, making it suitable for responding to ''fault precursors'' (for instance, elevated error rates from ECC memory or network CRCs, or elevated temperature from sensors). Preemption can also increase the efficiency of batch scheduling; for instance reducing idle cycles (by allowing for shutdown without any queue draining period or reallocation of resources to eliminate idle nodes when better fitting jobs are queued), and reducing the average queued time (by limiting large jobs to running during off-peak hours, without the need to limit the length of such jobs). Each of these potential uses makes BLCR a valuable tool for efficient resource management in Linux clusters

  18. The blockade of immune checkpoints in cancer immunotherapy

    Science.gov (United States)

    Pardoll, Drew M.

    2016-01-01

    Among the most promising approaches to activating therapeutic antitumour immunity is the blockade of immune checkpoints. Immune checkpoints refer to a plethora of inhibitory pathways hardwired into the immune system that are crucial for maintaining self-tolerance and modulating the duration and amplitude of physiological immune responses in peripheral tissues in order to minimize collateral tissue damage. It is now clear that tumours co-opt certain immune-checkpoint pathways as a major mechanism of immune resistance, particularly against T cells that are specific for tumour antigens. Because many of the immune checkpoints are initiated by ligand–receptor interactions, they can be readily blocked by antibodies or modulated by recombinant forms of ligands or receptors. Cytotoxic T-lymphocyte-associated antigen 4 (CTLA4) antibodies were the first of this class of immunotherapeutics to achieve US Food and Drug Administration (FDA) approval. Preliminary clinical findings with blockers of additional immune-checkpoint proteins, such as programmed cell death protein 1 (PD1), indicate broad and diverse opportunities to enhance antitumour immunity with the potential to produce durable clinical responses. PMID:22437870

  19. Checkpoint Inhibitors for the Treatment of Hodgkin Lymphoma.

    Science.gov (United States)

    Bennani-Baiti, Nabila; Thanarajasingam, Gita; Ansell, Stephen

    2016-06-01

    Hodgkin lymphoma's (HL) tumor composition is characterized by a paucity of malignant cells and a preponderance of immune and stromal cells. Despite the rich immune milieu within the tumor microenvironment, malignant cells are able to effectively evade the immune system and use immune support to promote lymphoma cell growth and proliferation. Recognizing this has led to the identification of checkpoint inhibitory signals that enable immune evasion and to opening the door to therapeutic strategies on how to exploit the immune system in targeting tumor cells. We discuss herein some of the tumor evasion mechanisms in HL with a particular focus on the immune checkpoint pathways and focus on recent clinical data of checkpoint blockade in HL treatment. PMID:26818843

  20. Immune checkpoint blockade therapy for bladder cancer treatment.

    Science.gov (United States)

    Kim, Jayoung

    2016-06-01

    Bladder cancer remains the most immunogenic and expensive malignant tumor in the United States today. As the 4th leading cause of death from cancer in United States, Immunotherapy blocking immune checkpoints have been recently been applied to many aggressive cancers and changed interventions of urological cancers including advanced bladder cancer. The applied inhibition of PD-1-PD-L1 interactions can restore antitumor T-cell activity and enhance the cellular immune attack on antigens. The overall goals of this short review article are to introduce current cancer immunotherapy and immune checkpoint inhibitors, and to provide new insight into the underlying mechanisms that block immune checkpoints in tumor microenvironment. Furthermore, this review will address the preclinical and clinical trials to determine whether bladder cancer patients could benefit from this new cancer therapy in near future. PMID:27326412

  1. Low-Overhead Non-Blocking Checkpointing Scheme for Mobile Computing Systems

    Institute of Scientific and Technical Information of China (English)

    MEN Chaoguang; CAO Liujuan; WANG Liwen; XU Zhenpeng

    2007-01-01

    When applied to mobile computing systems, checkpoint protocols for distributed computing systems would face many new challenges, such as low wireless bandwidth, frequent disconnections, and lack of stable storage at mobile hosts. This paper proposes a novel checkpoint protocol to effectively reduce the coordinating overhead. By using a communication vector, only a few processes participate in the checkpointing event. During checkpointing, the scheme can save the time used to trace the dependency tree by sending checkpoint requests to dependent processes at once. In addition, processes are non-blocking in this scheme, since the inconsistency is resolved by the piggyback technique. Hence the unnecessary and orphan messages can be avoided. Compared with the traditional coordinated checkpoint approach, the proposed non-blocking algorithm obtains a minimal number of processes to take checkpoints. It also reduces the checkpoint latency, which brings less overhead to mobile host with limited resources.

  2. Keeping it together in times of stress: checkpoint function at stalled replication forks

    OpenAIRE

    Berens, Theresa J.; David P Toczyski

    2012-01-01

    In this issue, De Piccoli et al. (2012) show that, contrary to current models of DNA replication checkpoint function, replication proteins remain associated with each other and with replicating DNA when replication is stressed in checkpoint-deficient cells.

  3. Calf-ovary protein kinases dependent on adenosine 3':5' -monophosphate. Analysis by electrophoresis and electro-focusing on polyacrylamide get.

    Science.gov (United States)

    Salokangas, A; Talmadge, K; Bechtel, E; Eppenberger, U; Chrambach, A

    1977-03-01

    High resolving power and quantitative application polyacrylamide-gel electrophopresis at various pore sizes and electrofocusing provide resolution of a calf-ovarian protein-kinase system at an increased level of magnification, as well as optimal preparative routes. Three protein kinases dependent on adenosine 3':5' -monophosphate are distinguished by polyacrylamide gel electrophoresis in calf ovarian cytosol. These enzymes which are observed in the pH range 7.5--10.2, appear to be aggregates of a commonsubmit or monomer. The three kinases are, by the criteria of polyacylamide gel electrophoresis, distinct from three adenosine-3':5' -monophosphate-binding proteins found in the calf ovarian system. Analysis by electrofocusing on polyacrylamide gel shows that conventionally purified preparations of the major kinase of cytosol contain an overwhelming majority of contaminant proteins. PMID:191253

  4. Immune-Checkpoint Blockade and Active Immunotherapy for Glioma

    Energy Technology Data Exchange (ETDEWEB)

    Ahn, Brian J. [Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States); Brain Tumor Program, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213 (United States); Pollack, Ian F. [Brain Tumor Program, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213 (United States); Department of Neurological Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States); Okada, Hideho, E-mail: okadah@upmc.edu [Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States); Brain Tumor Program, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213 (United States); Department of Neurological Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States); Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States)

    2013-11-01

    Cancer immunotherapy has made tremendous progress, including promising results in patients with malignant gliomas. Nonetheless, the immunological microenvironment of the brain and tumors arising therein is still believed to be suboptimal for sufficient antitumor immune responses for a variety of reasons, including the operation of “immune-checkpoint” mechanisms. While these mechanisms prevent autoimmunity in physiological conditions, malignant tumors, including brain tumors, actively employ these mechanisms to evade from immunological attacks. Development of agents designed to unblock these checkpoint steps is currently one of the most active areas of cancer research. In this review, we summarize recent progresses in the field of brain tumor immunology with particular foci in the area of immune-checkpoint mechanisms and development of active immunotherapy strategies. In the last decade, a number of specific monoclonal antibodies designed to block immune-checkpoint mechanisms have been developed and show efficacy in other cancers, such as melanoma. On the other hand, active immunotherapy approaches, such as vaccines, have shown encouraging outcomes. We believe that development of effective immunotherapy approaches should ultimately integrate those checkpoint-blockade agents to enhance the efficacy of therapeutic approaches. With these agents available, it is going to be quite an exciting time in the field. The eventual success of immunotherapies for brain tumors will be dependent upon not only an in-depth understanding of immunology behind the brain and brain tumors, but also collaboration and teamwork for the development of novel trials that address multiple layers of immunological challenges in gliomas.

  5. The absence of a DNA replication checkpoint in porcine zygotes

    Czech Academy of Sciences Publication Activity Database

    Vacková, I.; Křen, Radomír; Loi, P.; Krylov, V.; Fulka Jr., J.

    2006-01-01

    Roč. 14, 1 (2006), s. 33-37. ISSN 0967-1994 Institutional research plan: CEZ:AV0Z50450515 Keywords : checkpoint * DNA replication * fertilization Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.782, year: 2006

  6. Immune-Checkpoint Blockade and Active Immunotherapy for Glioma

    Directory of Open Access Journals (Sweden)

    Brian J. Ahn

    2013-11-01

    Full Text Available Cancer immunotherapy has made tremendous progress, including promising results in patients with malignant gliomas. Nonetheless, the immunological microenvironment of the brain and tumors arising therein is still believed to be suboptimal for sufficient antitumor immune responses for a variety of reasons, including the operation of “immune-checkpoint” mechanisms. While these mechanisms prevent autoimmunity in physiological conditions, malignant tumors, including brain tumors, actively employ these mechanisms to evade from immunological attacks. Development of agents designed to unblock these checkpoint steps is currently one of the most active areas of cancer research. In this review, we summarize recent progresses in the field of brain tumor immunology with particular foci in the area of immune-checkpoint mechanisms and development of active immunotherapy strategies. In the last decade, a number of specific monoclonal antibodies designed to block immune-checkpoint mechanisms have been developed and show efficacy in other cancers, such as melanoma. On the other hand, active immunotherapy approaches, such as vaccines, have shown encouraging outcomes. We believe that development of effective immunotherapy approaches should ultimately integrate those checkpoint-blockade agents to enhance the efficacy of therapeutic approaches. With these agents available, it is going to be quite an exciting time in the field. The eventual success of immunotherapies for brain tumors will be dependent upon not only an in-depth understanding of immunology behind the brain and brain tumors, but also collaboration and teamwork for the development of novel trials that address multiple layers of immunological challenges in gliomas.

  7. Attachment issues : kinetochore transformations and spindle checkpoint silencing

    NARCIS (Netherlands)

    Etemad, Banafsheh; Kops, Geert Jpl

    2016-01-01

    Cell division culminates in the segregation of duplicated chromosomes in opposite directions prior to cellular fission. This process is guarded by the spindle assembly checkpoint (SAC), which prevents the anaphase of cell division until stable connections between spindle microtubules and the kinetoc

  8. Immune-Checkpoint Blockade and Active Immunotherapy for Glioma

    International Nuclear Information System (INIS)

    Cancer immunotherapy has made tremendous progress, including promising results in patients with malignant gliomas. Nonetheless, the immunological microenvironment of the brain and tumors arising therein is still believed to be suboptimal for sufficient antitumor immune responses for a variety of reasons, including the operation of “immune-checkpoint” mechanisms. While these mechanisms prevent autoimmunity in physiological conditions, malignant tumors, including brain tumors, actively employ these mechanisms to evade from immunological attacks. Development of agents designed to unblock these checkpoint steps is currently one of the most active areas of cancer research. In this review, we summarize recent progresses in the field of brain tumor immunology with particular foci in the area of immune-checkpoint mechanisms and development of active immunotherapy strategies. In the last decade, a number of specific monoclonal antibodies designed to block immune-checkpoint mechanisms have been developed and show efficacy in other cancers, such as melanoma. On the other hand, active immunotherapy approaches, such as vaccines, have shown encouraging outcomes. We believe that development of effective immunotherapy approaches should ultimately integrate those checkpoint-blockade agents to enhance the efficacy of therapeutic approaches. With these agents available, it is going to be quite an exciting time in the field. The eventual success of immunotherapies for brain tumors will be dependent upon not only an in-depth understanding of immunology behind the brain and brain tumors, but also collaboration and teamwork for the development of novel trials that address multiple layers of immunological challenges in gliomas

  9. Checkpointing for graceful degradation in distributed embedded systems

    Science.gov (United States)

    Sababha, Belal Hussein

    Graceful degradation is an approach to developing dependable safety-critical embedded applications, where redundant active or standby resources are used to cope with faults through a system reconfiguration at run-time. Compared to traditional hardware and software redundancy, it is a promising technique that may achieve dependability with a significant reduction in cost, size, weight, and power requirements. Reconfiguration at run-time necessitates using proper checkpointing protocols to support state reservation to ensure correct task restarts after a system reconfiguration. One of the most common checkpointing protocols are communication induced checkpointing (CIC) protocols, which are well developed and understood for large parallel and information systems, but not much has been done for resource limited embedded systems. This work implements and evaluates some of the most common CIC protocols in a periodic resource constrained distributed embedded system for graceful degradation purposes. A test-bed has been developed and used for the evaluation of the various protocols. The implemented protocols are thoroughly studied and performances are contrasted. Specifically the periodicity property and how it benefits checkpointing in embedded systems is investigated. This work introduces a unique effort of CIC protocol implementation and evaluation in the field of distributed embedded systems. Other than providing a test-bed for graceful degradation support, this work shows that some checkpointing protocols that are not efficient in large information systems and supercomputers perform well in embedded systems. We show that a simple index-based CIC protocol, such as the BCS protocol, is more appropriate in embedded system applications compared to other protocols that piggyback a significant amount of information to reduce the number of forced checkpoints. Finally, this work proposes a whole graceful degradation approach to achieve fault tolerance in resource constrained

  10. Cell size checkpoint control by the retinoblastoma tumor suppressor pathway.

    Directory of Open Access Journals (Sweden)

    Su-Chiung Fang

    2006-10-01

    Full Text Available Size control is essential for all proliferating cells, and is thought to be regulated by checkpoints that couple cell size to cell cycle progression. The aberrant cell-size phenotypes caused by mutations in the retinoblastoma (RB tumor suppressor pathway are consistent with a role in size checkpoint control, but indirect effects on size caused by altered cell cycle kinetics are difficult to rule out. The multiple fission cell cycle of the unicellular alga Chlamydomonas reinhardtii uncouples growth from division, allowing direct assessment of the relationship between size phenotypes and checkpoint function. Mutations in the C. reinhardtii RB homolog encoded by MAT3 cause supernumerous cell divisions and small cells, suggesting a role for MAT3 in size control. We identified suppressors of an mat3 null allele that had recessive mutations in DP1 or dominant mutations in E2F1, loci encoding homologs of a heterodimeric transcription factor that is targeted by RB-related proteins. Significantly, we determined that the dp1 and e2f1 phenotypes were caused by defects in size checkpoint control and were not due to a lengthened cell cycle. Despite their cell division defects, mat3, dp1, and e2f1 mutants showed almost no changes in periodic transcription of genes induced during S phase and mitosis, many of which are conserved targets of the RB pathway. Conversely, we found that regulation of cell size was unaffected when S phase and mitotic transcription were inhibited. Our data provide direct evidence that the RB pathway mediates cell size checkpoint control and suggest that such control is not directly coupled to the magnitude of periodic cell cycle transcription.

  11. Review of some Minimum-Process Synchronous Checkpointing Schemes for Mobile Distributed Systems

    Directory of Open Access Journals (Sweden)

    Preeti Gupta,

    2010-07-01

    Full Text Available The term Distributed Systems is used to describe a system with the following characteristics: i it consists of several computers that do not share memory or a clock, ii the computers communicate with each other by exchanging messages over a communication network, iii each computer has its own memory and runs its own operating system. A mobile computing system is a distributed system where some of processes are running on mobile hosts (MHs, whose location in the network changes with time. The number of processes that take checkpoints is minimized to 1 avoid awakening of MHs in doze mode of operation, 2 minimize thrashing of MHs with checkpointing activity, 3 save limited battery life of MHs and low bandwidth of wireless channels. In minimum-process checkpointing protocols, some useless checkpoints are taken or blocking of processes takes place. To take a checkpoint, an MH has to transfer a large amount of checkpoint data to its local MSS over the wireless network. Since the wireless network has low bandwidth and MHs have low computation power, all-process checkpointing will waste the scarce resources of the mobile system on every checkpoint.Minimum-process coordinated checkpointing is a preferred approach for mobile distributed systems. In this paper, we discuss various existing minimum-process checkpointing protocols for mobile distributed systems.

  12. Synthetic Physical Interactions Map Kinetochore-Checkpoint Activation Regions.

    Science.gov (United States)

    Ólafsson, Guðjón; Thorpe, Peter H

    2016-01-01

    The spindle assembly checkpoint (SAC) is a key mechanism to regulate the timing of mitosis and ensure that chromosomes are correctly segregated to daughter cells. The recruitment of the Mad1 and Mad2 proteins to the kinetochore is normally necessary for SAC activation. This recruitment is coordinated by the SAC kinase Mps1, which phosphorylates residues at the kinetochore to facilitate binding of Bub1, Bub3, Mad1, and Mad2. There is evidence that the essential function of Mps1 is to direct recruitment of Mad1/2. To test this model, we have systematically recruited Mad1, Mad2, and Mps1 to most proteins in the yeast kinetochore, and find that, while Mps1 is sufficient for checkpoint activation, recruitment of either Mad1 or Mad2 is not. These data indicate an important role for Mps1 phosphorylation in SAC activation, beyond the direct recruitment of Mad1 and Mad2. PMID:27280788

  13. Epigenetic modifiers in immunotherapy: a focus on checkpoint inhibitors.

    Science.gov (United States)

    Terranova-Barberio, Manuela; Thomas, Scott; Munster, Pamela N

    2016-06-01

    Immune surveillance should be directed to suppress tumor development and progression, involving a balance of coinhibitory and costimulatory signals that amplify immune response without overwhelming the host. Immunotherapy confers durable clinical benefit in 'immunogenic tumors', whereas in other tumors the responses are modest. Thus, immune checkpoint inhibitors may need to be combined with strategies to boost immune response or increase the tumor immune profile. Epigenetic aberrations contribute significantly to carcinogenesis. Recent findings suggest that epigenetic drugs prime the immune response by increasing expression of tumor-associated antigens and immune-related genes, as well as modulating chemokines and cytokines involved in immune system activation. This review describes our current understanding regarding epigenetic and immunotherapy combination, focusing on immune response priming to checkpoint blockade. PMID:27197539

  14. Checkpoint signaling from a single DNA interstrand crosslink

    OpenAIRE

    Ben-Yehoyada, Merav; Wang, Lily C; Kozekov, Ivan D.; Rizzo, Carmelo J.; Gottesman, Max E.; Gautier, Jean

    2009-01-01

    DNA interstrand crosslinks (ICLs) are the most toxic lesions induced by chemotherapeutic agents such as Mitomycin C and Cisplatin. By covalently linking both DNA strands, ICLs prevent DNA melting, transcription, and replication. Studies on ICL signaling and repair have been limited because these drugs generate additional DNA lesions that trigger checkpoint signaling. Here, we monitor sensing, signaling from and repairing of a single, site-specific ICL in cell-free extract derived from Xenopus...

  15. Checkpointing for the RESTART problem in Markov networks

    OpenAIRE

    Lipsky, Lester; Doran, Derek; Gokhale, Swapna

    2011-01-01

    We apply the known formulae of the RESTART problem to Markov models of software (and many other) systems, and derive new equations. We show how checkpoints might be included, with their resultant performance under RESTART. The result is a complete procedure for finding the mean, variance, and tail behavior of the job completion time as a function of the failure rate. We also provide a detailed example.

  16. A Monitor for Bud Emergence in the Yeast Morphogenesis Checkpoint

    Science.gov (United States)

    Theesfeld, Chandra L.; Zyla, Trevin R.; Bardes, Elaine G.S.; Lew, Daniel J.

    2003-01-01

    Cell cycle transitions are subject to regulation by both external signals and internal checkpoints that monitor satisfactory progression of key cell cycle events. In budding yeast, the morphogenesis checkpoint arrests the cell cycle in response to perturbations that affect the actin cytoskeleton and bud formation. Herein, we identify a step in this checkpoint pathway that seems to be directly responsive to bud emergence. Activation of the kinase Hsl1p is dependent upon its recruitment to a cortical domain organized by the septins, a family of conserved filament-forming proteins. Under conditions that delayed or blocked bud emergence, Hsl1p recruitment to the septin cortex still took place, but hyperphosphorylation of Hsl1p and recruitment of the Hsl1p-binding protein Hsl7p to the septin cortex only occurred after bud emergence. At this time, the septin cortex spread to form a collar between mother and bud, and Hsl1p and Hsl7p were restricted to the bud side of the septin collar. We discuss models for translating cellular geometry (in this case, the emergence of a bud) into biochemical signals regulating cell proliferation. PMID:12925763

  17. Potential biomarker for checkpoint blockade immunotherapy and treatment strategy.

    Science.gov (United States)

    Dong, Zhong-Yi; Wu, Si-Pei; Liao, Ri-Qiang; Huang, Shu-Mei; Wu, Yi-Long

    2016-04-01

    Programmed cell death protein-1 (PD-1) and ligand (PD-L1) provide an important escape mechanism from immune attack, and blockade therapy of these proteins show promising clinical benefits in many types of cancer. PD-L1 can be induced by interferon-gamma (IFN-γ), hypoxia, or toll-like receptor (TLR)-mediated pathways that confer adaptive immune resistance, or upregulated by oncogenic signals leading to constitutive expression and resulting in intrinsic immune resistance. The PD-1/PD-L1 checkpoint blockade, which targets regulatory pathways in T cells to overcome immune resistance, is correlated to PD-L1 expression pattern and the presence of tumor-infiltrating lymphocytes (TILs). Meanwhile, immunogenic mutation loads show significant response to checkpoint blockade, which is probably due to PD-1/L1 status and TIL content. Finally, the clinical strategies to design effective checkpoint-targeting immunotherapies are based on the classification of inducible/constitutive expression of PD-L1 and the presence of TILs. PMID:26779629

  18. Complex Commingling: Nucleoporins and the Spindle Assembly Checkpoint

    Directory of Open Access Journals (Sweden)

    Ikram Mossaid

    2015-11-01

    Full Text Available The segregation of the chromosomes during mitosis is an important process, in which the replicated DNA content is properly allocated into two daughter cells. To ensure their genomic integrity, cells present an essential surveillance mechanism known as the spindle assembly checkpoint (SAC, which monitors the bipolar attachment of the mitotic spindle to chromosomes to prevent errors that would result in chromosome mis-segregation and aneuploidy. Multiple components of the nuclear pore complex (NPC, a gigantic protein complex that forms a channel through the nuclear envelope to allow nucleocytoplasmic exchange of macromolecules, were shown to be critical for faithful cell division and implicated in the regulation of different steps of the mitotic process, including kinetochore and spindle assembly as well as the SAC. In this review, we will describe current knowledge about the interconnection between the NPC and the SAC in an evolutional perspective, which primarily relies on the two mitotic checkpoint regulators, Mad1 and Mad2. We will further discuss the role of NPC constituents, the nucleoporins, in kinetochore and spindle assembly and the formation of the mitotic checkpoint complex during mitosis and interphase.

  19. Learning common lessons and checkpoints form human error incidents. Maintenance

    International Nuclear Information System (INIS)

    CRIEPI has been conducting detailed and structured analyses of all human error incidents collected from Japanese Licensee Event Reports using J-HPES (Japanese version of HPES) as an analysis method. Results obtained by the analyses have been stored in J-HPES database. This paper described the process to analyze J-HPES data concerning maintenance systematically to extract problems identified in the process of error action and checkpoints for preventing errors. Human error actions of these J-HPES data are classified by viewpoints of error mode and work type. As to each of these error categories, problems are extracted based on J-HPES causal relation charts in following three viewpoints: acts at workplace, activities, and preconditions of job. Moreover, checkpoints for preventing errors are developed referring proposed countermeasures in J-HPES database. In order to share these results, we started to issue 4-pages booklets 'Catch the Point' periodically. In future, based on these results, we will publish a teaching material. We also have a plan to store the contents of Catch the Point in a database, which facilitate users to find necessary checkpoints and hazards before they start their activities. (author)

  20. Depletion of RIPK3 or MLKL blocks TNF-driven necroptosis and switches towards a delayed RIPK1 kinase-dependent apoptosis.

    Science.gov (United States)

    Remijsen, Q; Goossens, V; Grootjans, S; Van den Haute, C; Vanlangenakker, N; Dondelinger, Y; Roelandt, R; Bruggeman, I; Goncalves, A; Bertrand, M J M; Baekelandt, V; Takahashi, N; Berghe, T V; Vandenabeele, P

    2014-01-01

    In human cells, the RIPK1-RIPK3-MLKL-PGAM5-Drp1 axis drives tumor necrosis factor (TNF)-induced necroptosis through mitochondrial fission, but whether this pathway is conserved among mammals is not known. To answer this question, we analyzed the presence and functionality of the reported necroptotic axis in mice. As in humans, knockdown of receptor-interacting kinase-3 (RIPK3) or mixed lineage kinase domain like (MLKL) blocks TNF-induced necroptosis in L929 fibrosarcoma cells. However, repression of either of these proteins did not protect the cells from death, but instead induced a switch from TNF-induced necroptosis to receptor-interacting kinase-1 (RIPK1) kinase-dependent apoptosis. In addition, although mitochondrial fission also occurs during TNF-induced necroptosis in L929 cells, we found that knockdown of phosphoglycerate mutase 5 (PGAM5) and dynamin 1 like protein (Drp1) did not markedly protect the cells from TNF-induced necroptosis. Depletion of Pink1, a reported interactor of both PGAM5 and Drp1, did not affect TNF-induced necroptosis. These results indicate that in these murine cells mitochondrial fission and Pink1 dependent processes, including Pink-Parkin dependent mitophagy, apparently do not promote necroptosis. Our data demonstrate that the core components of the necrosome (RIPK1, RIPK3 and MLKL) are crucial to induce TNF-dependent necroptosis both in human and in mouse cells, but the associated mechanisms may differ between the two species or cell types. PMID:24434512

  1. Emodnet Med Sea Check-Point - Indicators for decision- maker

    Science.gov (United States)

    Besnard, Sophie; Claverie, Vincent; Blanc, Frédérique

    2015-04-01

    The Emodnet Checkpoint projects aim is to assess the cost-effectiveness, reliability and utility of the existing monitoring at the sea basin level. This involves the development of monitoring system indicators and a GIS Platform to perform the assessment and make it available. Assessment or production of Check-Point information is made by developing targeted products based on the monitoring data and determining whether the products are meeting the needs of industry and public authorities. Check-point users are the research community, the 'institutional' policy makers for IMP and MSFD implementation, the 'intermediate users', i.e., users capable to understand basic raw data but that benefit from seeing the Checkpoint targeted products and the assessment of the fitness for purpose. We define assessment criteria aimed to characterize/depict the input datasets in terms of 3 territories capable to show performance and gaps of the present monitoring system, appropriateness, availability and fitness for purpose. • Appropriateness: What is made available to users? What motivate/decide them to select this observation rather than this one. • Availability: How this is made available to the user? Place to understand the readiness and service performance of the EU infrastructure • Fitness for use / fitness for purpose: Ability for non-expert user to appreciate the data exploitability (feedback on efficiency & reliability of marine data) For each territory (appropriateness, Availability and Fitness for purpose / for use), we define several indicators. For example, for Availability we define Visibility, Accessibility and Performance. And Visibility is itself defined by "Easily found" and "EU service". So these indicators can be classified according to their territory and sub-territory as seen above, but also according to the complexity to build them. Indicators are built from raw descriptors in 3 stages:  Stage 1: to give a neutral and basic status directly computed from

  2. Checkpoint Defects Leading to Premature Mitosis Also Cause Endoreplication of DNA in Aspergillus nidulans

    OpenAIRE

    De Souza, Colin P. C.; Ye, Xiang S.; Osmani, Stephen A.

    1999-01-01

    The G2 DNA damage and slowing of S-phase checkpoints over mitosis function through tyrosine phosphorylation of NIMXcdc2 in Aspergillus nidulans. We demonstrate that breaking these checkpoints leads to a defective premature mitosis followed by dramatic rereplication of genomic DNA. Two additional checkpoint functions, uvsB and uvsD, also cause the rereplication phenotype after their mutation allows premature mitosis in the presence of low concentrations of hydroxyurea. uvsB is shown to encode ...

  3. The Dynamics of the Unreplicated DNA Checkpoint in Xenopus laevis Embryos and Extracts.

    OpenAIRE

    Adjerid, Nassiba

    2008-01-01

    When unreplicated or damaged DNA is present, cell cycle checkpoint pathways cause cell cycle arrest by inhibiting cyclin-dependent kinases (Cdks). In Xenopus laevis, early embryonic development consists of twelve rapid cleavage cycles between DNA replication (S) and mitosis (M) without checkpoints or gap phases. However, checkpoints are engaged in Xenopus once the embryo reaches the midblastula transition (MBT). At this point, the embryo initiates transcription, acquires gap phases between S...

  4. Assays Used to Study the DNA Replication Checkpoint in Fission Yeast

    OpenAIRE

    Noguchi, Eishi; Ansbach, Alison B.; Noguchi, Chiaki; Russell, Paul

    2009-01-01

    The DNA replication checkpoint, also known as the intra-S or S-phase checkpoint, plays a central role in ensuring the accuracy of DNA replication. When replication is impeded by DNA damage or other conditions, this checkpoint delays cell cycle progression and coordinates resumption of replication with DNA repair pathways. One of its critical functions is to stabilize stalled replication forks in a replication-competent state, presumably by maintaining proper assembly of replisome components a...

  5. Action-oriented use of ergonomic checkpoints for healthy work design in different settings.

    Science.gov (United States)

    Kogi, Kazutaka

    2007-12-01

    Recent experiences in the action-oriented use of ergonomic checkpoints in different work settings are reviewed. The purpose is to know what features are useful for healthy work design adjusted to each local situation. Based on the review results, common features of ergonomic checkpoints used in participatory training programs for improving workplace conditions in small enterprises, construction sites, home work and agriculture in industrially developing countries in Asia are discussed. These checkpoints generally compile practical improvement options in a broad range of technical areas, such as materials handling, workstation design, physical environment and work organization. Usually, "action checklists" comprising the tiles of the checkpoints are used together. A clear focus is placed on readily applicable low-cost options. Three common features of these various checkpoints appear to be important. First, the checkpoints represent typical good practices in multiple areas. Second, each how-to section of these checkpoints presents simple improvements reflecting basic ergonomic principles. Examples of these principles include easy reach, fewer and faster transport, elbow-level work, coded displays, isolated or screened hazards and shared teamwork. Third, the illustrated checkpoints accompanied by corresponding checklists are used as group work tools in short-term training courses. Many practical improvements achieved are displayed in websites for inter-country work improvement networks. It is suggested to promote the use of locally adjusted checkpoints in various forms of participatory action-oriented training in small-scale workplaces and in agriculture particularly in industrially developing countries. PMID:18572793

  6. Activation of repair and checkpoints by double-strand breaks of DNA. Activational cascade of protein phosphorylation

    International Nuclear Information System (INIS)

    Molecular mechanisms of double-strand breaks repair and checkpoints include phosphorylations of repair and checkpoint-proteins by protein kinases. Chemical modification of proteins has different consequences including activation, changing of affinity to proteins and localization

  7. Centrosome-associated regulators of the G2/M checkpoint as targets for cancer therapy

    Directory of Open Access Journals (Sweden)

    Broaddus Russell R

    2009-02-01

    Full Text Available Abstract In eukaryotic cells, control mechanisms have developed that restrain cell-cycle transitions in response to stress. These regulatory pathways are termed cell-cycle checkpoints. The G2/M checkpoint prevents cells from entering mitosis when DNA is damaged in order to afford these cells an opportunity to repair the damaged DNA before propagating genetic defects to the daughter cells. If the damage is irreparable, checkpoint signaling might activate pathways that lead to apoptosis. Since alteration of cell-cycle control is a hallmark of tumorigenesis, cell-cycle regulators represent potential targets for therapy. The centrosome has recently come into focus as a critical cellular organelle that integrates G2/M checkpoint control and repairs signals in response to DNA damage. A growing number of G2/M checkpoint regulators have been found in the centrosome, suggesting that centrosome has an important role in G2/M checkpoint function. In this review, we discuss centrosome-associated regulators of the G2/M checkpoint, the dysregulation of this checkpoint in cancer, and potential candidate targets for cancer therapy.

  8. Twitter as a Tool to Warn Others about Sobriety Checkpoints: A Pilot Observational Study

    Science.gov (United States)

    Seitz, Christopher M.; Orsini, Muhsin Michael; Fearnow-Kenney, Melodie; Hatzudis, Kiki; Wyrick, David L.

    2012-01-01

    Anecdotal evidence suggests that young people use the website Twitter as a tool to warn drivers about the locations of sobriety checkpoints. Researchers investigated this claim by independently analyzing the website's content regarding a sample of 10 sobriety checkpoints that were conducted in cities throughout the United States during the weekend…

  9. Cdc20 and Cks direct the spindle checkpoint-independent destruction of cyclin A

    NARCIS (Netherlands)

    Wolthuis, Rob; Clay-Farrace, Lori; van Zon, Wouter; Yekezare, Mona; Koop, Lars; Ogink, Janneke; Medema, Rene; Pines, Jonathon

    2008-01-01

    Successful mitosis requires the right protein be degraded at the right time. Central to this is the spindle checkpoint that prevents the destruction of securin and cyclin 131 when there are improperly attached chromosomes. The principal target of the checkpoint is Cdc20, which activates the anaphase

  10. Space Reclamation for Uncoordinated Checkpointing in Message-Passing Systems. Ph.D. Thesis

    Science.gov (United States)

    Wang, Yi-Min

    1993-01-01

    Checkpointing and rollback recovery are techniques that can provide efficient recovery from transient process failures. In a message-passing system, the rollback of a message sender may cause the rollback of the corresponding receiver, and the system needs to roll back to a consistent set of checkpoints called recovery line. If the processes are allowed to take uncoordinated checkpoints, the above rollback propagation may result in the domino effect which prevents recovery line progression. Traditionally, only obsolete checkpoints before the global recovery line can be discarded, and the necessary and sufficient condition for identifying all garbage checkpoints has remained an open problem. A necessary and sufficient condition for achieving optimal garbage collection is derived and it is proved that the number of useful checkpoints is bounded by N(N+1)/2, where N is the number of processes. The approach is based on the maximum-sized antichain model of consistent global checkpoints and the technique of recovery line transformation and decomposition. It is also shown that, for systems requiring message logging to record in-transit messages, the same approach can be used to achieve optimal message log reclamation. As a final topic, a unifying framework is described by considering checkpoint coordination and exploiting piecewise determinism as mechanisms for bounding rollback propagation, and the applicability of the optimal garbage collection algorithm to domino-free recovery protocols is demonstrated.

  11. Cyclin F suppresses B-Myb activity to promote cell cycle checkpoint control

    DEFF Research Database (Denmark)

    Klein, Ditte Kjærsgaard; Hoffmann, Saskia; Ahlskog, Johanna K;

    2015-01-01

    an important role in checkpoint control following ionizing radiation. Cyclin F-depleted cells initiate checkpoint signalling after ionizing radiation, but fail to maintain G2 phase arrest and progress into mitosis prematurely. Importantly, cyclin F suppresses the B-Myb-driven transcriptional programme...

  12. Priming the pancreatic cancer tumor microenvironment for checkpoint-inhibitor immunotherapy

    OpenAIRE

    Lutz, Eric R.; Kinkead, Heather; Jaffee, Elizabeth M.; Zheng, Lei

    2014-01-01

    Single agent immunotherapy is effective against several cancers, but has failed against poorly immunogenic cancers, including pancreatic cancer. Evaluation of pancreatic tumors following treatment with an experimental vaccine (Lutz et al. Cancer Immunology Research 2014) suggests that vaccination primes the tumor microenvironment (TME) for checkpoint-inhibitor immunotherapy, and supports a new platform for evaluating checkpoint-inhibitors in poorly immunogenic cancers.

  13. Identification of inhibitors of checkpoint kinase 1 through template screening.

    Science.gov (United States)

    Matthews, Thomas P; Klair, Suki; Burns, Samantha; Boxall, Kathy; Cherry, Michael; Fisher, Martin; Westwood, Isaac M; Walton, Michael I; McHardy, Tatiana; Cheung, Kwai-Ming J; Van Montfort, Rob; Williams, David; Aherne, G Wynne; Garrett, Michelle D; Reader, John; Collins, Ian

    2009-08-13

    Checkpoint kinase 1 (CHK1) is an oncology target of significant current interest. Inhibition of CHK1 abrogates DNA damage-induced cell cycle checkpoints and sensitizes p53 deficient cancer cells to genotoxic therapies. Using template screening, a fragment-based approach to small molecule hit generation, we have identified multiple CHK1 inhibitor scaffolds suitable for further optimization. The sequential combination of in silico low molecular weight template selection, a high concentration biochemical assay and hit validation through protein-ligand X-ray crystallography provided 13 template hits from an initial in silico screening library of ca. 15000 compounds. The use of appropriate counter-screening to rule out nonspecific aggregation by test compounds was essential for optimum performance of the high concentration bioassay. One low molecular weight, weakly active purine template hit was progressed by iterative structure-based design to give submicromolar pyrazolopyridines with good ligand efficiency and appropriate CHK1-mediated cellular activity in HT29 colon cancer cells. PMID:19572549

  14. Cenp-meta is required for sustained spindle checkpoint

    Directory of Open Access Journals (Sweden)

    Thomas Rubin

    2014-05-01

    Full Text Available Cenp-E is a kinesin-like motor protein required for efficient end-on attachment of kinetochores to the spindle microtubules. Cenp-E immunodepletion in Xenopus mitotic extracts results in the loss of mitotic arrest and massive chromosome missegregation, whereas its depletion in mammalian cells leads to chromosome segregation defects despite the presence of a functional spindle assembly checkpoint (SAC. Cenp-meta has previously been reported to be the Drosophila homolog of vertebrate Cenp-E. In this study, we show that cenp-metaΔ mutant neuroblasts arrest in mitosis when treated with colchicine. cenp-metaΔ mutant cells display a mitotic delay. Yet, despite the persistence of the two checkpoint proteins Mad2 and BubR1 on unattached kinetochores, these cells eventually enter anaphase and give rise to highly aneuploid daughter cells. Indeed, we find that cenp-metaΔ mutant cells display a slow but continuous degradation of cyclin B, which eventually triggers the mitotic exit observed. Thus, our data provide evidence for a role of Cenp-meta in sustaining the SAC response.

  15. The budding yeast protein kinase Ipl1/Aurora allows the absence of tension to activate the spindle checkpoint

    OpenAIRE

    Biggins, Sue; Murray, Andrew W.

    2001-01-01

    The spindle checkpoint prevents cell cycle progression in cells that have mitotic spindle defects. Although several spindle defects activate the spindle checkpoint, the exact nature of the primary signal is unknown. We have found that the budding yeast member of the Aurora protein kinase family, Ipl1p, is required to maintain a subset of spindle checkpoint arrests. Ipl1p is required to maintain the spindle checkpoint that is induced by overexpression of the protein kinase Mps1. Inactivating I...

  16. Apelin-13 inhibits large-conductance Ca2+-activated K+ channels in cerebral artery smooth muscle cells via a PI3-kinase dependent mechanism.

    Directory of Open Access Journals (Sweden)

    Amit Modgil

    Full Text Available Apelin-13 causes vasoconstriction by acting directly on APJ receptors in vascular smooth muscle (VSM cells; however, the ionic mechanisms underlying this action at the cellular level remain unclear. Large-conductance Ca(2+-activated K(+ (BKCa channels in VSM cells are critical regulators of membrane potential and vascular tone. In the present study, we examined the effect of apelin-13 on BK(Ca channel activity in VSM cells, freshly isolated from rat middle cerebral arteries. In whole-cell patch clamp mode, apelin-13 (0.001-1 μM caused concentration-dependent inhibition of BK(Ca in VSM cells. Apelin-13 (0.1 µM significantly decreased BK(Ca current density from 71.25 ± 8.14 pA/pF to 44.52 ± 7.10 pA/pF (n=14 cells, P<0.05. This inhibitory effect of apelin-13 was confirmed by single channel recording in cell-attached patches, in which extracellular application of apelin-13 (0.1 µM decreased the open-state probability (NPo of BK(Ca channels in freshly isolated VSM cells. However, in inside-out patches, extracellular application of apelin-13 (0.1 µM did not alter the NPo of BK(Ca channels, suggesting that the inhibitory effect of apelin-13 on BKCa is not mediated by a direct action on BK(Ca. In whole cell patches, pretreatment of VSM cells with LY-294002, a PI3-kinase inhibitor, markedly attenuated the apelin-13-induced decrease in BK(Ca current density. In addition, treatment of arteries with apelin-13 (0.1 µM significantly increased the ratio of phosphorylated-Akt/total Akt, indicating that apelin-13 significantly increases PI3-kinase activity. Taken together, the data suggest that apelin-13 inhibits BK(Ca channel via a PI3-kinase-dependent signaling pathway in cerebral artery VSM cells, which may contribute to its regulatory action in the control of vascular tone.

  17. Detailed Modeling, Design, and Evaluation of a Scalable Multi-level Checkpointing System

    Energy Technology Data Exchange (ETDEWEB)

    Moody, A T; Bronevetsky, G; Mohror, K M; de Supinski, B R

    2010-04-09

    High-performance computing (HPC) systems are growing more powerful by utilizing more hardware components. As the system mean-time-before-failure correspondingly drops, applications must checkpoint more frequently to make progress. However, as the system memory sizes grow faster than the bandwidth to the parallel file system, the cost of checkpointing begins to dominate application run times. A potential solution to this problem is to use multi-level checkpointing, which employs multiple types of checkpoints with different costs and different levels of resiliency in a single run. The goal is to design light-weight checkpoints to handle the most common failure modes and rely on more expensive checkpoints for less common, but more severe failures. While this approach is theoretically promising, it has not been fully evaluated in a large-scale, production system context. To this end we have designed a system, called the Scalable Checkpoint/Restart (SCR) library, that writes checkpoints to storage on the compute nodes utilizing RAM, Flash, or disk, in addition to the parallel file system. We present the performance and reliability properties of SCR as well as a probabilistic Markov model that predicts its performance on current and future systems. We show that multi-level checkpointing improves efficiency on existing large-scale systems and that this benefit increases as the system size grows. In particular, we developed low-cost checkpoint schemes that are 100x-1000x faster than the parallel file system and effective against 85% of our system failures. This leads to a gain in machine efficiency of up to 35%, and it reduces the the load on the parallel file system by a factor of two on current and future systems.

  18. Parallelization and checkpointing of GPU applications through program transformation

    Energy Technology Data Exchange (ETDEWEB)

    Solano-Quinde, Lizandro Damian [Iowa State Univ., Ames, IA (United States)

    2012-01-01

    GPUs have emerged as a powerful tool for accelerating general-purpose applications. The availability of programming languages that makes writing general-purpose applications for running on GPUs tractable have consolidated GPUs as an alternative for accelerating general purpose applications. Among the areas that have benefited from GPU acceleration are: signal and image processing, computational fluid dynamics, quantum chemistry, and, in general, the High Performance Computing (HPC) Industry. In order to continue to exploit higher levels of parallelism with GPUs, multi-GPU systems are gaining popularity. In this context, single-GPU applications are parallelized for running in multi-GPU systems. Furthermore, multi-GPU systems help to solve the GPU memory limitation for applications with large application memory footprint. Parallelizing single-GPU applications has been approached by libraries that distribute the workload at runtime, however, they impose execution overhead and are not portable. On the other hand, on traditional CPU systems, parallelization has been approached through application transformation at pre-compile time, which enhances the application to distribute the workload at application level and does not have the issues of library-based approaches. Hence, a parallelization scheme for GPU systems based on application transformation is needed. Like any computing engine of today, reliability is also a concern in GPUs. GPUs are vulnerable to transient and permanent failures. Current checkpoint/restart techniques are not suitable for systems with GPUs. Checkpointing for GPU systems present new and interesting challenges, primarily due to the natural differences imposed by the hardware design, the memory subsystem architecture, the massive number of threads, and the limited amount of synchronization among threads. Therefore, a checkpoint/restart technique suitable for GPU systems is needed. The goal of this work is to exploit higher levels of parallelism and

  19. Fueling the engine and releasing the break:combinational therapy of cancer vaccines and immune checkpoint inhibitors

    Institute of Scientific and Technical Information of China (English)

    Jennifer Kleponis; Richard Skelton; Lei Zheng

    2015-01-01

    Immune checkpoint inhibitors are increasingly drawing much attention in the therapeutic development for cancer treatment. However, many cancer patients do not respond to treatments with immune checkpoint inhibitors, partly because of the lack of tumor-inifltrating effector T cells. Cancer vaccines may prime patients for treatments with immune checkpoint inhibitors by inducing effector T-cell infiltration into the tumors and immune checkpoint signals. The combination of cancer vaccine and an immune checkpoint inhibitor may function synergistically to induce more effective antitumor immune responses, and clinical trials to test the combination are currently ongoing.

  20. A novel role for the GTPase-activating protein Bud2 in the spindle position checkpoint.

    Directory of Open Access Journals (Sweden)

    Scott A Nelson

    Full Text Available The spindle position checkpoint (SPC ensures correct mitotic spindle position before allowing mitotic exit in the budding yeast Saccharomyces cerevisiae. In a candidate screen for checkpoint genes, we identified bud2Δ as deficient for the SPC. Bud2 is a GTPase activating protein (GAP, and the only known substrate of Bud2 was Rsr1/Bud1, a Ras-like GTPase and a central component of the bud-site-selection pathway. Mutants lacking Rsr1/Bud1 had no checkpoint defect, as did strains lacking and overexpressing Bud5, a guanine-nucleotide exchange factor (GEF for Rsr1/Bud1. Thus, the checkpoint function of Bud2 is distinct from its role in bud site selection. The catalytic activity of the Bud2 GAP domain was required for the checkpoint, based on the failure of the known catalytic point mutant Bud2(R682A to function in the checkpoint. Based on assays of heterozygous diploids, bud2(R682A, was dominant for loss of checkpoint but recessive for bud-site-selection failure, further indicating a separation of function. Tem1 is a Ras-like protein and is the critical regulator of mitotic exit, sitting atop the mitotic exit network (MEN. Tem1 is a likely target for Bud2, supported by genetic analyses that exclude other Ras-like proteins.

  1. Probabilistic Checkpointing Protocol to Sensor Network Fault-Tolerant

    Directory of Open Access Journals (Sweden)

    Titouna Faiza

    2012-09-01

    Full Text Available A wireless sensor network WSN is a collection of autonomous sensors nodes organized into a cooperative network. A sensor node transmits the data quantity to the sink. Indeed, a failed sink may abort the overall mission of the network. Due to their crucial functions, sinks must be designed and maintained to be robust enough in order to face trouble coming from the harsh environment. Thus, as a keystone of a WSN, a sink has to be provided with ability to recover from failures. In this paper, we propose a new protocol avoiding to the sink to be a central point of failure. First, we model a sensor node failure estimation problem through a causal network. Then, we show how the checkpointing process ensures the recovery of the network. This approach reduces both energy consumption and communication bandwidth requirements, and prolongs the lifetime of WSN. Interesting results are given by simulation

  2. EMODnet MedSea Checkpoint for sustainable Blue Growth

    Science.gov (United States)

    Moussat, Eric; Pinardi, Nadia; Manzella, Giuseppe; Blanc, Frederique

    2016-04-01

    The EMODNET checkpoint is a wide monitoring system assessment activity aiming to support the sustainable Blue Growth at the scale of the European Sea Basins by: 1) Clarifying the observation landscape of all compartments of the marine environment including Air, Water, Seabed, Biota and Human activities, pointing out to the existing programs, national, European and international 2) Evaluating fitness for use indicators that will show the accessibility and usability of observation and modeling data sets and their roles and synergies based upon selected applications by the European Marine Environment Strategy 3) Prioritizing the needs to optimize the overall monitoring Infrastructure (in situ and satellite data collection and assembling, data management and networking, modeling and forecasting, geo-infrastructure) and release recommendations for evolutions to better meet the application requirements in view of sustainable Blue Growth The assessment is designed for : - Institutional stakeholders for decision making on observation and monitoring systems - Data providers and producers to know how their data collected once for a given purpose could fit other user needs - End-users interested in a regional status and possible uses of existing monitoring data Selected end-user applications are of paramount importance for: (i) the blue economy sector (offshore industries, fisheries); (ii) marine environment variability and change (eutrophication, river inputs and ocean climate change impacts); (iii) emergency management (oil spills); and (iv) preservation of natural resources and biodiversity (Marine Protected Areas). End-user applications generate innovative products based on the existing observation landscape. The fitness for use assessment is made thanks to the comparison of the expected product specifications with the quality of the product derived from the selected data. This involves the development of checkpoint information and indicators based on Data quality and

  3. Attachment issues: kinetochore transformations and spindle checkpoint silencing.

    Science.gov (United States)

    Etemad, Banafsheh; Kops, Geert J P L

    2016-04-01

    Cell division culminates in the segregation of duplicated chromosomes in opposite directions prior to cellular fission. This process is guarded by the spindle assembly checkpoint (SAC), which prevents the anaphase of cell division until stable connections between spindle microtubules and the kinetochores of all chromosomes are established. The anaphase inhibitor is generated at unattached kinetochores and inhibitor production is prevented when microtubules are captured. Understanding the molecular changes in the kinetochore that are evoked by microtubule attachments is crucial for understanding the mechanisms of SAC signaling and silencing. Here, we highlight the most recent findings on these events, pinpoint some remaining mysteries, and argue for incorporating holistic views of kinetochore dynamics in order to understand SAC silencing. PMID:26947988

  4. Checkpoint inhibition for colorectal cancer: progress and possibilities.

    Science.gov (United States)

    Paul, Barry; O'Neil, Bert H; McRee, Autumn J

    2016-06-01

    Colorectal cancer (CRC) remains the third most common cause of cancer death in the USA. Despite an increase in the repertoire of treatment options available for CRC, median overall survival has plateaued at approximately 2.5 years. Strategies that engage the patient's native immune system to overcome checkpoint inhibition have proven to be promising in subsets of CRCs, specifically those with mismatch repair deficiency. Further studies are required to determine combinations of standard therapies with immunotherapy drugs and to discover the best biomarkers to predict response. This review provides insight into the progress made in treating patients with advanced CRC with immunotherapeutics and the areas that demand further research to make these drugs more effective in this patient population. PMID:27197538

  5. Checkpoint Blockade in Cancer Immunotherapy: Squaring the Circle

    Directory of Open Access Journals (Sweden)

    Maria A.V. Marzolini

    2015-03-01

    Full Text Available Manipulating the complex interaction between the immune system and tumour cells has been the focus of cancer research for many years, but it is only in the past decade that significant progress has been made in the field of cancer immunotherapy resulting in clinically effective treatments. The blockade of co-inhibitory immune checkpoints, essential for maintaining lymphocyte homeostasis and self-tolerance, by immunomodulatory monoclonal antibodies has resulted in the augmentation of anti-tumour responses. The greatest successes so far have been seen with the blockade of cytotoxic T lymphocyte associated antigen-4, which has resulted in the first Phase III clinical trial showing an overall survival benefit in metastatic melanoma, and in the blockade of the programmed cell death protein-1 axis. This concise review will focus on the clinical advances made by the blockade of these two pathways and their role in current cancer treatment strategies.

  6. A mitotic phosphorylation feedback network connects Cdk1, Plk1, 53BP1, and Chk2 to inactivate the G(2)/M DNA damage checkpoint

    DEFF Research Database (Denmark)

    van Vugt, Marcel A T M; Gardino, Alexandra K; Linding, Rune; Ostheimer, Gerard J; Reinhardt, H Christian; Ong, Shao-En; Tan, Chris S; Miao, Hua; Keezer, Susan M; Li, Jeijin; Pawson, Tony; Lewis, Timothy A; Carr, Steven A; Smerdon, Stephen J; Brummelkamp, Thijn R; Yaffe, Michael B

    2010-01-01

    DNA damage checkpoints arrest cell cycle progression to facilitate DNA repair. The ability to survive genotoxic insults depends not only on the initiation of cell cycle checkpoints but also on checkpoint maintenance. While activation of DNA damage checkpoints has been studied extensively, molecular...

  7. NEK11: linking CHK1 and CDC25A in DNA damage checkpoint signaling

    DEFF Research Database (Denmark)

    Sørensen, Claus Storgaard; Melixetian, Marina; Klein, Ditte Kjaersgaard;

    2010-01-01

    The DNA damage induced G(2)/M checkpoint is an important guardian of the genome that prevents cell division when DNA lesions are present. The checkpoint prevents cells from entering mitosis by degrading CDC25A, a key CDK activator. CDC25A proteolysis is controlled by direct phosphorylation events...... required for beta-TrCP mediated CDC25A polyubiquitylation and degradation. The activity of NEK11 is in turn controlled by CHK1 that activates NEK11 via phosphorylation on serine 273. Since inhibition of NEK11 activity forces checkpoint-arrested cells into mitosis and cell death, NEK11 is, like CHK1, a...

  8. The DNA damage and the DNA replication checkpoints converge at the MBF transcription factor

    OpenAIRE

    Ivanova, Tsvetomira Georgieva, 1978-; Alves-Rodrigues, Isabel; G??mez Escoda, Blanca; Dutta, Chaitali; DeCaprio, James A.; Rhind, Nick; Hidalgo Hernando, Elena; Ayt?? del Olmo, Jos??

    2013-01-01

    In fission yeast cells, Cds1 is the effector kinase of the DNA replication checkpoint. We previously showed that when the DNA replication checkpoint is activated, the repressor Yox1 is phosphorylated and inactivated by Cds1, resulting in activation of MluI-binding factor (MBF)-dependent transcription. This is essential to reinitiate DNA synthesis and for correct G1-to-S transition. Here we show that Cdc10, which is an essential part of the MBF core, is the target of the DNA damage checkpoint....

  9. Defining the optimal murine models to investigate immune checkpoint blockers and their combination with other immunotherapies.

    Science.gov (United States)

    Sanmamed, M F; Chester, C; Melero, I; Kohrt, H

    2016-07-01

    The recent success of checkpoint blockers to treat cancer has demonstrated that the immune system is a critical player in the war against cancer. Historically, anticancer therapeutics have been tested in syngeneic mouse models (with a fully murine immune system) or in immunodeficient mice that allow the engraftment of human xenografts. Animal models with functioning human immune systems are critically needed to more accurately recapitulate the complexity of the human tumor microenvironment. Such models are integral to better predict tumor responses to both immunomodulatory agents and directly antineoplastic therapies. In this regard, the development of humanized models is a promising, novel strategy that offers the possibility of testing checkpoint blockers' capacity and their combination with other antitumor drugs. In this review, we discuss the strengths and weaknesses of the available animal models regarding their capacity to evaluate checkpoint blockers and checkpoint blocker-based combination immunotherapy. PMID:26912558

  10. Coupling end resection with the checkpoint response at DNA double-strand breaks.

    Science.gov (United States)

    Villa, Matteo; Cassani, Corinne; Gobbini, Elisa; Bonetti, Diego; Longhese, Maria Pia

    2016-10-01

    DNA double-strand breaks (DSBs) are a nasty form of damage that needs to be repaired to ensure genome stability. The DSB ends can undergo a strand-biased nucleolytic processing (resection) to generate 3'-ended single-stranded DNA (ssDNA) that channels DSB repair into homologous recombination. Generation of ssDNA also triggers the activation of the DNA damage checkpoint, which couples cell cycle progression with DSB repair. The checkpoint response is intimately linked to DSB resection, as some checkpoint proteins regulate the resection process. The present review will highlight recent works on the mechanism and regulation of DSB resection and its interplays with checkpoint activation/inactivation in budding yeast. PMID:27141941

  11. The checkpoint Saccharomyces cerevisiae Rad9 protein contains a tandem tudor domain that recognizes DNA.

    OpenAIRE

    Lancelot, Nathalie; Charier, Gaëlle; Couprie, Joël; Duband-Goulet, Isabelle; Alpha-Bazin, Béatrice; Quémeneur, Eric; Ma, Emilie; Marsolier-Kergoat, Marie-Claude; Ropars, Virginie; Charbonnier, Jean-Baptiste; Miron, Simona; Craescu, Constantin,; Callebaut, Isabelle; Gilquin, Bernard; Zinn-Justin, Sophie

    2007-01-01

    DNA damage checkpoints are signal transduction pathways that are activated after genotoxic insults to protect genomic integrity. At the site of DNA damage, 'mediator' proteins are in charge of recruiting 'signal transducers' to molecules 'sensing' the damage. Budding yeast Rad9, fission yeast Crb2 and metazoan 53BP1 are presented as mediators involved in the activation of checkpoint kinases. Here we show that, despite low sequence conservation, Rad9 exhibits a tandem tudor domain structurally...

  12. Cambridge checkpoint English revision guide for the Cambridge secondary 1 test

    CERN Document Server

    Reynolds, John

    2013-01-01

    With Checkpoint English Revision Guide for the Cambridge Secondary 1 test you can aim for the best grade with the help of relevant and accessible notes, examiner advice plus questions and answers on each key topic. - Clear explanations of every topic covered in the Cambridge Secondary 1 Checkpoint English syllabus. - Builds revision skills you need for success in the test. - Exam tips wirtten by test setters and examiners giving you their expert advice. This text has not been through the Cambridge endorsement process.

  13. Cambridge checkpoint maths revision guide for the Cambridge secondary 1 test

    CERN Document Server

    Smith, Alan

    2013-01-01

    With Checkpoint Maths Revision Guide for the Cambridge Secondary 1 test you can aim for the best grade with the help of relevant and accessible notes, examiner advice plus questions and answers on each key topic. - Clear explanations of every topic covered in the Cambridge Secondary 1 Checkpoint Maths syllabus. - Builds revision skills you need for success in the test. - Exam tips wirtten by test setters and examiners giving you their expert advice. This text has not been through the Cambridge endorsement process.

  14. The pachytene checkpoint prevents accumulation and phosphorylation of the meiosis-specific transcription factor Ndt80

    OpenAIRE

    Tung, Kuei-Shu; Hong, Eun-Jin Erica; Roeder, G. Shirleen

    2000-01-01

    In budding yeast, many mutants defective in meiotic recombination and chromosome synapsis undergo checkpoint-mediated arrest at the pachytene stage of meiotic prophase. We recovered the NDT80 gene in a screen for genes whose overexpression bypasses the pachytene checkpoint. Ndt80 is a meiosis-specific transcription factor that promotes expression of genes required for exit from pachytene and entry into meiosis I. Herein, we show that the Ndt80 protein accumulates a...

  15. A role for Ddc1 in signaling meiotic double-strand breaks at the pachytene checkpoint

    OpenAIRE

    Hong, Eun-Jin Erica; Roeder, G. Shirleen

    2002-01-01

    The pachytene checkpoint prevents meiotic cell cycle progression in response to unrepaired recombination intermediates. We show that Ddc1 is required for the pachytene checkpoint in Saccharomyces cerevisiae. During meiotic prophase, Ddc1 localizes to chromosomes and becomes phosphorylated; these events depend on the formation and processing of double-strand breaks (DSBs). Ddc1 colocalizes with Rad51, a DSB-repair protein, indicating that Ddc1 associates with sites of DSB repair. The Rad24 che...

  16. A Minimum-Process Coordinated Checkpointing Protocol For Mobile Distributed System

    Directory of Open Access Journals (Sweden)

    Praveen Kumar

    2010-05-01

    Full Text Available While dealing with Mobile Distributed systems, we come across some issues like: mobility, low bandwidth of wireless channels and lack of stable storage on mobile nodes, disconnections, limited battery power and high failure rate of mobile nodes. These issues make traditional checkpointing techniques designed for Distributed systems unsuitable for Mobile environments. In this paper, we design a minimum process algorithm for Mobile Distributed systems, where no useless checkpoints are taken and an effort has been made to optimize the blocking of processes. We propose to delay the processing of selective messages at the receiver end only during the checkpointing period. A Process is allowed to perform its normal computations and send messages during its blocking period. In this way, we try to keep blocking of processes to bare minimum. We captured the transitive dependencies during the normal execution by piggybacking dependency vectors onto computational messages. In this way, we try to reduce the Checkpointing time by avoiding formation of Checkpointing tree. The Z-dependencies are well taken care of. The proposed scheme forces zero useless checkpoints at the cost of very small blocking.

  17. Human cytomegalovirus inhibits a DNA damage response by mislocalizing checkpoint proteins

    Science.gov (United States)

    Gaspar, Miguel; Shenk, Thomas

    2006-02-01

    The DNA damage checkpoint pathway responds to DNA damage and induces a cell cycle arrest to allow time for DNA repair. Several viruses are known to activate or modulate this cellular response. Here we show that the ataxia-telangiectasia mutated checkpoint pathway, which responds to double-strand breaks in DNA, is activated in response to human cytomegalovirus DNA replication. However, this activation does not propagate through the pathway; it is blocked at the level of the effector kinase, checkpoint kinase 2 (Chk2). Late after infection, several checkpoint proteins, including ataxia-telangiectasia mutated and Chk2, are mislocalized to a cytoplasmic virus assembly zone, where they are colocalized with virion structural proteins. This colocalization was confirmed by immunoprecipitation of virion proteins with an antibody that recognizes Chk2. Virus replication was resistant to ionizing radiation, which causes double-strand breaks in DNA. We propose that human CMV DNA replication activates the checkpoint response to DNA double-strand breaks, and the virus responds by altering the localization of checkpoint proteins to the cytoplasm and thereby inhibiting the signaling pathway. ionizing radiation | ataxia-telangiectasia mutated pathway

  18. A Cdc2 dependent checkpoint maintains diploidy in Drosophila.

    Science.gov (United States)

    Hayashi, S

    1996-04-01

    DNA replication in G2 does not normally occur due to the checkpoint control. To elucidate its mechanism, the functions of the escargot and Dmcdc2 genes of Drosophila were studied. When escargot function was eliminated, diploid imaginal cells that were arrested in G2 lost Cyclin A, a regulatory subunit of G2/M cdk, and entered an endocycle. escargot genetically interacted with Dmcdc2 which encodes a catalytic subunit of G2/M cdk. The mutant phenotypes of Dmcdc2 itself was similar to those of escargot: many diploid cells in imaginal discs, salivary glands and the central nervous system entered an endocycle and sometimes formed polytene chromosomes. Since mitotically quiescent abdominal histoblasts still required Dmcdc2 to remain diploid, the inhibitory activity of G2/M cdk on DNA replication appeared to be separable from its activity as the mitosis promoting factor. These results suggest that in G2, escargot is required to maintain a high level of G2/M cdk that actively inhibits the entry into S phase. PMID:8620832

  19. Checkpoint Activation of an Unconventional DNA Replication Program in Tetrahymena.

    Science.gov (United States)

    Sandoval, Pamela Y; Lee, Po-Hsuen; Meng, Xiangzhou; Kapler, Geoffrey M

    2015-07-01

    The intra-S phase checkpoint kinase of metazoa and yeast, ATR/MEC1, protects chromosomes from DNA damage and replication stress by phosphorylating subunits of the replicative helicase, MCM2-7. Here we describe an unprecedented ATR-dependent pathway in Tetrahymena thermophila in which the essential pre-replicative complex proteins, Orc1p, Orc2p and Mcm6p are degraded in hydroxyurea-treated S phase cells. Chromosomes undergo global changes during HU-arrest, including phosphorylation of histone H2A.X, deacetylation of histone H3, and an apparent diminution in DNA content that can be blocked by the deacetylase inhibitor sodium butyrate. Most remarkably, the cell cycle rapidly resumes upon hydroxyurea removal, and the entire genome is replicated prior to replenishment of ORC and MCMs. While stalled replication forks are elongated under these conditions, DNA fiber imaging revealed that most replicating molecules are produced by new initiation events. Furthermore, the sole origin in the ribosomal DNA minichromosome is inactive and replication appears to initiate near the rRNA promoter. The collective data raise the possibility that replication initiation occurs by an ORC-independent mechanism during the recovery from HU-induced replication stress. PMID:26218270

  20. The Meiotic Recombination Checkpoint Suppresses NHK-1 Kinase to Prevent Reorganisation of the Oocyte Nucleus in Drosophila

    OpenAIRE

    Lancaster, Oscar M.; Breuer, Manuel; Cullen, C. Fiona; Ito, Takashi; Ohkura, Hiroyuki

    2010-01-01

    The meiotic recombination checkpoint is a signalling pathway that blocks meiotic progression when the repair of DNA breaks formed during recombination is delayed. In comparison to the signalling pathway itself, however, the molecular targets of the checkpoint that control meiotic progression are not well understood in metazoans. In Drosophila, activation of the meiotic checkpoint is known to prevent formation of the karyosome, a meiosis-specific organisation of chromosomes, but the molecular ...

  1. Smurf2 as a novel mitotic regulator: From the spindle assembly checkpoint to tumorigenesis

    Directory of Open Access Journals (Sweden)

    Moore Finola E

    2009-07-01

    Full Text Available Abstract The execution of the mitotic program with high fidelity is dependent upon precise spatiotemporal regulation of posttranslational protein modifications. For example, the timely polyubiquitination of critical mitotic regulators by Anaphase Promoting Complex/Cyclosome (APC/C is essential for the metaphase to anaphase transition and mitotic exit. The spindle assembly checkpoint prevents unscheduled activity of APC/C-Cdc20 in early mitosis, allowing bipolar attachment of kinetochores to mitotic spindle and facilitating equal segregation of sister chromatids. The critical effector of the spindle checkpoint, Mitotic arrest deficient 2 (Mad2, is recruited to unattached kinetochores forming a complex with other regulatory proteins to efficiently and cooperatively inhibit APC/C-Cdc20. A weakened and/or dysfunctional spindle checkpoint has been linked to the development of genomic instability in both cell culture and animal models, and evidence suggests that aberrant regulation of the spindle checkpoint plays a critical role in human carcinogenesis. Recent studies have illuminated a network of both degradative and non-degradative ubiquitination events that regulate the metaphase to anaphase transition and mitotic exit. Within this context, our recent work showed that the HECT (Homologous to E6-AP C-terminus-family E3 ligase Smurf2 (Smad specific ubiquitin regulatory factor 2, known as a negative regulator of transforming growth factor-beta (TGF-β signaling, is required for a functional spindle checkpoint by promoting the functional localization and stability of Mad2. Here we discuss putative models explaining the role of Smurf2 as a new regulator in the spindle checkpoint. The dynamic mitotic localization of Smurf2 to the centrosome and other critical mitotic structures provides implications about mitotic checkpoint control dependent on various ubiquitination events. Finally, deregulated Smurf2 activity may contribute to carcinogenesis by

  2. In-silico modeling of the mitotic spindle assembly checkpoint.

    Directory of Open Access Journals (Sweden)

    Bashar Ibrahim

    Full Text Available BACKGROUND: The Mitotic Spindle Assembly Checkpoint ((MSAC is an evolutionary conserved mechanism that ensures the correct segregation of chromosomes by restraining cell cycle progression from entering anaphase until all chromosomes have made proper bipolar attachments to the mitotic spindle. Its malfunction can lead to cancer. PRINCIPLE FINDINGS: We have constructed and validated for the human (MSAC mechanism an in silico dynamical model, integrating 11 proteins and complexes. The model incorporates the perspectives of three central control pathways, namely Mad1/Mad2 induced Cdc20 sequestering based on the Template Model, MCC formation, and APC inhibition. Originating from the biochemical reactions for the underlying molecular processes, non-linear ordinary differential equations for the concentrations of 11 proteins and complexes of the (MSAC are derived. Most of the kinetic constants are taken from literature, the remaining four unknown parameters are derived by an evolutionary optimization procedure for an objective function describing the dynamics of the APC:Cdc20 complex. MCC:APC dissociation is described by two alternatives, namely the "Dissociation" and the "Convey" model variants. The attachment of the kinetochore to microtubuli is simulated by a switching parameter silencing those reactions which are stopped by the attachment. For both, the Dissociation and the Convey variants, we compare two different scenarios concerning the microtubule attachment dependent control of the dissociation reaction. Our model is validated by simulation of ten perturbation experiments. CONCLUSION: Only in the controlled case, our models show (MSAC behaviour at meta- to anaphase transition in agreement with experimental observations. Our simulations revealed that for (MSAC activation, Cdc20 is not fully sequestered; instead APC is inhibited by MCC binding.

  3. Preserving Yeast Genetic Heritage through DNA Damage Checkpoint Regulation and Telomere Maintenance

    Directory of Open Access Journals (Sweden)

    Huilin Zhou

    2012-10-01

    Full Text Available In order to preserve genome integrity, extrinsic or intrinsic DNA damages must be repaired before they accumulate in cells and trigger other mutations and genome rearrangements. Eukaryotic cells are able to respond to different genotoxic stresses as well as to single DNA double strand breaks (DSBs, suggesting highly sensitive and robust mechanisms to detect lesions that trigger a signal transduction cascade which, in turn, controls the DNA damage response (DDR. Furthermore, cells must be able to distinguish natural chromosomal ends from DNA DSBs in order to prevent inappropriate checkpoint activation, DDR and chromosomal rearrangements. Since the original discovery of RAD9, the first DNA damage checkpoint gene identified in Saccharomyces cerevisiae, many genes that have a role in this pathway have been identified, including MRC1, MEC3, RAD24, RAD53, DUN1, MEC1 and TEL1. Extensive studies have established most of the genetic basis of the DNA damage checkpoint and uncovered its different functions in cell cycle regulation, DNA replication and repair, and telomere maintenance. However, major questions concerning the regulation and functions of the DNA damage checkpoint remain to be answered. First, how is the checkpoint activity coupled to DNA replication and repair? Second, how do cells distinguish natural chromosome ends from deleterious DNA DSBs? In this review we will examine primarily studies performed using Saccharomyces cerevisiae as a model system.

  4. Immune checkpoints aberrations and gastric cancer; assessment of prognostic value and evaluation of therapeutic potentials.

    Science.gov (United States)

    Abdel-Rahman, Omar

    2016-01-01

    Till now, the prognosis of advanced gastric cancer looked dreadful; thus the search for newer better approaches for this lethal disease has been a strategic target for cancer researchers. In recent years, important immunobiological aspects of the tumor have been revealed with the subsequent proposal of immune check point inhibitors to target these pathways. Clinically, unselected use of immune checkpoint inhibitors in gastric cancer has been deemed with failure; in contrast to the clear success of more recent studies reporting on the use of pembrolizumab in molecularly selected patients. This may illustrate that any future use of immune checkpoint inhibitors in gastric cancer has to be molecularly supported. This review provides a delicate dissection of the clinical and immunobiological considerations underlying the use of these agents in addition to a thorough review of the published clinical data of immune checkpoint inhibitors in gastric cancer. PMID:26321371

  5. The Replication Checkpoint Prevents Two Types of Fork Collapse without Regulating Replisome Stability.

    Science.gov (United States)

    Dungrawala, Huzefa; Rose, Kristie L; Bhat, Kamakoti P; Mohni, Kareem N; Glick, Gloria G; Couch, Frank B; Cortez, David

    2015-09-17

    The ATR replication checkpoint ensures that stalled forks remain stable when replisome movement is impeded. Using an improved iPOND protocol combined with SILAC mass spectrometry, we characterized human replisome dynamics in response to fork stalling. Our data provide a quantitative picture of the replisome and replication stress response proteomes in 32 experimental conditions. Importantly, rather than stabilize the replisome, the checkpoint prevents two distinct types of fork collapse. Unsupervised hierarchical clustering of protein abundance on nascent DNA is sufficient to identify protein complexes and place newly identified replisome-associated proteins into functional pathways. As an example, we demonstrate that ZNF644 complexes with the G9a/GLP methyltransferase at replication forks and is needed to prevent replication-associated DNA damage. Our data reveal how the replication checkpoint preserves genome integrity, provide insights into the mechanism of action of ATR inhibitors, and will be a useful resource for replication, DNA repair, and chromatin investigators. PMID:26365379

  6. Blocking CHK1 Expression Induces Apoptosis and Abrogates the G2 Checkpoint Mechanism

    Directory of Open Access Journals (Sweden)

    Yan Luo

    2001-01-01

    Full Text Available Checkpoint kinase 1 (Chki is a checkpoint gene that is activated after DNA damage. It phosphorylates and inactivates the Cdc2 activating phosphatase Cdc25C. This in turn inactivates Cdc2, which leads to G2/M arrest. We report that blocking Chki expression by antisense or ribozymes in mammalian cells induces apoptosis and interferes with the G2/M arrest induced by adriamycin. The Chki inhibitor UCN-01 also blocks the G2 arrest after DNA damage and renders cells more susceptible to adriamycin. These results indicate that Chki is an essential gene for the checkpoint mechanism during normal cell proliferation as well as in the DNA damage response.

  7. Hierarchical Non-blocking Coordinated Checkpointing Algorithms for Mobile Distributed Computing

    Directory of Open Access Journals (Sweden)

    Parveen Kumar

    2010-01-01

    Full Text Available Mobile system typically uses wireless communication which is based on electromagnetic waves and utilizes a shared broadcast medium. This has made possible creating a mobile distributed computing environment and has brought us several new challenges in distributed protocol design. So many issues such as range of transmission, limited power supply due to battery capacity and mobility of processes. These new issue makes traditional recovery algorithm unsuitable. In this paper, we propose hierarchical non blocking coordinated checkpointing algorithms suitable for mobile distributed computing. The algorithm is non-blocking, requires minimum message logging, has minimum stable storage requirement and produce a consistent set of checkpoints. This algorithm requires minimum number of processes to take checkpoint.

  8. Combining targeted therapy and immune checkpoint inhibitors in the treatment of metastatic melanoma

    Institute of Scientific and Technical Information of China (English)

    Teresa Kim; Rodabe N Amaria; Christine Spencer; Alexandre Reuben; Zachary A Cooper; Jennifer A Wargo

    2014-01-01

    Melanoma is the deadliest form of skin cancer and has an incidence that is rising faster than any other solid tumor. Metastatic melanoma treatment has considerably progressed in the past ifve years with the introduction of targeted therapy (BARF and MEK inhibitors) and immune checkpoint blockade (anti-CTLA4, anti-PD-1, and anti-PD-L1). However, each treatment modality has limitations. Treatment with targeted therapy has been associated with a high response rate, but with short-term responses. Conversely, treatment with immune checkpoint blockade has a lower response rate, but with long-term responses. Targeted therapy affects antitumor immunity, and synergy may exist when targeted therapy is combined with immunotherapy. hTis article presents a brief review of the rationale and evidence for the potential synergy between targeted therapy and immune checkpoint blockade. Challenges and directions for future studies are also proposed.

  9. Use of checkpoint-restart for complex HEP software on traditional architectures and Intel MIC

    International Nuclear Information System (INIS)

    Process checkpoint-restart is a technology with great potential for use in HEP workflows. Use cases include debugging, reducing the startup time of applications both in offline batch jobs and the High Level Trigger, permitting job preemption in environments where spare CPU cycles are being used opportunistically and efficient scheduling of a mix of multicore and single-threaded jobs. We report on tests of checkpoint-restart technology using CMS software, Geant4-MT (multi-threaded Geant4), and the DMTCP (Distributed Multithreaded Checkpointing) package. We analyze both single- and multi-threaded applications and test on both standard Intel x86 architectures and on Intel MIC. The tests with multi-threaded applications on Intel MIC are used to consider scalability and performance. These are considered an indicator of what the future may hold for many-core computing

  10. RNA interference regulates the cell cycle checkpoint through the RNA export factor, Ptr1, in fission yeast

    International Nuclear Information System (INIS)

    Highlights: ► RNAi is linked to the cell cycle checkpoint in fission yeast. ► Ptr1 co-purifies with Ago1. ► The ptr1-1 mutation impairs the checkpoint but does not affect gene silencing. ► ago1+ and ptr1+ regulate the cell cycle checkpoint via the same pathway. ► Mutations in ago1+ and ptr1+ lead to the nuclear accumulation of poly(A)+ RNAs. -- Abstract: Ago1, an effector protein of RNA interference (RNAi), regulates heterochromatin silencing and cell cycle arrest in fission yeast. However, the mechanism by which Ago1 controls cell cycle checkpoint following hydroxyurea (HU) treatment has not been elucidated. In this study, we show that Ago1 and other RNAi factors control cell cycle checkpoint following HU treatment via a mechanism independent of silencing. While silencing requires dcr1+, the overexpression of ago1+ alleviated the cell cycle defect in dcr1Δ. Ago1 interacted with the mRNA export factor, Ptr1. The ptr1-1 mutation impaired cell cycle checkpoint but gene silencing was unaffected. Genetic analysis revealed that the regulation of cell cycle checkpoint by ago1+ is dependent on ptr1+. Nuclear accumulation of poly(A)+ RNAs was detected in mutants of ago1+ and ptr1+, suggesting there is a functional link between the cell cycle checkpoint and RNAi-mediated RNA quality control.

  11. The anaphase inhibitor of Saccharomyces cerevisiae Pds1p is a target of the DNA damage checkpoint pathway

    International Nuclear Information System (INIS)

    Inhibition of DNA replication and physical DNA damage induce checkpoint responses that arrest cell cycle progression at two different stages. In Saccharomyces cerevisiae, the execution of both checkpoint responses requires the Mec1 and Rad53 proteins. This observation led to the suggestion that these checkpoint responses are mediated through a common signal transduction pathway. However, because the checkpoint-induced arrests occur at different cell cycle stages, the downstream effectors mediating these arrests are likely to be distinct. We have previously shown that the S. cerevisiae protein Pds1p is an anaphase inhibitor and is essential for cell cycle arrest in mitosis in the presence DNA damage. Herein we show that DNA damage, but not inhibition of DNA replication, induces the phosphorylation of Pds1p. Analyses of Pds1p phosphorylation in different checkpoint mutants reveal that in the presence of DNA damage, Pds1p is phosphorylated in a Mec1p- and Rad9p-dependent hut Rad53p-independent manner. Our data place Pds1p and Rad53p on parallel branches of the DNA damage checkpoint pathway. We suggest that Pds1p is a downstream target of the DNA damage checkpoint pathway and that it is involved in implementing the DNA damage checkpoint arrest specifically in mitosis

  12. Functions of spindle check-point and its relationship to chromosome instability

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    It is generally believed that the equal distribution of genetic materials to two daughter cells during mitosis is the key to cell health and development. During the dynamic process, spindle checkpoint plays a very important role in chromosome movements and final sister chromatid separation. The equal and precise segregation of chromosomes contributes to the genomic stability while aberrant separations result in chromosome instability that causes pathogenesis of certain diseases such as Down's syndrome and cancers. Kinetochore and its regulatory proteins consist of the spindle checkpoint and determine the spatial and temporal orders of chromosome segregation.

  13. Constitutive Cdk2 activity promotes aneuploidy while altering the spindle assembly and tetraploidy checkpoints

    DEFF Research Database (Denmark)

    Jahn, Stephan C; Corsino, Patrick E; Davis, Bradley J;

    2013-01-01

    The cell has many mechanisms for protecting the integrity of its genome. These mechanisms are often weakened or absent in many cancers, leading to high rates of chromosomal instability in tumors. Control of the cell cycle is crucial for the function of these checkpoints, and is frequently lost in...... instability. Expression of these complexes in the MCF10A cell line leads to retinoblastoma protein (Rb) hyperphosphorylation, a subsequent increase in proliferation rate, and increased expression of the spindle assembly checkpoint protein Mad2. This results in a strengthening of the spindle assembly...

  14. Role for Rif1 in the checkpoint response to damaged DNA in Xenopus egg extracts

    OpenAIRE

    Kumar, Sanjay; Yoo, Hae Yong; Kumagai, Akiko; Shevchenko, Anna; Shevchenko, Andrej; Dunphy, William G.

    2012-01-01

    TopBP1 is critical for both DNA replication and checkpoint regulation in vertebrate cells. In this study, we have identified Rif1 as a binding partner of TopBP1 in Xenopus egg extracts. In addition, Rif1 also interacts with both ATM and the Mre11-Rad50-Nbs1 (MRN) complex, which are key regulators of checkpoint responses to double-stranded DNA breaks (DSBs). Depletion of Rif1 from egg extracts compromises the activation of Chk1 in response to DSBs but not stalled replication forks. Removal of ...

  15. Defective DNA repair increases susceptibility to senescence through extension of Chk1-mediated G2 checkpoint activation.

    Science.gov (United States)

    Johmura, Yoshikazu; Yamashita, Emiri; Shimada, Midori; Nakanishi, Keiko; Nakanishi, Makoto

    2016-01-01

    Susceptibility to senescence caused by defective DNA repair is a major hallmark of progeroid syndrome patients, but molecular mechanisms of how defective DNA repair predisposes to senescence are largely unknown. We demonstrate here that suppression of DNA repair pathways extends the duration of Chk1-dependent G2 checkpoint activation and sensitizes cells to senescence through enhancement of mitosis skipping. Extension of G2 checkpoint activation by introduction of the TopBP1 activation domain and the nondegradable mutant of Claspin sensitizes cells to senescence. In contrast, a shortening of G2 checkpoint activation by expression of SIRT6 or depletion of OTUB2 reduces susceptibility to senescence. Fibroblasts from progeroid syndromes tested shows a correlation between an extension of G2 checkpoint activation and an increase in the susceptibility to senescence. These results suggest that extension of G2 checkpoint activation caused by defective DNA repair is critical for senescence predisposition in progeroid syndrome patients. PMID:27507734

  16. Cells bearing chromosome aberrations lacking one telomere are selectively blocked at the G2/M checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Rodriguez, Pilar [Unitat de Biologia Cel.lular, Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain); Barquinero, Joan Francesc [Unitat d' Antropologia Biologica, Departament de Biologia Animal, Biologia Vegetal i Ecologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain); Duran, Assumpta [Unitat de Biologia Cel.lular, Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain); Caballin, Maria Rosa [Unitat d' Antropologia Biologica, Departament de Biologia Animal, Biologia Vegetal i Ecologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain); Ribas, Montserrat [Servei de Radiofisica i Radioproteccio de l' Hospital de la Santa Creu i Sant Pau, 08025 Barcelona (Spain); Barrios, Leonardo, E-mail: Lleonard.Barrios@uab.cat [Unitat de Biologia Cel.lular, Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain)

    2009-11-02

    Cell cycle checkpoints are part of the cellular mechanisms to maintain genomic integrity. After ionizing radiation exposure, the cells can show delay or arrest in their progression through the cell cycle, as well as an activation of the DNA repair machinery in order to reduce the damage. The G2/M checkpoint prevents G2 cells entering mitosis until the DNA damage has been reduced. The present study evaluates which G0 radiation-induced chromosome aberrations are negatively selected in the G2/M checkpoint. For this purpose, peripheral blood samples were irradiated at 1 and 3 Gy of {gamma}-rays, and lymphocytes were cultured for 48 h. Calyculin-A and Colcemid were used to analyze, in the same slide, cells in G2 and M. Chromosome spreads were consecutively analyzed by solid stain, pancentromeric and pantelomeric FISH and mFISH. The results show that the frequency of incomplete chromosome elements, those lacking a telomeric signal at one end, decreases abruptly from G2 to M. This indicates that cells with incomplete chromosome elements can progress from G0 to G2, but at the G2/M checkpoint suffer a strong negative selection.

  17. Differential impact of diverse anticancer chemotherapeutics on the Cdc25A-degradation checkpoint pathway

    International Nuclear Information System (INIS)

    When exposed to DNA-damaging insults such as ionizing radiation (IR) or ultraviolet light (UV), mammalian cells activate checkpoint pathways to halt cell cycle progression or induce cell death. Here we examined the ability of five commonly used anticancer drugs with different mechanisms of action to activate the Chk1/Chk2-Cdc25A-CDK2/cyclin E cell cycle checkpoint pathway, previously shown to be induced by IR or UV. Whereas exposure of human cells to topoisomerase inhibitors camptothecin, etoposide, or adriamycin resulted in rapid (within 1 h) activation of the pathway including degradation of the Cdc25A phosphatase and inhibition of cyclin E/CDK2 kinase activity, taxol failed to activate this checkpoint even after a prolonged treatment. Unexpectedly, although the alkylating agent cisplatin also induced degradation of Cdc25A (albeit delayed, after 8-12 h), cyclin E/CDK2 activity was elevated and DNA synthesis continued, a phenomena that correlated with increased E2F1 protein levels and consequently enhanced expression of cyclin E. These results reveal a differential impact of various classes of anticancer chemotherapeutics on the Cdc25A-degradation pathway, and indicate that the kinetics of checkpoint induction, and the relative balance of key components within the DNA damage response network may dictate whether the treated cells arrest their cell cycle progression

  18. A phospho-proteomic screen identifies substrates of the checkpoint kinase Chk1

    DEFF Research Database (Denmark)

    Blasius, Melanie; Forment, Josep V; Thakkar, Neha;

    2011-01-01

    BACKGROUND: The cell-cycle checkpoint kinase Chk1 is essential in mammalian cells due to its roles in controlling processes such as DNA replication, mitosis and DNA-damage responses. Despite its paramount importance, how Chk1 controls these functions remains unclear, mainly because very few Chk1...

  19. Tumor suppressor protein C53 antagonizes checkpoint kinases to promote cyclin-dependent kinase 1 activation

    Institute of Scientific and Technical Information of China (English)

    Hai Jiang; Jianchun Wu; Chen He; Wending Yang; Honglin Li

    2009-01-01

    Cyclin-dependent kinase 1 (Cdk1)/cyclin B1 complex is the driving force for mitotic entry, and its activation is tightly regulated by the G2/M checkpoint. We originally reported that a novel protein C53 (also known as Cdk5rap3 and LZAP) potentiates DNA damage-induced cell death by modulating the G2/M checkpoint. More recently, Wang et al. (2007) found that C53/LZAP may function as a tumor suppressor by way of inhibiting NF-kB signaling. We report here the identification of C53 protein as a novel regulator of Cdk1 activation. We found that knockdown of C53 protein causes delayed Cdkl activation and mitotic entry. During DNA damage response, activation of checkpoint kinase 1 and 2 (Chk1 and Chk2) is partially inhibited by C53 overexpression. Intriguingly, we found that C53 interacts with Chkl and antagonizes its function. Moreover, a portion of C53 protein is localized at the centrosome, and centrosome-targeting C53 potently promotes local Cdk1 activation. Taken together, our results strongly suggest that C53 is a novel negative regulator of checkpoint response. By counteracting Chk1, C53 promotes Cdk1 activation and mitotic entry in both unperturbed cell-cycle progression and DNA damage response.

  20. Clonal neoantigens elicit T cell immunoreactivity and sensitivity to immune checkpoint blockade

    DEFF Research Database (Denmark)

    McGranahan, Nicholas; Furness, Andrew J S; Rosenthal, Rachel;

    2016-01-01

    As tumors grow, they acquire mutations, some of which create neoantigens that influence the response of patients to immune checkpoint inhibitors. We explored the impact of neoantigen intratumor heterogeneity (ITH) on antitumor immunity. Through integrated analysis of ITH and neoantigen burden, we...

  1. The DNA damage and the DNA replication checkpoints converge at the MBF transcription factor

    Science.gov (United States)

    Ivanova, Tsvetomira; Alves-Rodrigues, Isabel; Gómez-Escoda, Blanca; Dutta, Chaitali; DeCaprio, James A.; Rhind, Nick; Hidalgo, Elena; Ayté, José

    2013-01-01

    In fission yeast cells, Cds1 is the effector kinase of the DNA replication checkpoint. We previously showed that when the DNA replication checkpoint is activated, the repressor Yox1 is phosphorylated and inactivated by Cds1, resulting in activation of MluI-binding factor (MBF)–dependent transcription. This is essential to reinitiate DNA synthesis and for correct G1-to-S transition. Here we show that Cdc10, which is an essential part of the MBF core, is the target of the DNA damage checkpoint. When fission yeast cells are treated with DNA-damaging agents, Chk1 is activated and phosphorylates Cdc10 at its carboxy-terminal domain. This modification is responsible for the repression of MBF-dependent transcription through induced release of MBF from chromatin. This inactivation of MBF is important for survival of cells challenged with DNA-damaging agents. Thus Yox1 and Cdc10 couple normal cell cycle regulation in unperturbed conditions and the DNA replication and DNA damage checkpoints into a single transcriptional complex. PMID:24006488

  2. The DNA damage and the DNA replication checkpoints converge at the MBF transcription factor.

    Science.gov (United States)

    Ivanova, Tsvetomira; Alves-Rodrigues, Isabel; Gómez-Escoda, Blanca; Dutta, Chaitali; DeCaprio, James A; Rhind, Nick; Hidalgo, Elena; Ayté, José

    2013-11-01

    In fission yeast cells, Cds1 is the effector kinase of the DNA replication checkpoint. We previously showed that when the DNA replication checkpoint is activated, the repressor Yox1 is phosphorylated and inactivated by Cds1, resulting in activation of MluI-binding factor (MBF)-dependent transcription. This is essential to reinitiate DNA synthesis and for correct G1-to-S transition. Here we show that Cdc10, which is an essential part of the MBF core, is the target of the DNA damage checkpoint. When fission yeast cells are treated with DNA-damaging agents, Chk1 is activated and phosphorylates Cdc10 at its carboxy-terminal domain. This modification is responsible for the repression of MBF-dependent transcription through induced release of MBF from chromatin. This inactivation of MBF is important for survival of cells challenged with DNA-damaging agents. Thus Yox1 and Cdc10 couple normal cell cycle regulation in unperturbed conditions and the DNA replication and DNA damage checkpoints into a single transcriptional complex. PMID:24006488

  3. Synthesis of Fault-Tolerant Embedded Systems with Checkpointing and Replication

    DEFF Research Database (Denmark)

    Izosimov, Viacheslav; Pop, Paul; Eles, Petru; Peng, Zebo

    We present an approach to the synthesis of fault-tolerant hard real-time systems for safety-critical applications. We use checkpointing with rollback recovery and active replication for tolerating transient faults. Processes are statically scheduled and communications are performed using the time...

  4. Cells bearing chromosome aberrations lacking one telomere are selectively blocked at the G2/M checkpoint

    International Nuclear Information System (INIS)

    Cell cycle checkpoints are part of the cellular mechanisms to maintain genomic integrity. After ionizing radiation exposure, the cells can show delay or arrest in their progression through the cell cycle, as well as an activation of the DNA repair machinery in order to reduce the damage. The G2/M checkpoint prevents G2 cells entering mitosis until the DNA damage has been reduced. The present study evaluates which G0 radiation-induced chromosome aberrations are negatively selected in the G2/M checkpoint. For this purpose, peripheral blood samples were irradiated at 1 and 3 Gy of γ-rays, and lymphocytes were cultured for 48 h. Calyculin-A and Colcemid were used to analyze, in the same slide, cells in G2 and M. Chromosome spreads were consecutively analyzed by solid stain, pancentromeric and pantelomeric FISH and mFISH. The results show that the frequency of incomplete chromosome elements, those lacking a telomeric signal at one end, decreases abruptly from G2 to M. This indicates that cells with incomplete chromosome elements can progress from G0 to G2, but at the G2/M checkpoint suffer a strong negative selection.

  5. Importance of immunopharmacogenomics in cancer treatment: Patient selection and monitoring for immune checkpoint antibodies.

    Science.gov (United States)

    Choudhury, Noura; Nakamura, Yusuke

    2016-02-01

    In the last 5 years, immune checkpoint antibodies have become established as anticancer agents for various types of cancer. These antibody drugs, namely cytotoxic T-lymphocyte-associated antigen, programmed death-1, and programmed death ligand-1 antibodies, have revealed relatively high response rates, the ability to induce durable responses, and clinical efficacy in malignancies not previously thought to be susceptible to immune-based strategies. However, because of its unique mechanisms of activating the host immune system against cancer as well as expensive cost, immune checkpoint blockade faces novel challenges in selecting appropriate patient populations, monitoring clinical responses, and predicting immune adverse events. The development of objective criteria for selecting patient populations that are likely to have benefit from these therapies has been vigorously investigated but still remains unclear. In this review, we describe immune checkpoint inhibition-specific challenges with patient selection and monitoring, and focus on approaches to remedy these challenges. We also discuss applications of the emerging field of immunopharmacogenomics for guiding selection and monitoring for anti-immune checkpoint treatment. PMID:26678880

  6. Preserved DNA Damage Checkpoint Pathway Protects against Complications in Long-Standing Type 1 Diabetes

    Energy Technology Data Exchange (ETDEWEB)

    Bhatt, Shweta; Gupta, Manoj; Khamaisi, Mogher; Martinez, Rachael; Gritsenko, Marina A.; Wagner, Bridget; Guye, Patrick; Busskamp, Volker; Shirakawa, Jun; Wu, Gongxiong; Liew, Chong Wee; Clauss, Therese RW; Valdez, Ivan; EL Ouaaman, Abdelfattah; Dirice, Ercument; Takatani, Tomozumi; Keenan, Hillary; Smith, Richard D.; Church, George; Weiss, Ron; Wagers, Amy J.; Qian, Weijun; King, George L.; Kulkami, Rohit N.

    2015-08-04

    Themechanisms underlying the development of complications in type 1 diabetes (T1D) are poorly understood. Disease modeling of induced pluripotent stem cells (iPSCs) from patients with longstanding T1D(disease durationR50 years) with severe (Medalist +C) or absent to mild complications (Medalist *C) revealed impaired growth, reprogramming, and differentiation in Medalist +C. Genomics and proteomics analyses suggested differential regulation of DNA damage checkpoint proteins favoring protection from cellular apoptosis in Medalist *C. In silico analyses showed altered expression patterns of DNA damage checkpoint factors among the Medalist groups to be targets of miR200, whose expression was significantly elevated in Medalist +C serum. Notably, neurons differentiated from Medalist +C iPSCs exhibited enhanced susceptibility to genotoxic stress that worsened upon miR200 overexpression. Furthermore, knockdown of miR200 in Medalist +C fibroblasts and iPSCs rescued checkpoint protein expression and reduced DNA damage.WeproposemiR200-regulated DNA damage checkpoint pathway as a potential therapeutic target for treating complications of diabetes.

  7. The effect of heavy ion on DNA damage checkpoint, and the discovery of its sensitizing compounds

    International Nuclear Information System (INIS)

    In the present study, we demonstrated that a persimmon leaf extract (PLE) promoted cytotoxic effect of cancer cells by chemotherapeutic agents inhibiting the DNA checkpoint activity. Therefore, PLE is a strong possibility of sensitizing agents for heavy ion cancer therapy. Herein, we investigate the mechanism of cellular responses to DNA damage induced by heavy ion radiation with PLE. Human adenocarcinoma A549 cells were pre-incubated with PLE (0, 1, 10, 30 μg/mL) at one hour. After pre-incubation, the cells were irradiated with carbon-ion beams [135C (linear energy transfer (LET) 70 Kev/um)]. The phosphorylation of p53, Chk1 and SMC1 was increased by 2.5 Gy heavy ion exposure. PLE decreased the phosphorylation of p53, Chk1 and SMC1 in cells damaged by heavy ion with PLE dose-dependent manner. G2/M checkpoint was investigated in percentage of mitotic cell. The percentage of mitotic cell decreased in heavy ion beam treatment. Interestingly, heavy ion with PLE treatment was significantly increased. The result indicated that PLE treatment disrupted the G2/M checkpoint activated by heavy ion. These results indicated that DNA damage checkpoint system in heavy ion exposed cells were abrogated by PLE treatment through the inhibition of ataxia telangiectasia mutated (ATM) and/or AT and Rad3 related (ATR)-dependent signaling pathway. (author)

  8. Chemogenetic profiling identifies RAD17 as synthetically lethal with checkpoint kinase inhibition.

    Science.gov (United States)

    Shen, John Paul; Srivas, Rohith; Gross, Andrew; Li, Jianfeng; Jaehnig, Eric J; Sun, Su Ming; Bojorquez-Gomez, Ana; Licon, Katherine; Sivaganesh, Vignesh; Xu, Jia L; Klepper, Kristin; Yeerna, Huwate; Pekin, Daniel; Qiu, Chu Ping; van Attikum, Haico; Sobol, Robert W; Ideker, Trey

    2015-11-01

    Chemical inhibitors of the checkpoint kinases have shown promise in the treatment of cancer, yet their clinical utility may be limited by a lack of molecular biomarkers to identify specific patients most likely to respond to therapy. To this end, we screened 112 known tumor suppressor genes for synthetic lethal interactions with inhibitors of the CHEK1 and CHEK2 checkpoint kinases. We identified eight interactions, including the Replication Factor C (RFC)-related protein RAD17. Clonogenic assays in RAD17 knockdown cell lines identified a substantial shift in sensitivity to checkpoint kinase inhibition (3.5-fold) as compared to RAD17 wild-type. Additional evidence for this interaction was found in a large-scale functional shRNA screen of over 100 genotyped cancer cell lines, in which CHEK1/2 mutant cell lines were unexpectedly sensitive to RAD17 knockdown. This interaction was widely conserved, as we found that RAD17 interacts strongly with checkpoint kinases in the budding yeast Saccharomyces cerevisiae. In the setting of RAD17 knockdown, CHEK1/2 inhibition was found to be synergistic with inhibition of WEE1, another pharmacologically relevant checkpoint kinase. Accumulation of the DNA damage marker γH2AX following chemical inhibition or transient knockdown of CHEK1, CHEK2 or WEE1 was magnified by knockdown of RAD17. Taken together, our data suggest that CHEK1 or WEE1 inhibitors are likely to have greater clinical efficacy in tumors with RAD17 loss-of-function. PMID:26437225

  9. The meiotic recombination checkpoint suppresses NHK-1 kinase to prevent reorganisation of the oocyte nucleus in Drosophila.

    Directory of Open Access Journals (Sweden)

    Oscar M Lancaster

    2010-10-01

    Full Text Available The meiotic recombination checkpoint is a signalling pathway that blocks meiotic progression when the repair of DNA breaks formed during recombination is delayed. In comparison to the signalling pathway itself, however, the molecular targets of the checkpoint that control meiotic progression are not well understood in metazoans. In Drosophila, activation of the meiotic checkpoint is known to prevent formation of the karyosome, a meiosis-specific organisation of chromosomes, but the molecular pathway by which this occurs remains to be identified. Here we show that the conserved kinase NHK-1 (Drosophila Vrk-1 is a crucial meiotic regulator controlled by the meiotic checkpoint. An nhk-1 mutation, whilst resulting in karyosome defects, does so independent of meiotic checkpoint activation. Rather, we find unrepaired DNA breaks formed during recombination suppress NHK-1 activity (inferred from the phosphorylation level of one of its substrates through the meiotic checkpoint. Additionally DNA breaks induced by X-rays in cultured cells also suppress NHK-1 kinase activity. Unrepaired DNA breaks in oocytes also delay other NHK-1 dependent nuclear events, such as synaptonemal complex disassembly and condensin loading onto chromosomes. Therefore we propose that NHK-1 is a crucial regulator of meiosis and that the meiotic checkpoint suppresses NHK-1 activity to prevent oocyte nuclear reorganisation until DNA breaks are repaired.

  10. DNA damage activates a spatially distinct late cytoplasmic cell-cycle checkpoint network controlled by MK2-mediated RNA stabilization

    DEFF Research Database (Denmark)

    Reinhardt, H Christian; Hasskamp, Pia; Schmedding, Ingolf;

    2010-01-01

    Following genotoxic stress, cells activate a complex kinase-based signaling network to arrest the cell cycle and initiate DNA repair. p53-defective tumor cells rewire their checkpoint response and become dependent on the p38/MK2 pathway for survival after DNA damage, despite a functional ATR-Chk1...... pathway. We used functional genetics to dissect the contributions of Chk1 and MK2 to checkpoint control. We show that nuclear Chk1 activity is essential to establish a G(2)/M checkpoint, while cytoplasmic MK2 activity is critical for prolonged checkpoint maintenance through a process of...... posttranscriptional mRNA stabilization. Following DNA damage, the p38/MK2 complex relocalizes from nucleus to cytoplasm where MK2 phosphorylates hnRNPA0, to stabilize Gadd45α mRNA, while p38 phosphorylates and releases the translational inhibitor TIAR. In addition, MK2 phosphorylates PARN, blocking Gadd45α m...

  11. Ghost Process: a Sound Basis to Implement Process Duplication, Migration and Checkpoint-Restart in Linux Clusters

    OpenAIRE

    Vallée, Geoffroy; Lottiaux, Renaud; Margery, David; Morin, Christine; Berthou, Jean-Yves

    2005-01-01

    Process management mechanisms (process duplication, migration and checkpoint-restart) are very useful for high performance and high availability in clustering systems. The single system image approach aims at providing a global process management service with mechanisms for process checkpoint, process migration and process duplication. In this context, a common mechanism for process virtualization is highly desirable but traditional operating systems do not provide such a mechanism. This pape...

  12. DNA damage-induced metaphase I arrest is mediated by the spindle assembly checkpoint and maternal age

    OpenAIRE

    Marangos, P; Stevense, M.; Niaka, K.; Lagoudaki, M.; Nabti, I.; Jessberger, R.; Carroll, J.

    2015-01-01

    In mammalian oocytes DNA damage can cause chromosomal abnormalities that potentially lead to infertility and developmental disorders. However, there is little known about the response of oocytes to DNA damage. Here we find that oocytes with DNA damage arrest at metaphase of the first meiosis (MI). The MI arrest is induced by the spindle assembly checkpoint (SAC) because inhibiting the SAC overrides the DNA damage-induced MI arrest. Furthermore, this MI checkpoint is compromised in oocytes fro...

  13. Telomerase expression is sufficient for chromosomal integrity in cells lacking p53 dependent G1 checkpoint function

    Directory of Open Access Journals (Sweden)

    Simpson Dennis

    2005-01-01

    Full Text Available Abstract Background Secondary cultures of human fibroblasts display a finite lifespan ending at senescence. Loss of p53 function by mutation or viral oncogene expression bypasses senescence, allowing cell division to continue for an additional 10 – 20 doublings. During this time chromosomal aberrations seen in mitotic cells increase while DNA damage and decatenation checkpoint functions in G2 cells decrease. Methods To explore this complex interplay between chromosomal instability and checkpoint dysfunction, human fibroblast lines were derived that expressed HPV16E6 oncoprotein or dominant-negative alleles of p53 (A143V and H179Q with or without the catalytic subunit of telomerase. Results Cells with normal p53 function displayed 86 – 93% G1 arrest after exposure to 1.5 Gy ionizing radiation (IR. Expression of HPV16E6 or p53-H179Q severely attenuated G1 checkpoint function (3 – 20% arrest while p53-A143V expression induced intermediate attenuation (55 – 57% arrest irrespective of telomerase expression. All cell lines, regardless of telomerase expression or p53 status, exhibited a normal DNA damage G2 checkpoint response following exposure to 1.5 Gy IR prior to the senescence checkpoint. As telomerase-negative cells bypassed senescence, the frequencies of chromosomal aberrations increased generally congruent with attenuation of G2 checkpoint function. Telomerase expression allowed cells with defective p53 function to grow >175 doublings without chromosomal aberrations or attenuation of G2 checkpoint function. Conclusion Thus, chromosomal instability in cells with defective p53 function appears to depend upon telomere erosion not loss of the DNA damage induced G1 checkpoint.

  14. RNA interference regulates the cell cycle checkpoint through the RNA export factor, Ptr1, in fission yeast

    Energy Technology Data Exchange (ETDEWEB)

    Iida, Tetsushi, E-mail: tiida@nig.ac.jp [Division of Cytogenetics, National Institute of Genetics, Mishima, 1111 Yata, Mishima 411-8540 (Japan); The Graduate University for Advanced Studies, Sokendai, Mishima, 1111 Yata, Mishima 411-8540 (Japan); Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency (JST), 4-1-8, Honcho, Kawaguchi-shi, Saitama 332-0012 (Japan); Iida, Naoko [Division of Mutagenesis, National Institute of Genetics, Mishima, 1111 Yata, Mishima 411-8540 (Japan); Tsutsui, Yasuhiro [Department of Life Science, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Nagatsuda-cho, Midori-ku, Yokohama 226-8501 (Japan); Yamao, Fumiaki [Division of Mutagenesis, National Institute of Genetics, Mishima, 1111 Yata, Mishima 411-8540 (Japan); The Graduate University for Advanced Studies, Sokendai, Mishima, 1111 Yata, Mishima 411-8540 (Japan); Kobayashi, Takehiko [Division of Cytogenetics, National Institute of Genetics, Mishima, 1111 Yata, Mishima 411-8540 (Japan); The Graduate University for Advanced Studies, Sokendai, Mishima, 1111 Yata, Mishima 411-8540 (Japan)

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer RNAi is linked to the cell cycle checkpoint in fission yeast. Black-Right-Pointing-Pointer Ptr1 co-purifies with Ago1. Black-Right-Pointing-Pointer The ptr1-1 mutation impairs the checkpoint but does not affect gene silencing. Black-Right-Pointing-Pointer ago1{sup +} and ptr1{sup +} regulate the cell cycle checkpoint via the same pathway. Black-Right-Pointing-Pointer Mutations in ago1{sup +} and ptr1{sup +} lead to the nuclear accumulation of poly(A){sup +} RNAs. -- Abstract: Ago1, an effector protein of RNA interference (RNAi), regulates heterochromatin silencing and cell cycle arrest in fission yeast. However, the mechanism by which Ago1 controls cell cycle checkpoint following hydroxyurea (HU) treatment has not been elucidated. In this study, we show that Ago1 and other RNAi factors control cell cycle checkpoint following HU treatment via a mechanism independent of silencing. While silencing requires dcr1{sup +}, the overexpression of ago1{sup +} alleviated the cell cycle defect in dcr1{Delta}. Ago1 interacted with the mRNA export factor, Ptr1. The ptr1-1 mutation impaired cell cycle checkpoint but gene silencing was unaffected. Genetic analysis revealed that the regulation of cell cycle checkpoint by ago1{sup +} is dependent on ptr1{sup +}. Nuclear accumulation of poly(A){sup +} RNAs was detected in mutants of ago1{sup +} and ptr1{sup +}, suggesting there is a functional link between the cell cycle checkpoint and RNAi-mediated RNA quality control.

  15. Intestinal microbiome analyses identify melanoma patients at risk for checkpoint-blockade-induced colitis.

    Science.gov (United States)

    Dubin, Krista; Callahan, Margaret K; Ren, Boyu; Khanin, Raya; Viale, Agnes; Ling, Lilan; No, Daniel; Gobourne, Asia; Littmann, Eric; Huttenhower, Curtis; Pamer, Eric G; Wolchok, Jedd D

    2016-01-01

    The composition of the intestinal microbiota influences the development of inflammatory disorders. However, associating inflammatory diseases with specific microbial members of the microbiota is challenging, because clinically detectable inflammation and its treatment can alter the microbiota's composition. Immunologic checkpoint blockade with ipilimumab, a monoclonal antibody that blocks cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) signalling, is associated with new-onset, immune-mediated colitis. Here we conduct a prospective study of patients with metastatic melanoma undergoing ipilimumab treatment and correlate the pre-inflammation faecal microbiota and microbiome composition with subsequent colitis development. We demonstrate that increased representation of bacteria belonging to the Bacteroidetes phylum is correlated with resistance to the development of checkpoint-blockade-induced colitis. Furthermore, a paucity of genetic pathways involved in polyamine transport and B vitamin biosynthesis is associated with an increased risk of colitis. Identification of these biomarkers may enable interventions to reduce the risk of inflammatory complications following cancer immunotherapy. PMID:26837003

  16. Social and ethical checkpoints for bottom-up synthetic biology, or protocells.

    Science.gov (United States)

    Bedau, Mark A; Parke, Emily C; Tangen, Uwe; Hantsche-Tangen, Brigitte

    2009-12-01

    An alternative to creating novel organisms through the traditional "top-down" approach to synthetic biology involves creating them from the "bottom up" by assembling them from non-living components; the products of this approach are called "protocells." In this paper we describe how bottom-up and top-down synthetic biology differ, review the current state of protocell research and development, and examine the unique ethical, social, and regulatory issues raised by bottom-up synthetic biology. Protocells have not yet been developed, but many expect this to happen within the next five to ten years. Accordingly, we identify six key checkpoints in protocell development at which particular attention should be given to specific ethical, social and regulatory issues concerning bottom-up synthetic biology, and make ten recommendations for responsible protocell science that are tied to the achievement of these checkpoints. PMID:19816801

  17. An Overview of the Spindle Assembly Checkpoint Status in Oral Cancer

    Science.gov (United States)

    Teixeira, José Henrique; Silva, Patrícia Manuela; Reis, Rita Margarida; Moura, Inês Moranguinho; Marques, Sandra; Fonseca, Joana; Monteiro, Luís Silva; Bousbaa, Hassan

    2014-01-01

    Abnormal chromosome number, or aneuploidy, is a common feature of human solid tumors, including oral cancer. Deregulated spindle assembly checkpoint (SAC) is thought as one of the mechanisms that drive aneuploidy. In normal cells, SAC prevents anaphase onset until all chromosomes are correctly aligned at the metaphase plate thereby ensuring genomic stability. Significantly, the activity of this checkpoint is compromised in many cancers. While mutations are rather rare, many tumors show altered expression levels of SAC components. Genomic alterations such as aneuploidy indicate a high risk of oral cancer and cancer-related mortality, and the molecular basis of these alterations is largely unknown. Yet, our knowledge on the status of SAC components in oral cancer remains sparse. In this review, we address the state of our knowledge regarding the SAC defects and the underlying molecular mechanisms in oral cancer, and discuss their therapeutic relevance, focusing our analysis on the core components of SAC and its target Cdc20. PMID:24995269

  18. Cloud object store for checkpoints of high performance computing applications using decoupling middleware

    Energy Technology Data Exchange (ETDEWEB)

    Bent, John M.; Faibish, Sorin; Grider, Gary

    2016-04-19

    Cloud object storage is enabled for checkpoints of high performance computing applications using a middleware process. A plurality of files, such as checkpoint files, generated by a plurality of processes in a parallel computing system are stored by obtaining said plurality of files from said parallel computing system; converting said plurality of files to objects using a log structured file system middleware process; and providing said objects for storage in a cloud object storage system. The plurality of processes may run, for example, on a plurality of compute nodes. The log structured file system middleware process may be embodied, for example, as a Parallel Log-Structured File System (PLFS). The log structured file system middleware process optionally executes on a burst buffer node.

  19. Dynein Light Intermediate Chain 2 Facilitates the Metaphase to Anaphase Transition by Inactivating the Spindle Assembly Checkpoint.

    Directory of Open Access Journals (Sweden)

    Sagar P Mahale

    Full Text Available The multi-functional molecular motor cytoplasmic dynein performs diverse essential roles during mitosis. The mechanistic importance of the dynein Light Intermediate Chain homologs, LIC1 and LIC2 is unappreciated, especially in the context of mitosis. LIC1 and LIC2 are believed to exist in distinct cytoplasmic dynein complexes as obligate subunits. LIC1 had earlier been reported to be required for metaphase to anaphase progression by inactivating the kinetochore-microtubule attachment-sensing arm of the spindle assembly checkpoint (SAC. However, the functional importance of LIC2 during mitosis remains elusive. Here we report prominent novel roles for the LIC2 subunit of cytoplasmic dynein in regulating the spindle assembly checkpoint. LIC2 depletion in mammalian cells led to prolonged metaphase arrest in the presence of an active SAC and also to stretched kinetochores, thus implicating it in SAC inactivation. Quantitative fluorescence microscopy of SAC components revealed accumulation of both attachment- and tension-sensing checkpoint proteins at metaphase kinetochores upon LIC2 depletion. These observations support a stronger and more diverse role in checkpoint inactivation for LIC2 in comparison to its close homolog LIC1. Our study uncovers a novel functional hierarchy during mitotic checkpoint inactivation between the closely related but homologous LIC subunits of cytoplasmic dynein. These subtle functional distinctions between dynein subpopulations could be exploited to study specific aspects of the spindle assembly checkpoint, which is a key mediator of fidelity in eukaryotic cell division.

  20. Lyn tyrosine kinase promotes silencing of ATM-dependent checkpoint signaling during recovery from DNA double-strand breaks

    International Nuclear Information System (INIS)

    Highlights: • Inhibition of Src family kinases decreased γ-H2AX signal. • Inhibition of Src family increased ATM-dependent phosphorylation of Chk2 and Kap1. • shRNA-mediated knockdown of Lyn increased phosphorylation of Kap1 by ATM. • Ectopic expression of Src family kinase suppressed ATM-mediated Kap1 phosphorylation. • Src is involved in upstream signaling for inactivation of ATM signaling. - Abstract: DNA damage activates the DNA damage checkpoint and the DNA repair machinery. After initial activation of DNA damage responses, cells recover to their original states through completion of DNA repair and termination of checkpoint signaling. Currently, little is known about the process by which cells recover from the DNA damage checkpoint, a process called checkpoint recovery. Here, we show that Src family kinases promote inactivation of ataxia telangiectasia mutated (ATM)-dependent checkpoint signaling during recovery from DNA double-strand breaks. Inhibition of Src activity increased ATM-dependent phosphorylation of Chk2 and Kap1. Src inhibition increased ATM signaling both in G2 phase and during asynchronous growth. shRNA knockdown of Lyn increased ATM signaling. Src-dependent nuclear tyrosine phosphorylation suppressed ATM-mediated Kap1 phosphorylation. These results suggest that Src family kinases are involved in upstream signaling that leads to inactivation of the ATM-dependent DNA damage checkpoint

  1. Differential activation of intra-S-phase checkpoint in response to tripchlorolide and its effects on DNA replication

    Institute of Scientific and Technical Information of China (English)

    Yan REN; Jia Rui WU

    2004-01-01

    DNA replication is tightly regulated during the S phase of the cell cycle, and the activation of the intra-S-phase checkpoint due to DNA damage usually results in arrest of DNA synthesis. However, the molecular details about the correlation between the checkpoint and regulation of DNA replication are still unclear. To investigate the connections between DNA replication and DNA damage checkpoint, a DNA-damage reagent, tripchlorolide, was applied to CHO (Chinese ovary hamster) cells at early- or middle-stages of the S phase. The early-S-phase treatment with TC significantly delayed the progression of the S phase and caused the phosphorylation of the Chk1 checkpoint protein, whereas the middle-S-phase treatment only slightly slowed down the progression of the S phase. Furthermore, the analysis of DNA replication patterns revealed that replication pattern Ⅱ was greatly prolonged in the cells treated with the drug during the early-S phase, whereas the late-replication patterns of these cells were hardly detected, suggesting that the activation of the intra-S-phase checkpoint inhibits the late-origin firing of DNA replication. We conclude that cells at different stages of the S phase are differentially sensitive to the DNA-damage reagent, and the activation of the intra-Sphase checkpoint blocks the DNA replication progression in the late stage of S phase.

  2. Dynein Light Intermediate Chain 2 Facilitates the Metaphase to Anaphase Transition by Inactivating the Spindle Assembly Checkpoint

    Science.gov (United States)

    Mahale, Sagar P.; Sharma, Amit; Mylavarapu, Sivaram V. S.

    2016-01-01

    The multi-functional molecular motor cytoplasmic dynein performs diverse essential roles during mitosis. The mechanistic importance of the dynein Light Intermediate Chain homologs, LIC1 and LIC2 is unappreciated, especially in the context of mitosis. LIC1 and LIC2 are believed to exist in distinct cytoplasmic dynein complexes as obligate subunits. LIC1 had earlier been reported to be required for metaphase to anaphase progression by inactivating the kinetochore-microtubule attachment-sensing arm of the spindle assembly checkpoint (SAC). However, the functional importance of LIC2 during mitosis remains elusive. Here we report prominent novel roles for the LIC2 subunit of cytoplasmic dynein in regulating the spindle assembly checkpoint. LIC2 depletion in mammalian cells led to prolonged metaphase arrest in the presence of an active SAC and also to stretched kinetochores, thus implicating it in SAC inactivation. Quantitative fluorescence microscopy of SAC components revealed accumulation of both attachment- and tension-sensing checkpoint proteins at metaphase kinetochores upon LIC2 depletion. These observations support a stronger and more diverse role in checkpoint inactivation for LIC2 in comparison to its close homolog LIC1. Our study uncovers a novel functional hierarchy during mitotic checkpoint inactivation between the closely related but homologous LIC subunits of cytoplasmic dynein. These subtle functional distinctions between dynein subpopulations could be exploited to study specific aspects of the spindle assembly checkpoint, which is a key mediator of fidelity in eukaryotic cell division. PMID:27441562

  3. Lyn tyrosine kinase promotes silencing of ATM-dependent checkpoint signaling during recovery from DNA double-strand breaks

    Energy Technology Data Exchange (ETDEWEB)

    Fukumoto, Yasunori, E-mail: fukumoto@faculty.chiba-u.jp; Kuki, Kazumasa; Morii, Mariko; Miura, Takahito; Honda, Takuya; Ishibashi, Kenichi; Hasegawa, Hitomi; Kubota, Sho; Ide, Yudai; Yamaguchi, Noritaka; Nakayama, Yuji; Yamaguchi, Naoto, E-mail: nyama@faculty.chiba-u.jp

    2014-09-26

    Highlights: • Inhibition of Src family kinases decreased γ-H2AX signal. • Inhibition of Src family increased ATM-dependent phosphorylation of Chk2 and Kap1. • shRNA-mediated knockdown of Lyn increased phosphorylation of Kap1 by ATM. • Ectopic expression of Src family kinase suppressed ATM-mediated Kap1 phosphorylation. • Src is involved in upstream signaling for inactivation of ATM signaling. - Abstract: DNA damage activates the DNA damage checkpoint and the DNA repair machinery. After initial activation of DNA damage responses, cells recover to their original states through completion of DNA repair and termination of checkpoint signaling. Currently, little is known about the process by which cells recover from the DNA damage checkpoint, a process called checkpoint recovery. Here, we show that Src family kinases promote inactivation of ataxia telangiectasia mutated (ATM)-dependent checkpoint signaling during recovery from DNA double-strand breaks. Inhibition of Src activity increased ATM-dependent phosphorylation of Chk2 and Kap1. Src inhibition increased ATM signaling both in G2 phase and during asynchronous growth. shRNA knockdown of Lyn increased ATM signaling. Src-dependent nuclear tyrosine phosphorylation suppressed ATM-mediated Kap1 phosphorylation. These results suggest that Src family kinases are involved in upstream signaling that leads to inactivation of the ATM-dependent DNA damage checkpoint.

  4. Intestinal microbiome analyses identify melanoma patients at risk for checkpoint-blockade-induced colitis

    OpenAIRE

    Dubin, Krista; Callahan, Margaret K.; Ren, Boyu; Khanin, Raya; Viale, Agnes; Ling, Lilan; No, Daniel; Gobourne, Asia; Littmann, Eric; Huttenhower, Curtis; Pamer, Eric G.; Wolchok, Jedd D.

    2016-01-01

    The composition of the intestinal microbiota influences the development of inflammatory disorders. However, associating inflammatory diseases with specific microbial members of the microbiota is challenging, because clinically detectable inflammation and its treatment can alter the microbiota's composition. Immunologic checkpoint blockade with ipilimumab, a monoclonal antibody that blocks cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) signalling, is associated with new-onset, immune-med...

  5. CDC28, NETI, and HFII are required for checkpoints in Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    The involvement of SRM genes selected as genes affecting genetic stability and radiosensitivity in a cell cycle arrest under the action of damaging agents was studied. It was shown that the srm5/cdc28-srm, srm8/netI-srm, and srmI2/hfiI-srm mutations prevent checkpoint activation by DNA damage, particularly the G0/S (srm5, srm8), G1/S (srm5, srm8, srm12), S (srm8, srm12) and S/G2 (srm5) checkpoints. It seems that in budding yeast the CDC28, HFII/ADAI, and NETI genes mediate cellular response induced by DNA damage with checkpoint control. The well-known checkpoint-genes RAD9, RAD17, RAD24, and RAD53, and the genes CDC28, and NETI have been found to belong to one epistasis group named RAD9-group as regards cell sensitivity to γ radiation. An analysis of the radiosensitivity of double mutants has revealed that the mutation cdc-28-srm is hypostatic to each of mutations rad9Δ, and rad24Δ, and additive to rad17Δ. The mutation netI-srm is hypostatic to the mutations rad9Δ but additive to rad17Δ, rad24Δ, and rad53. The mutation hfiI-srm has an additive effect in compound with the mutations rad24Δ and rad9Δ. So, investigations of epistatic interactions have demonstrated a branched RAD9-dependent pathway. The analyzed genes can also participate in a minor mechanism involved in determining cell radiation sensitivity independently of the mentioned RAD9-dependent pathway

  6. Handoff Based Secure Checkpointing and Log Based Rollback Recovery for Mobile Hosts

    Directory of Open Access Journals (Sweden)

    Priyanka Dey and Suparna Biswas

    2012-10-01

    Full Text Available An efficient fault tolerant algorithm based on movement-based secure checkpointing and logging formobile computing system is proposed here. The recovery scheme proposed here combines independentcheckpointing and message logging. Here we consider mobility rate of the user in checkpointing so thatmobile host can manage recovery information such as checkpoints and logs properly so that a mobilehost takes less recovery time after failure. Mobile hosts save checkpoints when number of hand-offexceeds a predefined hand-off threshold value. Current approaches save logs in base station. But thisapproach maximizes recovery time if message passing frequency is large. If a mobile host saves log in itsown memory, recovery cost will be less because log retrieval time will be small after failure. But there isa probability of memory crash of a mobile host. In that case logs can not be retrieved if it is saved only inmobile node. If the failure is transient then logs can be retrieved from the memory of mobile node.Hence in this algorithm mobile hosts also save log in own memory and base station. In case of crashrecovery, log will be retrieved from base station and in case of transient failure recovery logs will beretrieved from mobile host. In this algorithm recovery probability is optimized and total recovery time isreduced in comparison to existing works. Logs are very small in size. Hence saving logs in mobile hostsdoes not cause much memory overhead. Hand-off threshold is a function of mobility rate, messagepassing frequency and failure rate of mobile hosts. This algorithm describes a secure check pointingtechnique as a method for providing fault tolerance while preventing information leakage through thecheckpoint data.

  7. Sudden Telomere Lengthening Triggers a Rad53-dependent Checkpoint in Saccharomyces cerevisiae

    OpenAIRE

    Viscardi, Valeria; Baroni, Enrico; Romano, Michele; Lucchini, Giovanna; Longhese, Maria Pia

    2003-01-01

    Telomeres are specialized functional complexes that ensure chromosome stability by protecting chromosome ends from fusions and degradation and avoiding chromosomal termini from being sensed as DNA breaks. Budding yeast Tel1 is required both for telomere metabolism and for a Rad53-dependent checkpoint responding to unprocessed double-strand breaks. We show that overexpression of a GAL1-TEL1 fusion causes transient telomere lengthening and activation of a Rad53-dependent...

  8. In vivo role of checkpoint kinase 2 in signaling telomere dysfunction

    OpenAIRE

    García-Beccaria, María; Martínez, Paula; Flores, Juana M.; Blasco, Maria A.

    2014-01-01

    Checkpoint kinase 2 (CHK2) is a downstream effector of the DNA damage response (DDR). Dysfunctional telomeres, either owing to critical shortening or disruption of the shelterin complex, activate a DDR, which eventually results in cell cycle arrest, senescence and/or apoptosis. Successive generations of telomerase-deficient (Terc) mice show accelerated aging and shorter lifespan due to tissue atrophy and impaired organ regeneration associated to progressive telomere shortening. In contrast, m...

  9. Cdk2 is required for p53-independent G2/M checkpoint control.

    Directory of Open Access Journals (Sweden)

    Jon H Chung

    2010-02-01

    Full Text Available The activation of phase-specific cyclin-dependent kinases (Cdks is associated with ordered cell cycle transitions. Among the mammalian Cdks, only Cdk1 is essential for somatic cell proliferation. Cdk1 can apparently substitute for Cdk2, Cdk4, and Cdk6, which are individually dispensable in mice. It is unclear if all functions of non-essential Cdks are fully redundant with Cdk1. Using a genetic approach, we show that Cdk2, the S-phase Cdk, uniquely controls the G(2/M checkpoint that prevents cells with damaged DNA from initiating mitosis. CDK2-nullizygous human cells exposed to ionizing radiation failed to exclude Cdk1 from the nucleus and exhibited a marked defect in G(2/M arrest that was unmasked by the disruption of P53. The DNA replication licensing protein Cdc6, which is normally stabilized by Cdk2, was physically associated with the checkpoint regulator ATR and was required for efficient ATR-Chk1-Cdc25A signaling. These findings demonstrate that Cdk2 maintains a balance of S-phase regulatory proteins and thereby coordinates subsequent p53-independent G(2/M checkpoint activation.

  10. Biological significance of the focus on DNA damage checkpoint factors remained after irradiation of ionizing radiation

    International Nuclear Information System (INIS)

    This paper reviews recent reports on the focus formation and participation to checkpoint of (such phosphorylated (P-d) as below) ATM and H2AX, MDC1, 53BP1 and NBS1, and discusses their role in DNA damage checkpoint induction mainly around authors' studies. When the cell is irradiated by ionizing radiation, the subtype histone like H2AX is P-d and the formed focus', seen in the nucleus on immuno-fluorographic observation, represents the P-d H2AX at the damaged site of DNA. The role of P-d ATM (the product of causative gene of ataxia-telangiectasia mutation, a protein kinase) has been first shown by laser beam irradiation. Described are discussions on the roles and functions after irradiation in focus formation and DNA damage checkpoint of P-d H2AX (a specific histone product by the radiation like γ-ray as above), P-d ATM, MDC1 (a mediator of DNA damage check point protein 1), 53BP1, (a p53 binding protein) and NBS1 (the product of the causative gene of Nijmegen Breakage Syndrome). Authors have come to point out the remained focal size increase as implications of the efficient repair of damaged DNA, and the second cycled p53 accumulation, of tumor suppression. Thus evaluation of biological significance of these aspects, scarcely noted hitherto, is concluded important. (S.I.)

  11. Efficient checkpointing schemes for depletion perturbation solutions on memory-limited architectures

    International Nuclear Information System (INIS)

    We describe a methodology for decreasing the memory footprint and machine I/O load associated with the need to access a forward solution during an adjoint solve. Specifically, we are interested in the depletion perturbation equations, where terms in the adjoint Bateman and transport equations depend on the forward flux solution. Checkpointing is the procedure of storing snapshots of the forward solution to disk and using these snapshots to recompute the parts of the forward solution that are necessary for the adjoint solve. For large problems, however, the storage cost of just a few copies of an angular flux vector can exceed the available RAM on the host machine. We propose a methodology that does not checkpoint the angular flux vector; instead, we write and store converged source moments, which are typically of a much lower dimension than the angular flux solution. This reduces the memory footprint and I/O load of the problem, but requires that we perform single sweeps to reconstruct flux vectors on demand. We argue that this trade-off is exactly the kind of algorithm that will scale on advanced, memory-limited architectures. We analyze the cost, in terms of FLOPS and memory footprint, of five checkpointing schemes. We also provide computational results that support the analysis and show that the memory-for-work trade off does improve time to solution. (authors)

  12. Efficient checkpointing schemes for depletion perturbation solutions on memory-limited architectures

    Energy Technology Data Exchange (ETDEWEB)

    Stripling, H. F.; Adams, M. L.; Hawkins, W. D. [Texas A and M University, Department of Nuclear Engineering, 3133 TAMU, College Station, TX 77843-3133 (United States)

    2013-07-01

    We describe a methodology for decreasing the memory footprint and machine I/O load associated with the need to access a forward solution during an adjoint solve. Specifically, we are interested in the depletion perturbation equations, where terms in the adjoint Bateman and transport equations depend on the forward flux solution. Checkpointing is the procedure of storing snapshots of the forward solution to disk and using these snapshots to recompute the parts of the forward solution that are necessary for the adjoint solve. For large problems, however, the storage cost of just a few copies of an angular flux vector can exceed the available RAM on the host machine. We propose a methodology that does not checkpoint the angular flux vector; instead, we write and store converged source moments, which are typically of a much lower dimension than the angular flux solution. This reduces the memory footprint and I/O load of the problem, but requires that we perform single sweeps to reconstruct flux vectors on demand. We argue that this trade-off is exactly the kind of algorithm that will scale on advanced, memory-limited architectures. We analyze the cost, in terms of FLOPS and memory footprint, of five checkpointing schemes. We also provide computational results that support the analysis and show that the memory-for-work trade off does improve time to solution. (authors)

  13. Immune checkpoint blockade therapy: The 2014 Tang prize in biopharmaceutical science

    Directory of Open Access Journals (Sweden)

    Ya-Shan Chen

    2015-02-01

    Full Text Available The first Tang Prize for Biopharmaceutical Science has been awarded to Prof. James P. Allison and Prof. Tasuku Honjo for their contributions leading to an entirely new way to treat cancer by blocking the molecules cytotoxic T lymphocyte-associated antigen 4 (CTLA-4 and programmed cell death protein 1 (PD-1 that turn off immune response. The treatment, called "immune checkpoint blockade therapy," has opened a new therapeutic era. Here the discoveries of the immune checkpoints and how they contribute to the maintenance of self-tolerance, as well as how to protect tissues from the excess immune responses causing damage are reviewed. The efforts made by Prof. Allison and Prof. Honjo for developing the most promising approaches to activate therapeutic antitumor immunity are also summarized. Since these certain immune checkpoint pathways appear to be one of the major mechanisms resulting in immune escape of tumors, the presence of anti-CTLA-4 and/or anti-PD-1 should contribute to removal of the inhibition signals for T cell activation. Subsequently, it will enhance specific T cell activation and, therefore, strengthen antitumor immunity.

  14. Immunologic checkpoints in cancer therapy: focus on the programmed death-1 (PD-1 receptor pathway

    Directory of Open Access Journals (Sweden)

    Momtaz P

    2014-11-01

    Full Text Available Parisa Momtaz,1,2 Michael A Postow1,2 1Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, NY, USA; 2Weill Cornell Medical College, New York, NY, USA Abstract: T-lymphocytes have the potential to recognize cancer antigens as foreign and therefore eliminate them. However, immune checkpoints such as cytotoxic T-lymphocyte-associated antigen (CTLA-4 and programmed cell death (PD-1 receptor and its ligands (PD-L1, PD-L2 suppress the activity of T-lymphocytes. Advances in the understanding of immunology and its role in cancer have led to the development of immune checkpoint inhibitors that block CTLA-4 and PD-1 and result in durable responses in patients with a wide range of cancers. PD-1 and PD-L1 inhibitors are currently in many stages of clinical investigation, and the anti-PD-1 antibody, pembrolizumab, was recently approved by the US Food and Drug Administration. Many questions remain to be answered, such as the optimal administration schedule, biomarkers that associate with benefit, and potential for use of PD-1 agents in combination approaches. Nonetheless, immunotherapy with PD-1 blocking antibodies is now becoming an integral part in the management of cancer. Keyword: immune checkpoints, immunotherapy, programmed cell death protein-1, cytotoxic T-lymphocyte antigen 4

  15. Eavesdropping on the cytoskeleton: progress and controversy in the yeast morphogenesis checkpoint.

    Science.gov (United States)

    Keaton, Mignon A; Lew, Daniel J

    2006-12-01

    The morphogenesis checkpoint provides a link between bud formation and mitosis in yeast. In this pathway, insults affecting the actin or septin cytoskeleton trigger a cell cycle arrest, mediated by the Wee1 homolog Swe1p, which catalyzes the inhibitory phosphorylation of the mitosis-promoting cyclin-dependent kinase (CDK) on a conserved tyrosine residue. Analyses of Swe1p phosphorylation have mapped 61 sites targeted by CDKs and Polo-related kinases, which control both Swe1p activity and Swe1p degradation. Although the sites themselves are not evolutionarily conserved, the control of Swe1p degradation exhibits many conserved features, and is linked to DNA-responsive checkpoints in vertebrate cells. At the 'sensing' end of the checkpoint, recent work has begun to shed light on how septins are organized and how they impact Swe1p regulators. However, the means by which Swe1p responds to actin perturbations once a bud has formed remains controversial. PMID:17055334

  16. Multicellular tumor spheroid models to explore cell cycle checkpoints in 3D

    International Nuclear Information System (INIS)

    MultiCellular Tumor Spheroid (MCTS) mimics the organization of a tumor and is considered as an invaluable model to study cancer cell biology and to evaluate new antiproliferative drugs. Here we report how the characteristics of MCTS in association with new technological developments can be used to explore the regionalization and the activation of cell cycle checkpoints in 3D. Cell cycle and proliferation parameters were investigated in Capan-2 spheroids by immunofluorescence staining, EdU incorporation and using cells engineered to express Fucci-red and -green reporters. We describe in details the changes in proliferation and cell cycle parameters during spheroid growth and regionalization. We report the kinetics and regionalized aspects of cell cycle arrest in response to checkpoint activation induced by EGF starvation, lovastatin treatment and etoposide-induced DNA damage. Our data present the power and the limitation of spheroids made of genetically modified cells to explore cell cycle checkpoints. This study paves the way for the investigation of molecular aspects and dynamic studies of the response to novel antiproliferative agents in 3D models

  17. RNF8 Transduces the DNA-Damage Signal Via Histone Ubiquitylation And Checkpoint Protein Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Huen, M.S.Y.; Grant, R.; Manke, I.; Minn, K.; Yu, X.; Yaffe, M.B.; Chen, J.

    2009-06-01

    DNA-damage signaling utilizes a multitude of posttranslational modifiers as molecular switches to regulate cell-cycle checkpoints, DNA repair, cellular senescence, and apoptosis. Here we show that RNF8, a FHA/RING domain-containing protein, plays a critical role in the early DNA-damage response. We have solved the X-ray crystal structure of the FHA domain structure at 1.35 {angstrom}. We have shown that RNF8 facilitates the accumulation of checkpoint mediator proteins BRCA1 and 53BP1 to the damaged chromatin, on one hand through the phospho-dependent FHA domain-mediated binding of RNF8 to MDC1, on the other hand via its role in ubiquitylating H2AX and possibly other substrates at damage sites. Moreover, RNF8-depleted cells displayed a defective G2/M checkpoint and increased IR sensitivity. Together, our study implicates RNF8 as a novel DNA-damage-responsive protein that integrates protein phosphorylation and ubiquitylation signaling and plays a critical role in the cellular response to genotoxic stress.

  18. The Rho-GAP Bem2p plays a GAP-independent role in the morphogenesis checkpoint

    Science.gov (United States)

    Marquitz, Aron R.; Harrison, Jacob C.; Bose, Indrani; Zyla, Trevin R.; McMillan, John N.; Lew, Daniel J.

    2002-01-01

    The Saccharomyces cerevisiae morphogenesis checkpoint delays mitosis in response to insults that impair actin organization and/or bud formation. The delay is due to accumulation of the inhibitory kinase Swe1p, which phosphorylates the cyclin-dependent kinase Cdc28p. Having screened through a panel of yeast mutants with defects in cell morphogenesis, we report here that the polarity establishment protein Bem2p is required for the checkpoint response. Bem2p is a Rho-GTPase activating protein (GAP) previously shown to act on Rho1p, and we now show that it also acts on Cdc42p, the GTPase primarily responsible for establishment of cell polarity in yeast. Whereas the morphogenesis role of Bem2p required GAP activity, the checkpoint role of Bem2p did not. Instead, this function required an N-terminal Bem2p domain. Thus, this single protein has a GAP-dependent role in promoting cell polarity and a GAP-independent role in responding to defects in cell polarity by enacting the checkpoint. Surprisingly, Swe1p accumulation occurred normally in bem2 cells, but they were nevertheless unable to promote Cdc28p phosphorylation. Therefore, Bem2p defines a novel pathway in the morphogenesis checkpoint. PMID:12145202

  19. Dpb11/TopBP1 plays distinct roles in DNA replication, checkpoint response and homologous recombination

    DEFF Research Database (Denmark)

    Germann, Susanne Manuela; Østergaard, Vibe Hallundbæk; Haas, Caroline;

    2011-01-01

    DPB11/TopBP1 is an essential evolutionarily conserved gene involved in initiation of DNA replication and checkpoint signaling. Here, we show that Saccharomyces cerevisiae Dpb11 forms nuclear foci that localize to sites of DNA damage in G1, S and G2 phase, a recruitment that is conserved for its...... and Tel1, and of the checkpoint mediator Rad9. In a site-directed mutagenesis screen, we identify a separation-of-function mutant, dpb11-PF, that is sensitive to DSB-inducing agents yet remains proficient for DNA replication and the S-phase checkpoint at the permissive temperature. The dpb11-PF mutant...... homologue TopBP1 in Gallus gallus. Damage-induced Dpb11 foci are distinct from Sld3 replication initiation foci. Further, Dpb11 foci are dependent on the checkpoint proteins Mec3 (9-1-1 complex) and Rad24, and require the C-terminal domain of Dpb11. Dpb11 foci are independent of the checkpoint kinases Mec1...

  20. Drosophila MOF controls Checkpoint protein2 and regulates genomic stability during early embryogenesis

    Directory of Open Access Journals (Sweden)

    Pushpavalli Sreerangam NCVL

    2013-01-01

    Full Text Available Abstract Background In Drosophila embryos, checkpoints maintain genome stability by delaying cell cycle progression that allows time for damage repair or to complete DNA synthesis. Drosophila MOF, a member of MYST histone acetyl transferase is an essential component of male X hyperactivation process. Until recently its involvement in G2/M cell cycle arrest and defects in ionizing radiation induced DNA damage pathways was not well established. Results Drosophila MOF is highly expressed during early embryogenesis. In the present study we show that haplo-insufficiency of maternal MOF leads to spontaneous mitotic defects like mitotic asynchrony, mitotic catastrophe and chromatid bridges in the syncytial embryos. Such abnormal nuclei are eliminated and digested in the yolk tissues by nuclear fall out mechanism. MOF negatively regulates Drosophila checkpoint kinase 2 tumor suppressor homologue. In response to DNA damage the checkpoint gene Chk2 (Drosophila mnk is activated in the mof mutants, there by causing centrosomal inactivation suggesting its role in response to genotoxic stress. A drastic decrease in the fall out nuclei in the syncytial embryos derived from mof1/+; mnkp6/+ females further confirms the role of DNA damage response gene Chk2 to ensure the removal of abnormal nuclei from the embryonic precursor pool and maintain genome stability. The fact that mof mutants undergo DNA damage has been further elucidated by the increased number of single and double stranded DNA breaks. Conclusion mof mutants exhibited genomic instability as evidenced by the occurance of frequent mitotic bridges in anaphase, asynchronous nuclear divisions, disruption of cytoskeleton, inactivation of centrosomes finally leading to DNA damage. Our findings are consistent to what has been reported earlier in mammals that; reduced levels of MOF resulted in increased genomic instability while total loss resulted in lethality. The study can be further extended using

  1. The effect of heavy ion on DNA damage checkpoint, and the discovery of its sensitizing compounds

    International Nuclear Information System (INIS)

    We demonstrated that Schisandrin B (SchB) and persimmon leaf extract (PLE) inhibited activation of checkpoint signaling molecule of cancer cells by heavy ion radiation. Therefore, SchB and PLE raise the possibility of sensitizing agents for heavy ion cancer therapy. In the present study, we examined whether SchB and PLE can sensitize the effect of heavy ion on tumor growth by using xenograft study. The oral administration of SchB and PLE for 31 days did not clearly reduce a volume of tumor in Scid mice transplanted A549 human adenocarcinoma cells. Tumor growth was obviously inhibited by the irradiation of heavy ion (C290 MeV/u, spread-out Bragg peak (SOBP)). SchB and PLE reduced the tumor volume in 3 Gy heavy ion exposed mice, respectively. Similarly, we examined whether SchB and PLE can sensitize the effect of X-ray also. SchB did not sensitize the effect of X-ray. However, PLE reduced the tumor volume in 6 Gy X-ray exposed mice. These results indicated that tumor volume in heavy ion exposed mice was inhibited by SchB and PLE treatment through the inhibition of DNA damage checkpoint signaling pathway. On the other hand, SchB could not sensitize the effect of X-ray on tumor growth. These results suggest that the modulating effects of SchB and PLE on DNA damage checkpoint are targeting independent molecules. These mechanisms should be clarified in further study. (author)

  2. Acute renal allograft rejection after immune checkpoint inhibitor therapy for metastatic melanoma.

    Science.gov (United States)

    Spain, L; Higgins, R; Gopalakrishnan, K; Turajlic, S; Gore, M; Larkin, J

    2016-06-01

    Immune checkpoint inhibitors such as ipilimumab and nivolumab improve survival in patients with advanced melanoma and are increasingly available to clinicians for use in the clinic. Their safety in organ transplant recipients is not well defined but published case reports describing treatment with ipilimumab have not been complicated by graft rejection. No cases of anti-programmed cell death protein 1 administration are reported in this group. We describe a case of acute graft rejection in a kidney transplant recipient after treatment with nivolumab, after progression on ipilimumab. Potential factors increasing the risk of graft rejection in this case are discussed, in particular the contribution of nivolumab. PMID:26951628

  3. Human Embryonic Stem Cells Are Capable of Executing G1/S Checkpoint Activation

    Czech Academy of Sciences Publication Activity Database

    Bárta, Tomáš; Vinarský, Vladimír; Holubcová, Z.; Doležalová, Dáša; Verner, J.; Pospíšilová, Š.; Dvořák, Petr; Hampl, Aleš

    2010-01-01

    Roč. 28, č. 7 (2010), s. 1143-1152. ISSN 1066-5099 Grant ostatní: GA MŠk(CZ) MSM0021622430; GA MŠk(CZ) 1M0538; GA MZd(CZ) NS10439; GA MŠk(CZ) MUNIE/E/0118/2009; EC FP6(XE) LSHG-CT-2006-018739 Institutional research plan: CEZ:AV0Z50390512; CEZ:AV0Z50390703 Keywords : human embryonic stem cells * DNA damage * checkpoint activation Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.871, year: 2010

  4. The cell-cycle checkpoint kinase Chk1 is required for mammalian homologous recombination repair

    DEFF Research Database (Denmark)

    Sørensen, Claus Storgaard; Hansen, Lasse Tengbjerg; Dziegielewski, Jaroslaw;

    2005-01-01

    The essential checkpoint kinase Chk1 is required for cell-cycle delays after DNA damage or blocked DNA replication. However, it is unclear whether Chk1 is involved in the repair of damaged DNA. Here we establish that Chk1 is a key regulator of genome maintenance by the homologous recombination......, the essential recombination repair protein RAD51 is recruited to DNA repair foci performing a vital role in correct HRR. We demonstrate that Chk1 interacts with RAD51, and that RAD51 is phosphorylated on Thr 309 in a Chk1-dependent manner. Consistent with a functional interplay between Chk1 and RAD51...

  5. Transparent Checkpoint-Restart for Hardware-Accelerated 3D Graphics

    OpenAIRE

    Nafchi, Samaneh Kazemi; Garg, Rohan; Cooperman, Gene

    2013-01-01

    Providing fault-tolerance for long-running GPU-intensive jobs requires application-specific solutions, and often involves saving the state of complex data structures spread among many graphics libraries. This work describes a mechanism for transparent GPU-independent checkpoint-restart of 3D graphics. The approach is based on a record-prune-replay paradigm: all OpenGL calls relevant to the graphics driver state are recorded; calls not relevant to the internal driver state as of the last graph...

  6. DNA damage-induced metaphase I arrest is mediated by the spindle assembly checkpoint and maternal age.

    Science.gov (United States)

    Marangos, Petros; Stevense, Michelle; Niaka, Konstantina; Lagoudaki, Michaela; Nabti, Ibtissem; Jessberger, Rolf; Carroll, John

    2015-01-01

    In mammalian oocytes DNA damage can cause chromosomal abnormalities that potentially lead to infertility and developmental disorders. However, there is little known about the response of oocytes to DNA damage. Here we find that oocytes with DNA damage arrest at metaphase of the first meiosis (MI). The MI arrest is induced by the spindle assembly checkpoint (SAC) because inhibiting the SAC overrides the DNA damage-induced MI arrest. Furthermore, this MI checkpoint is compromised in oocytes from aged mice. These data lead us to propose that the SAC is a major gatekeeper preventing the progression of oocytes harbouring DNA damage. The SAC therefore acts to integrate protection against both aneuploidy and DNA damage by preventing production of abnormal mature oocytes and subsequent embryos. Finally, we suggest escaping this DNA damage checkpoint in maternal ageing may be one of the causes of increased chromosome anomalies in oocytes and embryos from older mothers. PMID:26522734

  7. The Geography of Deterrence: Exploring the Small Area Effects of Sobriety Checkpoints on Alcohol-Impaired Collision Rates within a City

    Science.gov (United States)

    Nunn, Samuel; Newby, William

    2011-01-01

    This article examines alcohol-impaired collision metrics around nine sobriety checkpoint locations in Indianapolis, Indiana, before and after implementation of 22 checkpoints, using a pre/post examination, a pre/post nonequivalent comparison group analysis, and an interrupted time series approach. Traffic safety officials used geographical…

  8. The Rho-GAP Bem2p plays a GAP-independent role in the morphogenesis checkpoint

    OpenAIRE

    Marquitz, Aron R.; Harrison, Jacob C.; Bose, Indrani; Zyla, Trevin R.; McMillan, John N.; Lew, Daniel J.

    2002-01-01

    The Saccharomyces cerevisiae morphogenesis checkpoint delays mitosis in response to insults that impair actin organization and/or bud formation. The delay is due to accumulation of the inhibitory kinase Swe1p, which phosphorylates the cyclin-dependent kinase Cdc28p. Having screened through a panel of yeast mutants with defects in cell morphogenesis, we report here that the polarity establishment protein Bem2p is required for the checkpoint response. Bem2p is a Rho-GTPase activating protein (G...

  9. Role of Intrinsic and Extrinsic Factors in the Regulation of the Mitotic Checkpoint Kinase Bub1

    Science.gov (United States)

    Breit, Claudia; Bange, Tanja; Petrovic, Arsen; Weir, John R.; Müller, Franziska; Vogt, Doro; Musacchio, Andrea

    2015-01-01

    The spindle assembly checkpoint (SAC) monitors microtubule attachment to kinetochores to ensure accurate sister chromatid segregation during mitosis. The SAC members Bub1 and BubR1 are paralogs that underwent significant functional specializations during evolution. We report an in-depth characterization of the kinase domains of Bub1 and BubR1. BubR1 kinase domain binds nucleotides but is unable to deliver catalytic activity in vitro. Conversely, Bub1 is an active kinase regulated by intra-molecular phosphorylation at the P+1 loop. The crystal structure of the phosphorylated Bub1 kinase domain illustrates a hitherto unknown conformation of the P+1 loop docked into the active site of the Bub1 kinase. Both Bub1 and BubR1 bind Bub3 constitutively. A hydrodynamic characterization of Bub1:Bub3 and BubR1:Bub3 demonstrates both complexes to have 1:1 stoichiometry, with no additional oligomerization. Conversely, Bub1:Bub3 and BubR1:Bub3 combine to form a heterotetramer. Neither BubR1:Bub3 nor Knl1, the kinetochore receptor of Bub1:Bub3, modulate the kinase activity of Bub1 in vitro, suggesting autonomous regulation of the Bub1 kinase domain. We complement our study with an analysis of the Bub1 substrates. Our results contribute to the mechanistic characterization of a crucial cell cycle checkpoint. PMID:26658523

  10. Tumor-suppressor genes, cell cycle regulatory checkpoints, and the skin

    Directory of Open Access Journals (Sweden)

    Ana Maria Abreu Velez

    2015-01-01

    Full Text Available The cell cycle (or cell-division cycle is a series of events that take place in a cell, leading to its division and duplication. Cell division requires cell cycle checkpoints (CPs that are used by the cell to both monitor and regulate the progress of the cell cycle. Tumor-suppressor genes (TSGs or antioncogenes are genes that protect the cell from a single event or multiple events leading to cancer. When these genes mutate, the cell can progress to a cancerous state. We aimed to perform a narrative review, based on evaluation of the manuscripts published in MEDLINE-indexed journals using the Medical Subject Headings (MeSH terms "tumor suppressor′s genes," "skin," and "cell cycle regulatory checkpoints." We aimed to review the current concepts regarding TSGs, CPs, and their association with selected cutaneous diseases. It is important to take into account that in some cell cycle disorders, multiple genetic abnormalities may occur simultaneously. These abnormalities may include intrachromosomal insertions, unbalanced division products, recombinations, reciprocal deletions, and/or duplication of the inserted segments or genes; thus, these presentations usually involve several genes. Due to their complexity, these disorders require specialized expertise for proper diagnosis, counseling, personal and family support, and genetic studies. Alterations in the TSGs or CP regulators may occur in many benign skin proliferative disorders, neoplastic processes, and genodermatoses.

  11. Anti-TNF-refractory colitis after checkpoint inhibitor therapy: Possible role of CMV-mediated immunopathogenesis.

    Science.gov (United States)

    Lankes, Katharina; Hundorfean, Gheorghe; Harrer, Thomas; Pommer, Ansgar J; Agaimy, Abbas; Angelovska, Irena; Tajmir-Riahi, Azadeh; Göhl, Jonas; Schuler, Gerold; Neurath, Markus F; Hohenberger, Werner; Heinzerling, Lucie

    2016-06-01

    Immune-related adverse events (irAEs) induced by checkpoint inhibitors are well known. Since fatal outcomes have been reported early detection and adequate management are crucial. In particular, colitis is frequently observed and can result in intestinal perforation. This is the first report of an autoimmune colitis that was treated according to algorithms but became resistant due to a CMV reactivation. The 32-y-old male patient with metastatic melanoma treated within an anti-PD-1/ipilimumab combination study developed severe immune-mediated colitis (CTCAE grade 3) with up to 18 watery stools per day starting 2 weeks after treatment initiation. After improving upon therapy with immunosuppressive treatment (high dose steroids and infliximab) combined with parenteral nutrition diarrhea again exacerbated. Additionally, the patient had asymptomatic grade 3 CTCAE amylase and lipase elevation. Colitis was monitored by weekly endoscopies and colon biopsies were analyzed histologically with CMV staining, multi-epitope ligand cartography (MELC) and qRT-PCR for inflammatory genes. In the course, CMV reactivation was detected in the colon and treated with antiviral medication in parallel to a reduction of corticosteroids. Subsequently, symptoms improved. The patient showed a complete response for 2 y now including regression of bone metastases. CMV reactivation under checkpoint inhibitor therapy in combination with immunosuppressive treatment for autoimmune side effects has to be considered in these patients and if present treated. Potentially, CMV reactivation is underdiagnosed. Treatment algorithms should include CMV diagnostics. PMID:27471608

  12. A2aR antagonists: Next generation checkpoint blockade for cancer immunotherapy

    Directory of Open Access Journals (Sweden)

    Robert D. Leone

    2015-01-01

    Full Text Available The last several years have witnessed exciting progress in the development of immunotherapy for the treatment of cancer. This has been due in great part to the development of so-called checkpoint blockade. That is, antibodies that block inhibitory receptors such as CTLA-4 and PD-1 and thus unleash antigen-specific immune responses against tumors. It is clear that tumors evade the immune response by usurping pathways that play a role in negatively regulating normal immune responses. In this regard, adenosine in the immune microenvironment leading to the activation of the A2a receptor has been shown to represent one such negative feedback loop. Indeed, the tumor microenvironment has relatively high concentrations of adenosine. To this end, blocking A2a receptor activation has the potential to markedly enhance anti-tumor immunity in mouse models. This review will present data demonstrating the ability of A2a receptor blockade to enhance tumor vaccines, checkpoint blockade and adoptive T cell therapy. Also, as several recent studies have demonstrated that under certain conditions A2a receptor blockade can enhance tumor progression, we will also explore the complexities of adenosine signaling in the immune response. Despite important nuances to the A2a receptor pathway that require further elucidation, studies to date strongly support the development of A2a receptor antagonists (some of which have already been tested in phase III clinical trials for Parkinson Disease as novel modalities in the immunotherapy armamentarium.

  13. Estimated Interval-Based Checkpointing (EIC on Spot Instances in Cloud Computing

    Directory of Open Access Journals (Sweden)

    Daeyong Jung

    2014-01-01

    Full Text Available In cloud computing, users can rent computing resources from service providers according to their demand. Spot instances are unreliable resources provided by cloud computing services at low monetary cost. When users perform tasks on spot instances, there is an inevitable risk of failures that causes the delay of task execution time, resulting in a serious deterioration of quality of service (QoS. To deal with the problem on spot instances, we propose an estimated interval-based checkpointing (EIC using weighted moving average. Our scheme sets the thresholds of price and execution time based on history. Whenever the actual price and the execution time cross over the thresholds, the system saves the state of spot instances. The Bollinger Bands is adopted to inform the ranges of estimated cost and execution time for user's discretion. The simulation results reveal that, compared to the HBC and REC, the EIC reduces the number of checkpoints and the rollback time. Consequently, the task execution time is decreased with EIC by HBC and REC. The EIC also provides the benefit of the cost reduction by HBC and REC, on average. We also found that the actual cost and execution time fall within the estimated ranges suggested by the Bollinger Bands.

  14. The impact of aviation checkpoint queues on optimizing security screening effectiveness

    International Nuclear Information System (INIS)

    Passenger screening at aviation security checkpoints is a critical component in protecting airports and aircraft from terrorist threats. Recent developments in screening device technology have increased the ability to detect these threats; however, the average amount of time it takes to screen a passenger still remains a concern. This paper models the queueing process for a multi-level airport checkpoint security system, where multiple security classes are formed through subsets of specialized screening devices. An optimal static assignment policy is obtained which minimizes the steady-state expected amount of time a passenger spends in the security system. Then, an optimal dynamic assignment policy is obtained through a transient analysis that balances the expected number of true alarms with the expected amount of time a passenger spends in the security system. Performance of a two-class system is compared to that of a selective security system containing primary and secondary levels of screening. The key contribution is that the resulting optimal assignment policies increase security and passenger throughput by efficiently and effectively utilizing available screening resources.

  15. Mad2 binding to Mad1 and Cdc20, rather than oligomerization, is required for the spindle checkpoint

    DEFF Research Database (Denmark)

    Sironi, L; Melixetian, M; Faretta, M;

    2001-01-01

    Mad2 is a key component of the spindle checkpoint, a device that controls the fidelity of chromosome segregation in mitosis. The ability of Mad2 to form oligomers in vitro has been correlated with its ability to block the cell cycle upon injection into Xenopus embryos. Here we show that Mad2 forms...

  16. The E3 ubiquitin ligase EDD regulates S-phase and G(2)/M DNA damage checkpoints.

    Science.gov (United States)

    Munoz, Marcia A; Saunders, Darren N; Henderson, Michelle J; Clancy, Jennifer L; Russell, Amanda J; Lehrbach, Gillian; Musgrove, Elizabeth A; Watts, Colin K W; Sutherland, Robert L

    2007-12-15

    The cellular response to DNA damage is critical for maintenance of genomic integrity and inhibition of tumorigenesis. Mutations or aberrant expression of the E3 ubiquitin ligase EDD have been observed in a number of carcinomas and we recently reported that EDD modulates activity of the DNA damage checkpoint kinase, CHK2. Here, we demonstrate that EDD is necessary for G(1)/S and intra S phase DNA damage checkpoint activation and for the maintenance of G(2)/M arrest after double strand DNA breaks. Defective checkpoint activation in EDD-depleted cells led to radio-resistant DNA synthesis, premature entry into mitosis, accumulation of polyploid cells, and cell death via mitotic catastrophe. In addition to decreased CHK2 activation in EDD-depleted cells, the expression of several key cell cycle mediators including Cdc25A/C and E2F1 was altered, suggesting that these checkpoint defects may be both CHK2-dependent and -independent. These data support a role for EDD in the maintenance of genomic stability, emphasising the potential importance of dysregulated EDD expression and/or function in the evolution of cancer. PMID:18073532

  17. Wip1 phosphatase is associated with chromatin and dephosphorylates gammaH2AX to promote checkpoint inhibition

    Czech Academy of Sciences Publication Activity Database

    Macůrek, Libor; Lindqvist, A.; Voets, O.; Kool, J.; Vos, H.R.; Medema, R.H.

    2010-01-01

    Roč. 29, č. 15 (2010), s. 2281-2291. ISSN 0950-9232 R&D Projects: GA ČR GPP305/10/P420 Institutional research plan: CEZ:AV0Z50520514 Keywords : DNA damage * checkpoint * phosphatase Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.414, year: 2010

  18. Killing the umpire: cooperative defects in mitotic checkpoint and BRCA2 genes on the road to transformation

    International Nuclear Information System (INIS)

    Recent findings from mouse models of BRCA2 genetic lesions have provided intriguing insights and important questions concerning modes of tumor development in familial breast and ovarian cancers. Fibroblasts from mice homozygous for the BRCA2Tr allele grow poorly and display an array of chromosomal abnormalities that are consistent with a role for BRCA2 in DNA repair. This growth defect can be overcome and cellular transformation promoted by the expression of defective, dominant negative alleles of p53 and of the mitotic checkpoint gene Bub1, both of which are known to induce chromosome instability. These findings are mirrored in the genetic lesions sustained in tumors found in the rare BRCA2Tr/Trmice that survive to adulthood, which include defects in p53 as well as the mitotic checkpoint proteins Bub1 and Mad3L. Together, these data hint that tumors in these mice evolve from an unusually intense selective pressure to remove DNA damage checkpoints, which in turn might be facilitated by chromosomal abolition of mitotic checkpoints and the consequent increase in shuffling of genetic information. How these genetic lesions co-operate to yield transformed cells and how these data relate to BRCA1 and BRCA2 defects in the human population are important questions raised by this work

  19. Toxicities of the anti-PD-1 and anti-PD-L1 immune checkpoint antibodies.

    Science.gov (United States)

    Naidoo, J; Page, D B; Li, B T; Connell, L C; Schindler, K; Lacouture, M E; Postow, M A; Wolchok, J D

    2015-12-01

    Immune checkpoint antibodies that augment the programmed cell death protein 1 (PD-1)/PD-L1 pathway have demonstrated antitumor activity across multiple malignancies, and gained recent regulatory approval as single-agent therapy for the treatment of metastatic malignant melanoma and nonsmall-cell lung cancer. Knowledge of toxicities associated with PD-1/PD-L1 blockade, as well as effective management algorithms for these toxicities, is pivotal in order to optimize clinical efficacy and safety. In this article, we review selected published and presented clinical studies investigating single-agent anti-PD-1/PD-L1 therapy and trials of combination approaches with other standard anticancer therapies, in multiple tumor types. We summarize the key adverse events reported in these studies and their management algorithms. PMID:26371282

  20. The Aurora B kinase in chromosome biorientation and spindle checkpoint signalling

    Directory of Open Access Journals (Sweden)

    Veronica eKrenn

    2015-10-01

    Full Text Available Aurora B, a member of the Aurora family of serine/threonine protein kinases, is a key player in chromosome segregation. As part of a macromolecular complex known as the chromosome passenger complex, Aurora B concentrates early during mitosis in the proximity of centromeres and kinetochores, the sites of attachment of chromosomes to spindle microtubules. There, it contributes to a number of processes that impart fidelity to cell division, including kinetochore stabilization, kinetochore-microtubule attachment, and the regulation of a surveillance mechanism named the spindle assembly checkpoint. In the regulation of these processes, Aurora B is the fulcrum of a remarkably complex network of interactions that feed back on its localization and activation state. In this review we discuss the multiple roles of Aurora B during mitosis, focusing in particular on its role at centromeres and kinetochores. Many details of the network of interactions at these locations remain poorly understood, and we focus here on several crucial outstanding questions.

  1. Clonal neoantigens elicit T cell immunoreactivity and sensitivity to immune checkpoint blockade.

    Science.gov (United States)

    McGranahan, Nicholas; Furness, Andrew J S; Rosenthal, Rachel; Ramskov, Sofie; Lyngaa, Rikke; Saini, Sunil Kumar; Jamal-Hanjani, Mariam; Wilson, Gareth A; Birkbak, Nicolai J; Hiley, Crispin T; Watkins, Thomas B K; Shafi, Seema; Murugaesu, Nirupa; Mitter, Richard; Akarca, Ayse U; Linares, Joseph; Marafioti, Teresa; Henry, Jake Y; Van Allen, Eliezer M; Miao, Diana; Schilling, Bastian; Schadendorf, Dirk; Garraway, Levi A; Makarov, Vladimir; Rizvi, Naiyer A; Snyder, Alexandra; Hellmann, Matthew D; Merghoub, Taha; Wolchok, Jedd D; Shukla, Sachet A; Wu, Catherine J; Peggs, Karl S; Chan, Timothy A; Hadrup, Sine R; Quezada, Sergio A; Swanton, Charles

    2016-03-25

    As tumors grow, they acquire mutations, some of which create neoantigens that influence the response of patients to immune checkpoint inhibitors. We explored the impact of neoantigen intratumor heterogeneity (ITH) on antitumor immunity. Through integrated analysis of ITH and neoantigen burden, we demonstrate a relationship between clonal neoantigen burden and overall survival in primary lung adenocarcinomas. CD8(+)tumor-infiltrating lymphocytes reactive to clonal neoantigens were identified in early-stage non-small cell lung cancer and expressed high levels of PD-1. Sensitivity to PD-1 and CTLA-4 blockade in patients with advanced NSCLC and melanoma was enhanced in tumors enriched for clonal neoantigens. T cells recognizing clonal neoantigens were detectable in patients with durable clinical benefit. Cytotoxic chemotherapy-induced subclonal neoantigens, contributing to an increased mutational load, were enriched in certain poor responders. These data suggest that neoantigen heterogeneity may influence immune surveillance and support therapeutic developments targeting clonal neoantigens. PMID:26940869

  2. Meiosis I in Xenopus oocytes is not error-prone despite lacking spindle assembly checkpoint.

    Science.gov (United States)

    Liu, Dandan; Shao, Hua; Wang, Hongmei; Liu, X Johné

    2014-01-01

    The spindle assembly checkpoint, SAC, is a surveillance mechanism to control the onset of anaphase during cell division. SAC prevents anaphase initiation until all chromosome pairs have achieved bipolar attachment and aligned at the metaphase plate of the spindle. In doing so, SAC is thought to be the key mechanism to prevent chromosome nondisjunction in mitosis and meiosis. We have recently demonstrated that Xenopus oocyte meiosis lacks SAC control. This prompted the question of whether Xenopus oocyte meiosis is particularly error-prone. In this study, we have karyotyped a total of 313 Xenopus eggs following in vitro oocyte maturation. We found no hyperploid egg, out of 204 metaphase II eggs with countable chromosome spreads. Therefore, chromosome nondisjunction is very rare during Xenopus oocyte meiosis I, despite the lack of SAC. PMID:24646611

  3. Clonal neoantigens elicit T cell immunoreactivity and sensitivity to immune checkpoint blockade

    Science.gov (United States)

    McGranahan, Nicholas; Furness, Andrew J. S.; Rosenthal, Rachel; Ramskov, Sofie; Lyngaa, Rikke; Saini, Sunil Kumar; Jamal-Hanjani, Mariam; Wilson, Gareth A.; Birkbak, Nicolai J.; Hiley, Crispin T.; Watkins, Thomas B. K.; Shafi, Seema; Murugaesu, Nirupa; Mitter, Richard; Akarca, Ayse U.; Linares, Joseph; Marafioti, Teresa; Henry, Jake Y.; Van Allen, Eliezer M.; Miao, Diana; Schilling, Bastian; Schadendorf, Dirk; Garraway, Levi A.; Makarov, Vladimir; Rizvi, Naiyer A.; Snyder, Alexandra; Hellmann, Matthew D.; Merghoub, Taha; Wolchok, Jedd D.; Shukla, Sachet A.; Wu, Catherine J.; Peggs, Karl S.; Chan, Timothy A.; Hadrup, Sine R.; Quezada, Sergio A.; Swanton, Charles

    2016-01-01

    As tumors grow, they acquire mutations, some of which create neoantigens that influence the response of patients to immune checkpoint inhibitors. We explored the impact of neoantigen intratumor heterogeneity (ITH) on antitumor immunity. Through integrated analysis of ITH and neoantigen burden, we demonstrate a relationship between clonal neoantigen burden and overall survival in primary lung adenocarcinomas. CD8+ tumor-infiltrating lymphocytes reactive to clonal neoantigens were identified in early-stage non–small cell lung cancer and expressed high levels of PD-1. Sensitivity to PD-1 and CTLA-4 blockade in patients with advanced NSCLC and melanoma was enhanced in tumors enriched for clonal neoantigens. T cells recognizing clonal neoantigens were detectable in patients with durable clinical benefit. Cytotoxic chemotherapy–induced subclonal neoantigens, contributing to an increased mutational load, were enriched in certain poor responders. These data suggest that neoantigen heterogeneity may influence immune surveillance and support therapeutic developments targeting clonal neoantigens. PMID:26940869

  4. Regulation of Sphingolipid Biosynthesis by the Morphogenesis Checkpoint Kinase Swe1.

    Science.gov (United States)

    Chauhan, Neha; Han, Gongshe; Somashekarappa, Niranjanakumari; Gable, Kenneth; Dunn, Teresa; Kohlwein, Sepp D

    2016-01-29

    Sphingolipid (SL) biosynthesis is negatively regulated by the highly conserved endoplasmic reticulum-localized Orm family proteins. Defective SL synthesis in Saccharomyces cerevisiae leads to increased phosphorylation and inhibition of Orm proteins by the kinase Ypk1. Here we present evidence that the yeast morphogenesis checkpoint kinase, Swe1, regulates SL biosynthesis independent of the Ypk1 pathway. Deletion of the Swe1 kinase renders mutant cells sensitive to serine palmitoyltransferase inhibition due to impaired sphingoid long-chain base synthesis. Based on these data and previous results, we suggest that Swe1 kinase perceives alterations in SL homeostasis, activates SL synthesis, and may thus represent the missing regulatory link that controls the SL rheostat during the cell cycle. PMID:26634277

  5. Regulation of Sphingolipid Biosynthesis by the Morphogenesis Checkpoint Kinase Swe1*

    Science.gov (United States)

    Chauhan, Neha; Han, Gongshe; Somashekarappa, Niranjanakumari; Gable, Kenneth; Dunn, Teresa; Kohlwein, Sepp D.

    2016-01-01

    Sphingolipid (SL) biosynthesis is negatively regulated by the highly conserved endoplasmic reticulum-localized Orm family proteins. Defective SL synthesis in Saccharomyces cerevisiae leads to increased phosphorylation and inhibition of Orm proteins by the kinase Ypk1. Here we present evidence that the yeast morphogenesis checkpoint kinase, Swe1, regulates SL biosynthesis independent of the Ypk1 pathway. Deletion of the Swe1 kinase renders mutant cells sensitive to serine palmitoyltransferase inhibition due to impaired sphingoid long-chain base synthesis. Based on these data and previous results, we suggest that Swe1 kinase perceives alterations in SL homeostasis, activates SL synthesis, and may thus represent the missing regulatory link that controls the SL rheostat during the cell cycle. PMID:26634277

  6. Checkpoint and Replication Oriented Fault Tolerant Mechanism for MapReduce Framework

    Directory of Open Access Journals (Sweden)

    Yang Liu

    2013-09-01

    Full Text Available MapReduce is an emerging programming paradigm and an associated implementation for processing and generating big data which has been widely applied in data-intensive systems. In cloud environment, node and task failure is no longer accidental but a common feature of large-scale systems. In MapReduce framework, although the rescheduling based fault-tolerant method is simple to implement, it failed to fully consider the location of distributed data, the computation and storage overhead. Thus, a single node failure will increase the completion time dramatically. In this paper, a Checkpoint and Replication Oriented Fault Tolerant scheduling algorithm (CROFT is proposed, which takes both task and node failure into consideration. Preliminary experiments show that with less storage and network overhead. CROFT will significantly reduce the completion time at failure time, and the overall performance of MapReduce can be improved at least over 30% than original mechanism in Hadoop.  

  7. Chromosome 17p deletion in human medulloblastoma: a missing checkpoint in the Hedgehog pathway.

    Science.gov (United States)

    De Smaele, Enrico; Di Marcotullio, Lucia; Ferretti, Elisabetta; Screpanti, Isabella; Alesse, Edoardo; Gulino, Alberto

    2004-10-01

    Although deregulation of Hedgehog signalling is considered to play a crucial oncogenic role and commonly occurrs in medulloblastoma, genetic lesions in components of this pathway are observed in a minority of cases. The recent identification of a novel putative tumor suppressor (REN(KCTD11)) on chromosome 17p13.2, a region most frequently lost in human medulloblastoma, highlights the role of allelic deletion of the gene in this brain malignancy, leading to the loss of growth inhibitory activity via suppression of Gli-dependent activation of Hedgehog target genes. The presence on 17p13 of another tumor suppressor gene (p53) whose inactivation cooperates with Hedgehog pathway for medulloblastoma formation, suggests that 17p deletion unveils haploinsufficiency conditions leading to abrogation of either direct and indirect checkpoints of Hedgehog signalling in cancer. PMID:15467454

  8. Immune checkpoint inhibitors enhance cytotoxicity of cytokine-induced killer cells against human myeloid leukaemic blasts.

    Science.gov (United States)

    Poh, Su Li; Linn, Yeh Ching

    2016-05-01

    We studied whether blockade of inhibitory receptors on cytokine-induced killer (CIK) cells by immune checkpoint inhibitors could increase its anti-tumour potency against haematological malignancies. CIK cultures were generated from seven normal donors and nine patients with acute myeloid leukaemia (AML), acute lymphoblastic leukaemia (ALL) or multiple myeloma (MM). The inhibitory receptors B and T lymphocyte attenuator, CD200 receptor, lymphocyte activation gene-3 (LAG-3) and T cell immunoglobulin and mucin-domain-containing-3 (TIM-3) were present at variable percentages in most CIK cultures, while cytotoxic T lymphocyte-associated protein 4 (CTLA-4), programmed death-1 (PD-1) and killer cell immunoglobulin-like receptors (KIR2DL1/2/3) were expressed at low level in most cultures. Without blockade, myeloid leukaemia cells were susceptible to autologous and allogeneic CIK-mediated cytotoxicity. Blockade of KIR, LAG-3, PD-1 and TIM-3 but not CTLA-4 resulted in remarkable increase in killing against these targets, even in those with poor baseline cytotoxicity. ALL and MM targets were resistant to CIK-mediated cytotoxicity, and blockade of receptors did not increase cytotoxicity to a meaningful extent. Combination of inhibitors against two receptors did not further increase cytotoxicity. Interestingly, potentiation of CIK killing by blocking antibodies was not predicted by expression of receptors on CIK and their respective ligands on the targets. Compared to un-activated T and NK cells, blockade potentiated the cytotoxicity of CIK cells to a greater degree and at a lower E:T ratio, but without significant increase in cytotoxicity against normal white cell. Our findings provide the basis for clinical trial combining autologous CIK cells with checkpoint inhibitors for patients with AML. PMID:26961084

  9. Genetic Control of the Trigger for the G2/M Checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Hall, Eric J. [Columbia University; Smilenov, Lubomir B. [Columbia University; Young, Erik F. [Columbia University

    2013-10-01

    The work undertaken in this project addressed two seminal areas of low dose radiation biology that are poorly understood and controversial. These areas are the challenge to the linear-no-threshold (LNT) paradigm at low doses of radiation and, the fundamental elements of radiation bystander effect biology Genetic contributions to low dose checkpoint engagement: The LNT paradigm is an extrapolation of known, measured cancer induction endpoints. Importantly, data for lower doses is often not available. Debatably, radiation protection standards have been introduced which are prudently contingent on the adherence of cancer risk to the established trend seen at higher doses. Intriguing findings from other labs have hinted at separate DNA damage response programs that engage at low or high levels of radiation. Individual radiation sensitivity commensurate with hemizygosity for a radiation sensitivity gene has been estimated at 1-2% in the U.S.. Careful interrogation of the DNA damage response at low doses of radiation became important and served as the basis for this grant. Several genes were tested in combinations to determine if combined haploinsufficiency for multiple radiosensitizing genes could render a cell more sensitive to lower levels of acute radiation exposure. We measured a classical radiation response endpoint, cell cycle arrest prior to mitosis. Mouse embryo fibroblasts were used and provided a uniform, rapidly dividing and genetically manipulable population of study. Our system did not report checkpoint engagement at acute doses of gamma rays below 100 mGy. The system did report checkpoint engagement reproducibly at 500 mGy establishing a threshold for activation between 100 and 500 mGy. Engagement of the checkpoint was ablated in cells nullizygous for ATM but was otherwise unperturbed in cells combinatorially haploinsufficient for ATM and Rad9, ATM and PTEN or PTEN and Rad9. Taken together, these experiments tell us that, in a sensitive fibroblast culture

  10. Xenopus Cds1 Is Regulated by DNA-Dependent Protein Kinase and ATR during the Cell Cycle Checkpoint Response to Double-Stranded DNA Ends

    OpenAIRE

    McSherry, Troy D.; Mueller, Paul R.

    2004-01-01

    The checkpoint kinase Cds1 (Chk2) plays a key role in cell cycle checkpoint responses with functions in cell cycle arrest, DNA repair, and induction of apoptosis. Proper regulation of Cds1 is essential for appropriate cellular responses to checkpoint-inducing insults. While the kinase ATM has been shown to be important in the regulation of human Cds1 (hCds1), here we report that the kinases ATR and DNA-dependent protein kinase (DNA-PK) play more significant roles in the regulation of Xenopus ...

  11. Ipl1p-dependent phosphorylation of Mad3p is required for the spindle checkpoint response to lack of tension at kinetochores

    OpenAIRE

    King, Emma M J; Rachidi, Najma; Morrice, Nick; Hardwick, Kevin G.; Stark, Michael J. R.

    2007-01-01

    The spindle checkpoint delays anaphase onset until all chromosomes are correctly attached to microtubules. Ipl1 protein kinase (Aurora B) is required to correct inappropriate kinetochore-microtubule attachments and for the response to lack of tension between sister kinetochores. Here we identify residues in the checkpoint protein Mad3p that are phosphorylated by Ipl1p. When phosphorylation of Mad3p at two sites is prevented, the cell's response to reduced kinetochore tension is dramatically c...

  12. Ipl1p-dependent phosphorylation of Mad3p is required for the spindle checkpoint response to lack of tension at kinetochores

    OpenAIRE

    King, Emma M J; Rachidi, Najma; Morrice, Nick; Hardwick, Kevin G.; Stark, Michael J. R.

    2007-01-01

    The spindle checkpoint delays anaphase onset until all chromosomes are correctly attached to microtubules. Ipl1 protein kinase (Aurora B) is required to correct inappropriate kinetochore–microtubule attachments and for the response to lack of tension between sister kinetochores. Here we identify residues in the checkpoint protein Mad3p that are phosphorylated by Ipl1p. When phosphorylation of Mad3p at two sites is prevented, the cell’s response to reduced kinetochore tension is dramatically c...

  13. In vitro expression levels of cell-cycle checkpoint proteins are associated with cellular DNA repair capacity in peripheral blood lymphocytes: a multivariate analysis

    OpenAIRE

    Fan, You-Hong; Hu, Zhibin; Li, Chunying; Wang, Li-E; Guo, Zhaozheng; Qiao, Yawei; Zhang, Li; Zhang, Wei; Mao, Li; Wei, Qingyi

    2007-01-01

    DNA repair should occur after cells sense DNA damage signals and undergo cell-cycle arrest to provide sufficient time for DNA repair, and suboptimal DNA repair capacity (DRC) in peripheral lymphocytes has been suggested as a cancer susceptibility marker. Numerous studies showed a functional link between DNA damage sensing, cell-cycle checkpoint and DNA repair. We hypothesized that in vitro cell-cycle checkpoint-related protein expression levels in stimulated lymphocytes predict DRC levels. To...

  14. Direct and indirect control of the initiation of meiotic recombination by DNA damage checkpoint mechanisms in budding yeast.

    Directory of Open Access Journals (Sweden)

    Bilge Argunhan

    Full Text Available Meiotic recombination plays an essential role in the proper segregation of chromosomes at meiosis I in many sexually reproducing organisms. Meiotic recombination is initiated by the scheduled formation of genome-wide DNA double-strand breaks (DSBs. The timing of DSB formation is strictly controlled because unscheduled DSB formation is detrimental to genome integrity. Here, we investigated the role of DNA damage checkpoint mechanisms in the control of meiotic DSB formation using budding yeast. By using recombination defective mutants in which meiotic DSBs are not repaired, the effect of DNA damage checkpoint mutations on DSB formation was evaluated. The Tel1 (ATM pathway mainly responds to unresected DSB ends, thus the sae2 mutant background in which DSB ends remain intact was employed. On the other hand, the Mec1 (ATR pathway is primarily used when DSB ends are resected, thus the rad51 dmc1 double mutant background was employed in which highly resected DSBs accumulate. In order to separate the effect caused by unscheduled cell cycle progression, which is often associated with DNA damage checkpoint defects, we also employed the ndt80 mutation which permanently arrests the meiotic cell cycle at prophase I. In the absence of Tel1, DSB formation was reduced in larger chromosomes (IV, VII, II and XI whereas no significant reduction was found in smaller chromosomes (III and VI. On the other hand, the absence of Rad17 (a critical component of the ATR pathway lead to an increase in DSB formation (chromosomes VII and II were tested. We propose that, within prophase I, the Tel1 pathway facilitates DSB formation, especially in bigger chromosomes, while the Mec1 pathway negatively regulates DSB formation. We also identified prophase I exit, which is under the control of the DNA damage checkpoint machinery, to be a critical event associated with down-regulating meiotic DSB formation.

  15. Spindle checkpoint activation at meiosis I advances anaphase II onset via meiosis-specific APC/C regulation

    OpenAIRE

    Yamamoto, Ayumu; Kitamura, Kenji; Hihara, Daisuke; Hirose, Yukinobu; Katsuyama, Satoshi; Hiraoka, Yasushi

    2008-01-01

    During mitosis, the spindle assembly checkpoint (SAC) inhibits the Cdc20-activated anaphase-promoting complex/cyclosome (APC/CCdc20), which promotes protein degradation, and delays anaphase onset to ensure accurate chromosome segregation. However, the SAC function in meiotic anaphase regulation is poorly understood. Here, we examined the SAC function in fission yeast meiosis. As in mitosis, a SAC factor, Mad2, delayed anaphase onset via Slp1 (fission yeast Cdc20) when chromosomes attach to th...

  16. Targeting of Carbon Ion-Induced G2 Checkpoint Activation in Lung Cancer Cells Using Wee-1 Inhibitor MK-1775.

    Science.gov (United States)

    Ma, Hongyu; Takahashi, Akihisa; Sejimo, Yukihiko; Adachi, Akiko; Kubo, Nobuteru; Isono, Mayu; Yoshida, Yukari; Kanai, Tatsuaki; Ohno, Tatsuya; Nakano, Takashi

    2015-12-01

    The potent inhibitor of the cell cycle checkpoint regulatory factor Wee-1, MK-1775, has been reported to enhance non-small cell lung cancer (NSCLC) cell sensitivity to photon radiation by abrogating radiation-induced G2 arrest. However, little is known about the effects of this sensitizer after exposure to carbon (C)-ion radiation. The purpose of this study was therefore to investigate the effects of C ions in combination with MK-1775 on the killing of NSCLC cells. Human NSCLC H1299 cells were exposed to X rays or C ions (290 MeV/n, 50 keV/μm at the center of a 6 cm spread-out Bragg peak) in the presence of MK-1775. The cell cycle was analyzed using flow cytometry and Western blotting. Radiosensitivity was determined using clonogenic survival assays. The mechanisms underlying MK-1775 radiosensitization were studied by observing H2AX phosphorylation and mitotic catastrophe. G2 checkpoint arrest was enhanced 2.3-fold by C-ion exposure compared with X-ray exposure. Radiation-induced G2 checkpoint arrest was abrogated by MK-1775. Exposure to radiation resulted in a significant reduction in the mitotic ratio and increased phosphorylation of cyclin-dependent kinase 1 (Cdk1), the primary downstream mediator of Wee-1-induced G2 arrest. The Wee-1 inhibitor, MK-1775 restored the mitotic ratio and suppressed Cdk1 phosphorylation. In addition, MK-1775 increased H1299 cell sensitivity to C ions and X rays independent of TP53 status. MK-1775 also significantly increased H2AX phosphorylation and mitotic catastrophe in irradiated cells. These results suggest that the G2 checkpoint inhibitor MK-1775 can enhance the sensitivity of human NSCLC cells to C ions as well as X rays. PMID:26645158

  17. Current status and perspectives in translational biomarker research for PD-1/PD-L1 immune checkpoint blockade therapy.

    Science.gov (United States)

    Ma, Weijie; Gilligan, Barbara M; Yuan, Jianda; Li, Tianhong

    2016-01-01

    Modulating immune inhibitory pathways has been a major recent breakthrough in cancer treatment. Checkpoint blockade antibodies targeting cytotoxic T-lymphocyte antigen 4 (CTLA-4) and programed cell-death protein 1 (PD-1) have demonstrated acceptable toxicity, promising clinical responses, durable disease control, and improved survival in some patients with advanced melanoma, non-small cell lung cancer (NSCLC), and other tumor types. About 20 % of advanced NSCLC patients and 30 % of advanced melanoma patients experience tumor responses from checkpoint blockade monotherapy, with better clinical responses seen with the combination of anti-PD-1 and anti-CTLA-4 antibodies. Given the power of these new therapies, it is important to understand the complex and dynamic nature of host immune responses and the regulation of additional molecules in the tumor microenvironment and normal organs in response to the checkpoint blockade therapies. In this era of precision oncology, there remains a largely unmet need to identify the patients who are most likely to benefit from immunotherapy, to optimize the monitoring assays for tumor-specific immune responses, to develop strategies to improve clinical efficacy, and to identify biomarkers so that immune-related adverse events can be avoided. At this time, PD-L1 immunohistochemistry (IHC) staining using 22C3 antibody is the only FDA-approved companion diagnostic for patients with NSCLC-treated pembrolizumab, but more are expected to come to market. We here summarize the current knowledge, clinical efficacy, potential immune biomarkers, and associated assays for immune checkpoint blockade therapies in advanced solid tumors. PMID:27234522

  18. The tumor suppressor homolog in fission yeast, myh1+, displays a strong interaction with the checkpoint gene rad1+

    International Nuclear Information System (INIS)

    The DNA glycosylase MutY is strongly conserved in evolution, and homologs are found in most eukaryotes and prokaryotes examined. This protein is implicated in repair of oxidative DNA damage, in particular adenine mispaired opposite 7,8-dihydro-8-oxoguanine. Previous investigations in Escherichia coli, fission yeast, and mammalian cells show an association of mutations in MutY homologs with a mutator phenotype and carcinogenesis. Eukaryotic MutY homologs physically associate with several proteins with a role in replication, DNA repair, and checkpoint signaling, specifically the trimeric 9-1-1 complex. In a genetic investigation of the fission yeast MutY homolog, myh1+, we show that the myh1 mutation confers a moderately increased UV sensitivity alone and in combination with mutations in several DNA repair genes. The myh1 rad1, and to a lesser degree myh1 rad9, double mutants display a synthetic interaction resulting in enhanced sensitivity to DNA damaging agents and hydroxyurea. UV irradiation of myh1 rad1 double mutants results in severe chromosome segregation defects and visible DNA fragmentation, and a failure to activate the checkpoint. Additionally, myh1 rad1 double mutants exhibit morphological defects in the absence of DNA damaging agents. We also found a moderate suppression of the slow growth and UV sensitivity of rhp51 mutants by the myh1 mutation. Our results implicate fission yeast Myh1 in repair of a wider range of DNA damage than previously thought, and functionally link it to the checkpoint pathway

  19. Variations in genes involved in immune response checkpoints and association with outcomes in patients with resected colorectal liver metastases.

    Science.gov (United States)

    Stremitzer, S; Sunakawa, Y; Zhang, W; Yang, D; Ning, Y; Stintzing, S; Sebio, A; Yamauchi, S; Matsusaka, S; El-Khoueiry, R; Stift, J; Wrba, F; Gruenberger, T; Lenz, H-J

    2015-12-01

    In patients with colorectal liver metastases (CLM), liver resection offers the possibility of cure and long-term survival. The liver is a highly immunogenic organ harboring ~80% of the body's tissue macrophages. Emerging data demonstrate a critical role of the immune response for cancer treatment. We investigated variations within genes involved in immune response checkpoints and their association with outcomes in patients with CLM who underwent neoadjuvant chemotherapy including bevacizumab and liver resection. Single-nucleotide polymorphisms (SNPs) in nine genes (CCL2, CCR2, LAG3, NT5E, PDCD1, CD274, IDO1, CTLA4 and CD24) were analyzed in genomic DNA from 149 patients with resected bevacizumab-pretreated CLM by direct Sanger DNA sequencing, and correlated with response, recurrence-free survival (RFS), overall survival (OS), probability of cure and recurrence patterns. IDO1 (indoleamine 2, 3-dioxygenase) rs3739319 G>A and CD24 rs8734 G>A showed a significant difference in 3-year OS rates. In addition, IDO1 rs3739319 G>A was significantly associated with extrahepatic recurrence. Recursive partitioning analyses revealed that IDO1 rs3739319 G>A was the dominant SNP predicting RFS and OS. Our data suggest that variants within genes involved in immune response checkpoints are associated with outcomes in patients with resected CLM and might lead to improved treatment strategies modulating anti-tumor immune response by targeting novel immune checkpoints. PMID:25752522

  20. Immune checkpoint blockade in cancer treatment: a double-edged sword cross-targeting the host as an "innocent bystander".

    Science.gov (United States)

    Gelao, Lucia; Criscitiello, Carmen; Esposito, Angela; Goldhirsch, Aron; Curigliano, Giuseppe

    2014-03-01

    Targeted immune checkpoint blockade augments anti-tumor immunity and induces durable responses in patients with melanoma and other solid tumors. It also induces specific "immune-related adverse events" (irAEs). IrAEs mainly include gastrointestinal, dermatological, hepatic and endocrinological toxicities. Off-target effects that arise appear to account for much of the toxicity of the immune checkpoint blockade. These unique "innocent bystander" effects are likely a direct result of breaking immune tolerance upon immune check point blockade and require specific treatment guidelines that include symptomatic therapies or systemic corticosteroids. What do we need going forward to limit immune checkpoint blockade-induced toxicity? Most importantly, we need a better understanding of the roles played by these agents in normal tissues, so that we can begin to predict potentially problematic side effects on the basis of their selectivity profile. Second, we need to focus on the predictive factors of the response and toxicity of the host rather than serially focusing on individual agents. Third, rigorous biomarker-driven clinical trials are needed to further elucidate the mechanisms of both the benefit and toxicity. We will summarize the double-edged sword effect of immunotherapeutics in cancer treatment. PMID:24594636

  1. Silencing of poly(ADP-ribose) glycohydrolase sensitizes lung cancer cells to radiation through the abrogation of DNA damage checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Nakadate, Yusuke [Shien-Lab, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Department of Bioengineering, Graduate School of Engineering, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); Kodera, Yasuo; Kitamura, Yuka [Shien-Lab, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Tachibana, Taro [Department of Bioengineering, Graduate School of Engineering, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); Tamura, Tomohide [Division of Thoracic Oncology, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Koizumi, Fumiaki, E-mail: fkoizumi@ncc.go.jp [Division of Thoracic Oncology, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan)

    2013-11-29

    Highlights: •Radiosensitization by PARG silencing was observed in multiple lung cancer cells. •PAR accumulation was enhanced by PARG silencing after DNA damage. •Radiation-induced G2/M arrest and checkpoint activation were impaired by PARG siRNA. -- Abstract: Poly(ADP-ribose) glycohydrolase (PARG) is a major enzyme that plays a role in the degradation of poly(ADP-ribose) (PAR). PARG deficiency reportedly sensitizes cells to the effects of radiation. In lung cancer, however, it has not been fully elucidated. Here, we investigated whether PARG siRNA contributes to an increased radiosensitivity using 8 lung cancer cell lines. Among them, the silencing of PARG induced a radiosensitizing effect in 5 cell lines. Radiation-induced G2/M arrest was largely suppressed by PARG siRNA in PC-14 and A427 cells, which exhibited significantly enhanced radiosensitivity in response to PARG knockdown. On the other hand, a similar effect was not observed in H520 cells, which did not exhibit a radiosensitizing effect. Consistent with a cell cycle analysis, radiation-induced checkpoint signals were not well activated in the PC-14 and A427 cells when treated with PARG siRNA. These results suggest that the increased sensitivity to radiation induced by PARG knockdown occurs through the abrogation of radiation-induced G2/M arrest and checkpoint activation in lung cancer cells. Our findings indicate that PARG could be a potential target for lung cancer treatments when used in combination with radiotherapy.

  2. Poly(ADP-ribose) binding to Chk1 at stalled replication forks is required for S-phase checkpoint activation

    Science.gov (United States)

    Min, Wookee; Bruhn, Christopher; Grigaravicius, Paulius; Zhou, Zhong-Wei; Li, Fu; Krüger, Anja; Siddeek, Bénazir; Greulich, Karl-Otto; Popp, Oliver; Meisezahl, Chris; Calkhoven, Cornelis F.; Bürkle, Alexander; Xu, Xingzhi; Wang, Zhao-Qi

    2013-12-01

    Damaged replication forks activate poly(ADP-ribose) polymerase 1 (PARP1), which catalyses poly(ADP-ribose) (PAR) formation; however, how PARP1 or poly(ADP-ribosyl)ation is involved in the S-phase checkpoint is unknown. Here we show that PAR, supplied by PARP1, interacts with Chk1 via a novel PAR-binding regulatory (PbR) motif in Chk1, independent of ATR and its activity. iPOND studies reveal that Chk1 associates readily with the unperturbed replication fork and that PAR is required for efficient retention of Chk1 and phosphorylated Chk1 at the fork. A PbR mutation, which disrupts PAR binding, but not the interaction with its partners Claspin or BRCA1, impairs Chk1 and the S-phase checkpoint activation, and mirrors Chk1 knockdown-induced hypersensitivity to fork poisoning. We find that long chains, but not short chains, of PAR stimulate Chk1 kinase activity. Collectively, we disclose a previously unrecognized mechanism of the S-phase checkpoint by PAR metabolism that modulates Chk1 activity at the replication fork.

  3. Cryo-EM of Mitotic Checkpoint Complex-Bound APC/C Reveals Reciprocal and Conformational Regulation of Ubiquitin Ligation.

    Science.gov (United States)

    Yamaguchi, Masaya; VanderLinden, Ryan; Weissmann, Florian; Qiao, Renping; Dube, Prakash; Brown, Nicholas G; Haselbach, David; Zhang, Wei; Sidhu, Sachdev S; Peters, Jan-Michael; Stark, Holger; Schulman, Brenda A

    2016-08-18

    The mitotic checkpoint complex (MCC) coordinates proper chromosome biorientation on the spindle with ubiquitination activities of CDC20-activated anaphase-promoting complex/cyclosome (APC/C(CDC20)). APC/C(CDC20) and two E2s, UBE2C and UBE2S, catalyze ubiquitination through distinct architectures for linking ubiquitin (UB) to substrates and elongating polyUB chains, respectively. MCC, which contains a second molecule of CDC20, blocks APC/C(CDC20)-UBE2C-dependent ubiquitination of Securin and Cyclins, while differentially determining or inhibiting CDC20 ubiquitination to regulate spindle surveillance, checkpoint activation, and checkpoint termination. Here electron microscopy reveals conformational variation of APC/C(CDC20)-MCC underlying this multifaceted regulation. MCC binds APC/C-bound CDC20 to inhibit substrate access. However, rotation about the CDC20-MCC assembly and conformational variability of APC/C modulate UBE2C-catalyzed ubiquitination of MCC's CDC20 molecule. Access of UBE2C is limiting for subsequent polyubiquitination by UBE2S. We propose that conformational dynamics of APC/C(CDC20)-MCC modulate E2 activation and determine distinctive ubiquitination activities as part of a response mechanism ensuring accurate sister chromatid segregation. PMID:27522463

  4. Fin1-PP1 Helps Clear Spindle Assembly Checkpoint Protein Bub1 from Kinetochores in Anaphase.

    Science.gov (United States)

    Bokros, Michael; Gravenmier, Curtis; Jin, Fengzhi; Richmond, Daniel; Wang, Yanchang

    2016-02-01

    The spindle assembly checkpoint (SAC) monitors chromosome attachment defects, and the assembly of SAC proteins at kinetochores is essential for its activation, but the SAC disassembly process remains unknown. We found that deletion of a 14-3-3 protein, Bmh1, or hyperactivation of Cdc14 early anaphase release (FEAR) allows premature SAC silencing in budding yeast, which depends on a kinetochore protein Fin1 that forms a complex with protein phosphatase PP1. Previous works suggest that FEAR-dependent Fin1 dephosphorylation promotes Bmh1-Fin1 dissociation, which enables kinetochore recruitment of Fin1-PP1. We found persistent kinetochore association of SAC protein Bub1 in fin1Δ mutants after anaphase entry. Therefore, we revealed a mechanism that clears SAC proteins from kinetochores. After anaphase entry, FEAR activation promotes kinetochore enrichment of Fin1-PP1, resulting in SAC disassembly at kinetochores. This mechanism is required for efficient SAC silencing after SAC is challenged, and untimely Fin1-kinetochore association causes premature SAC silencing and chromosome missegregation. PMID:26832405

  5. The flavonoid eupatorin inactivates the mitotic checkpoint leading to polyploidy and apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Salmela, Anna-Leena [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Graduate School of Biomedical Sciences, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Pouwels, Jeroen; Kukkonen-Macchi, Anu [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Waris, Sinikka; Toivonen, Pauliina [Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Jaakkola, Kimmo [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Maeki-Jouppila, Jenni [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Drug Discovery Graduate School, University of Turku (Finland); Kallio, Lila, E-mail: lila.kallio@vtt.fi [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Kallio, Marko J. [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Centre of Excellence for Translational Genome-Scale Biology, P.O. Box 106, Academy of Finland (Finland)

    2012-03-10

    The spindle assembly checkpoint (SAC) is a conserved mechanism that ensures the fidelity of chromosome distribution in mitosis by preventing anaphase onset until the correct bipolar microtubule-kinetochore attachments are formed. Errors in SAC function may contribute to tumorigenesis by inducing numerical chromosome anomalies (aneuploidy). On the other hand, total disruption of SAC can lead to massive genomic imbalance followed by cell death, a phenomena that has therapeutic potency. We performed a cell-based high-throughput screen with a compound library of 2000 bioactives for novel SAC inhibitors and discovered a plant-derived phenolic compound eupatorin (3 Prime ,5-dihydroxy-4 Prime ,6,7-trimethoxyflavone) as an anti-mitotic flavonoid. The premature override of the microtubule drug-imposed mitotic arrest by eupatorin is dependent on microtubule-kinetochore attachments but not interkinetochore tension. Aurora B kinase activity, which is essential for maintenance of normal SAC signaling, is diminished by eupatorin in cells and in vitro providing a mechanistic explanation for the observed forced mitotic exit. Eupatorin likely has additional targets since eupatorin treatment of pre-mitotic cells causes spindle anomalies triggering a transient M phase delay followed by impaired cytokinesis and polyploidy. Finally, eupatorin potently induces apoptosis in multiple cancer cell lines and suppresses cancer cell proliferation in organotypic 3D cell culture model.

  6. BRCA1 and its phosphorylation involved in caffeine-inhibitable event upstream of G2 checkpoint

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Caffeine,which specifically inhibits ATM/ATR kinases,efficiently abrogates the ionizing radiation(IR)-induced G2 arrest and increases the sensitivity of various tumor cells to IR.Mechanisms for the effect of caffeine remain to be elucidated.As a target of ATM/ATR kinases,BRCA1 becomes activated and phosphorylated in response to IR.Thus,in this work,we investigated the possible role of BRCA1 in the effect of caffeine on G2 checkpoint and observed how BRCA1 phosphorylation was regulated in this process.For these purposes,the BRCA1 protein level and the phosphorylation states were analyzed by Western blotting by using an antibody against BRCA1 and phospho-specific antibodies against Ser-1423 and Ser-1524 residues in cells exposed to a combination of IR and caffeine.The results showed that caffeine down-regulated IR-induced BRCA1 expression and specifically abolished BRCA1 phosphorylation of Ser-1524,which was followed by an override of G2 arrest by caffeine.In addition,the ability of BRCA1 to transactivate p21 may be required for MCF-7 but not necessary for Hela response to caffeine.These data suggest that BRCA1 may be a potential target of caffeine.BRCA1 and its phosphorylation are most likely to be involved in the caffeine-inhibitable event upstream of G2 arrest.

  7. Inhibition of checkpoint kinase 1 sensitizes lung cancer brain metastases to radiotherapy

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Heekyoung [Department of Neurosurgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); Samsung Biomedical Research Institute, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); Cancer Stem Cell Research Center, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); Yoon, Su Jin [Samsung Biomedical Research Institute, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); Jin, Juyoun [Department of Neurosurgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); Samsung Biomedical Research Institute, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); Cancer Stem Cell Research Center, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); Choi, Seung Ho [Samsung Biomedical Research Institute, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); Seol, Ho Jun; Lee, Jung-Il [Department of Neurosurgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul 135-710 (Korea, Republic of); and others

    2011-03-04

    Research highlights: {yields} The most important therapeutic tool in brain metastasis is radiation therapy. {yields} Radiosensitivity of cancer cells was enhanced with treatment of Chk1 inhibitor. {yields} Depletion of Chk1 in cancer cells showed an enhancement of sensitivity to radiation. {yields} Chk1 can be a good target for enhancement of radiosensitivity. -- Abstract: The most important therapeutic tool in brain metastasis is radiation therapy. However, resistance to radiation is a possible cause of recurrence or treatment failure. Recently, signal pathways about DNA damage checkpoints after irradiation have been noticed. We investigated the radiosensitivity can be enhanced with treatment of Chk1 inhibitor, AZD7762 in lung cancer cell lines and xenograft models of lung cancer brain metastasis. Clonogenic survival assays showed enhancement of radiosensitivity with AZD7762 after irradiation of various doses. AZD7762 increased ATR/ATM-mediated Chk1 phosphorylation and stabilized Cdc25A, suppressed cyclin A expression in lung cancer cell lines. In xenograft models of lung cancer (PC14PE6) brain metastasis, AZD7762 significantly prolonged the median survival time in response to radiation. Depletion of Chk1 using shRNA also showed an enhancement of sensitivity to radiation in PC14PE6 cells. The results of this study support that Chk1 can be a good target for enhancement of radiosensitivity.

  8. Stable kinetochore–microtubule attachment is sufficient to silence the spindle assembly checkpoint in human cells

    Science.gov (United States)

    Tauchman, Eric C.; Boehm, Frederick J.; DeLuca, Jennifer G.

    2015-01-01

    During mitosis, duplicated sister chromatids attach to microtubules emanating from opposing sides of the bipolar spindle through large protein complexes called kinetochores. In the absence of stable kinetochore–microtubule attachments, a cell surveillance mechanism known as the spindle assembly checkpoint (SAC) produces an inhibitory signal that prevents anaphase onset. Precisely how the inhibitory SAC signal is extinguished in response to microtubule attachment remains unresolved. To address this, we induced formation of hyper-stable kinetochore–microtubule attachments in human cells using a non-phosphorylatable version of the protein Hec1, a core component of the attachment machinery. We find that stable attachments are sufficient to silence the SAC in the absence of sister kinetochore bi-orientation and strikingly in the absence of detectable microtubule pulling forces or tension. Furthermore, we find that SAC satisfaction occurs despite the absence of large changes in intra-kinetochore distance, suggesting that substantial kinetochore stretching is not required for quenching the SAC signal. PMID:26620470

  9. The flavonoid eupatorin inactivates the mitotic checkpoint leading to polyploidy and apoptosis

    International Nuclear Information System (INIS)

    The spindle assembly checkpoint (SAC) is a conserved mechanism that ensures the fidelity of chromosome distribution in mitosis by preventing anaphase onset until the correct bipolar microtubule–kinetochore attachments are formed. Errors in SAC function may contribute to tumorigenesis by inducing numerical chromosome anomalies (aneuploidy). On the other hand, total disruption of SAC can lead to massive genomic imbalance followed by cell death, a phenomena that has therapeutic potency. We performed a cell-based high-throughput screen with a compound library of 2000 bioactives for novel SAC inhibitors and discovered a plant-derived phenolic compound eupatorin (3′,5-dihydroxy-4′,6,7-trimethoxyflavone) as an anti-mitotic flavonoid. The premature override of the microtubule drug-imposed mitotic arrest by eupatorin is dependent on microtubule–kinetochore attachments but not interkinetochore tension. Aurora B kinase activity, which is essential for maintenance of normal SAC signaling, is diminished by eupatorin in cells and in vitro providing a mechanistic explanation for the observed forced mitotic exit. Eupatorin likely has additional targets since eupatorin treatment of pre-mitotic cells causes spindle anomalies triggering a transient M phase delay followed by impaired cytokinesis and polyploidy. Finally, eupatorin potently induces apoptosis in multiple cancer cell lines and suppresses cancer cell proliferation in organotypic 3D cell culture model.

  10. Checkpoint-Inhibitoren in der Immuntherapie: Ein Meilenstein in der Behandlung des malignen Melanoms.

    Science.gov (United States)

    Wilden, Sophia M; Lang, Berenice M; Mohr, Peter; Grabbe, Stephan

    2016-07-01

    Seit Jahrzehnten ist bekannt, dass Tumoren vom Immunsystem erkannt und zerstört werden können. Diese, vor allem in Tierversuchen gewonnene Erkenntnis konnte jedoch in der Vergangenheit nicht zum Nutzen unserer Patienten umgesetzt werden, da immunonkologische Therapieansätze in den letzten Jahrzehnten in der Anwendung beim Menschen stets versagt haben. Daher hat, mit Ausnahme der adjuvanten Interferontherapie, keines dieser Verfahren den Einzug in die klinische Versorgung gefunden. Langzeitüberleben unter guter Lebensqualität war dabei sehr wenigen Patienten vorbehalten. Mit den neuen immunologischen Therapieansätzen wird jedoch sowohl das Langzeitüberleben als auch die Lebensqualität onkologischer Patienten neu definiert. Auf die neuen "Immun-Checkpoint-Inhibitoren" spricht erstmals ein relevanter Teil der behandelten Patienten an und diese zeigen in der Regel langandauernde Remissionen bis hin zur Heilung. Schon jetzt ist klar, dass die Immuntherapie in Zukunft eine der wesentlichen Therapiesäulen bei der Behandlung des metastasierten Melanoms und auch vieler anderer fortgeschrittener Tumoren bilden wird. In dieser Übersicht werden die wichtigsten neuen Therapiemodalitäten besprochen und sowohl deren Wirkprinzip als auch klinische Daten zum Therapieansprechen und zu erwartenden Nebenwirkungen der Therapie referiert. PMID:27373243

  11. The Spindle Assembly Checkpoint Safeguards Genomic Integrity of Skeletal Muscle Satellite Cells

    Directory of Open Access Journals (Sweden)

    Swapna Kollu

    2015-06-01

    Full Text Available To ensure accurate genomic segregation, cells evolved the spindle assembly checkpoint (SAC, whose role in adult stem cells remains unknown. Inducible perturbation of a SAC kinase, Mps1, and its downstream effector, Mad2, in skeletal muscle stem cells shows the SAC to be critical for normal muscle growth, repair, and self-renewal of the stem cell pool. SAC-deficient muscle stem cells arrest in G1 phase of the cell cycle with elevated aneuploidy, resisting differentiation even under inductive conditions. p21CIP1 is responsible for these SAC-deficient phenotypes. Despite aneuploidy’s correlation with aging, we find that aged proliferating muscle stem cells display robust SAC activity without elevated aneuploidy. Thus, muscle stem cells have a two-step mechanism to safeguard their genomic integrity. The SAC prevents chromosome missegregation and, if it fails, p21CIP1-dependent G1 arrest limits cellular propagation and tissue integration. These mechanisms ensure that muscle stem cells with compromised genomes do not contribute to tissue homeostasis.

  12. Optimising measles virus-guided radiovirotherapy with external beam radiotherapy and specific checkpoint kinase 1 inhibition

    International Nuclear Information System (INIS)

    Background and purpose: We previously reported a therapeutic strategy comprising replication-defective NIS-expressing adenovirus combined with radioiodide, external beam radiotherapy (EBRT) and DNA repair inhibition. We have now evaluated NIS-expressing oncolytic measles virus (MV-NIS) combined with NIS-guided radioiodide, EBRT and specific checkpoint kinase 1 (Chk1) inhibition in head and neck and colorectal models. Materials and methods: Anti-proliferative/cytotoxic effects of individual agents and their combinations were measured by MTS, clonogenic and Western analysis. Viral gene expression was measured by radioisotope uptake and replication by one-step growth curves. Potential synergistic interactions were tested in vitro by Bliss independence analysis and in in vivo therapeutic studies. Results: EBRT and MV-NIS were synergistic in vitro. Furthermore, EBRT increased NIS expression in infected cells. SAR-020106 was synergistic with EBRT, but also with MV-NIS in HN5 cells. MV-NIS mediated 131I-induced cytotoxicity in HN5 and HCT116 cells and, in the latter, this was enhanced by SAR-020106. In vivo studies confirmed that MV-NIS, EBRT and Chk1 inhibition were effective in HCT116 xenografts. The quadruplet regimen of MV-NIS, virally-directed 131I, EBRT and SAR-020106 had significant anti-tumour activity in HCT116 xenografts. Conclusion: This study strongly supports translational and clinical research on MV-NIS combined with radiation therapy and radiosensitising agents

  13. Recognition of a Single Transmembrane Degron by Sequential Quality Control Checkpoints

    Science.gov (United States)

    Fayadat, Laurence; Kopito, Ron R.

    2003-01-01

    To understand the relationship between conformational maturation and quality control–mediated proteolysis in the secretory pathway, we engineered the well-characterized degron from the α-subunit of the T-cell antigen receptor (TCRα) into the α-helical transmembrane domain of homotrimeric type I integral membrane protein, influenza hemagglutinin (HA). Although the membrane degron does not appear to interfere with acquisition of native secondary structure, as assessed by the formation of native intrachain disulfide bonds, only ∼50% of nascent mutant HA chains (HA++) become membrane-integrated and acquire complex N-linked glycans indicative of transit to a post-ER compartment. The remaining ∼50% of nascent HA++ chains fail to integrate into the lipid bilayer and are subject to proteasome-dependent degradation. Site-specific cleavage by extracellular trypsin and reactivity with conformation-specific monoclonal antibodies indicate that membrane-integrated HA++ molecules are able to mature to the plasma membrane with a conformation indistinguishable from that of HAwt. These apparently native HA++ molecules are, nevertheless, rapidly degraded by a process that is insensitive to proteasome inhibitors but blocked by lysosomotropic amines. These data suggest the existence in the secretory pathway of at least two sequential quality control checkpoints that recognize the same transmembrane degron, thereby ensuring the fidelity of protein deployment to the plasma membrane. PMID:12631739

  14. BRCA1 and its phosphorylation involved in caffeine-inhibitable event upstream of G2 checkpoint

    Science.gov (United States)

    Li, Ning; Zhang, Hong; Wang, Yanling; Hao, Jifang

    2010-07-01

    Caffeine, which specifically inhibits ATM/ATR kinases, efficiently abrogates the ionizing radiation (IR)-induced G2 arrest and increases the sensitivity of various tumor cells to IR. Mechanisms for the effect of caffeine remain to be elucidated. As a target of ATM/ATR kinases, BRCA1 becomes activated and phosphorylated in response to IR. Thus, in this work, we investigated the possible role of BRCA1 in the effect of caffeine on G2 checkpoint and observed how BRCA1 phosphorylation was regulated in this process. For these purposes, the BRCA1 protein level and the phosphorylation states were analyzed by Western blotting by using an antibody against BRCA1 and phospho-specific antibodies against Ser-1423 and Ser-1524 residues in cells exposed to a combination of IR and caffeine. The results showed that caffeine down-regulated IR-induced BRCA1 expression and specifically abolished BRCA1 phosphorylation of Ser-1524, which was followed by an override of G2 arrest by caffeine. In addition, the ability of BRCA1 to transactivate p21 may be required for MCF-7 but not necessary for Hela response to caffeine. These data suggest that BRCA1 may be a potential target of caffeine. BRCA1 and its phosphorylation are most likely to be involved in the caffeine-inhibitable event upstream of G2 arrest.

  15. Morphogenesis checkpoint kinase Swe1 is the executor of lipolysis-dependent cell-cycle progression.

    Science.gov (United States)

    Chauhan, Neha; Visram, Myriam; Cristobal-Sarramian, Alvaro; Sarkleti, Florian; Kohlwein, Sepp D

    2015-03-10

    Cell growth and division requires the precise duplication of cellular DNA content but also of membranes and organelles. Knowledge about the cell-cycle-dependent regulation of membrane and storage lipid homeostasis is only rudimentary. Previous work from our laboratory has shown that the breakdown of triacylglycerols (TGs) is regulated in a cell-cycle-dependent manner, by activation of the Tgl4 lipase by the major cyclin-dependent kinase Cdc28. The lipases Tgl3 and Tgl4 are required for efficient cell-cycle progression during the G1/S (Gap1/replication phase) transition, at the onset of bud formation, and their absence leads to a cell-cycle delay. We now show that defective lipolysis activates the Swe1 morphogenesis checkpoint kinase that halts cell-cycle progression by phosphorylation of Cdc28 at tyrosine residue 19. Saturated long-chain fatty acids and phytosphingosine supplementation rescue the cell-cycle delay in the Tgl3/Tgl4 lipase-deficient strain, suggesting that Swe1 activity responds to imbalanced sphingolipid metabolism, in the absence of TG degradation. We propose a model by which TG-derived sphingolipids are required to activate the protein phosphatase 2A (PP2A(Cdc55)) to attenuate Swe1 phosphorylation and its inhibitory effect on Cdc28 at the G1/S transition of the cell cycle. PMID:25713391

  16. Dictyostelium nucleomorphin is a member of the BRCT-domain family of cell cycle checkpoint proteins.

    Science.gov (United States)

    Myre, Michael A; O'Day, Danton H

    2004-11-18

    A search of the Dictyostelium genome project database (http://dictybase.org/db/cgi-bin/blast.pl) with nucleomorphin, a protein that regulates the nuclear number, predicted it to be encoded by a larger gene containing a putative breast cancer carboxy-terminus domain (BRCT). Using RT-PCR, Northern and Western blotting we have identified a differentially expressed, 2318 bp cDNA encoding a protein isoform of Dictyostelium NumA with an apparent molecular weight of 70 kDa that we have called NumB. It contains a single amino-terminal BRCT-domain spanning residues 125-201. Starvation of shaking cultures reduces NumA expression by approximately 88+/-5.6%, whereas NumB expression increases approximately 35+/-3.5% from vegetative levels. NumC, a third isoform that is also expressed during development but not growth, remains to be characterized. These findings suggest NumB may be a member of the BRCT-domain containing cell cycle checkpoint proteins. PMID:15535983

  17. Inhibition of checkpoint kinase 1 sensitizes lung cancer brain metastases to radiotherapy

    International Nuclear Information System (INIS)

    Research highlights: → The most important therapeutic tool in brain metastasis is radiation therapy. → Radiosensitivity of cancer cells was enhanced with treatment of Chk1 inhibitor. → Depletion of Chk1 in cancer cells showed an enhancement of sensitivity to radiation. → Chk1 can be a good target for enhancement of radiosensitivity. -- Abstract: The most important therapeutic tool in brain metastasis is radiation therapy. However, resistance to radiation is a possible cause of recurrence or treatment failure. Recently, signal pathways about DNA damage checkpoints after irradiation have been noticed. We investigated the radiosensitivity can be enhanced with treatment of Chk1 inhibitor, AZD7762 in lung cancer cell lines and xenograft models of lung cancer brain metastasis. Clonogenic survival assays showed enhancement of radiosensitivity with AZD7762 after irradiation of various doses. AZD7762 increased ATR/ATM-mediated Chk1 phosphorylation and stabilized Cdc25A, suppressed cyclin A expression in lung cancer cell lines. In xenograft models of lung cancer (PC14PE6) brain metastasis, AZD7762 significantly prolonged the median survival time in response to radiation. Depletion of Chk1 using shRNA also showed an enhancement of sensitivity to radiation in PC14PE6 cells. The results of this study support that Chk1 can be a good target for enhancement of radiosensitivity.

  18. DNA damage response and spindle assembly checkpoint function throughout the cell cycle to ensure genomic integrity.

    Directory of Open Access Journals (Sweden)

    Katherine S Lawrence

    2015-04-01

    Full Text Available Errors in replication or segregation lead to DNA damage, mutations, and aneuploidies. Consequently, cells monitor these events and delay progression through the cell cycle so repair precedes division. The DNA damage response (DDR, which monitors DNA integrity, and the spindle assembly checkpoint (SAC, which responds to defects in spindle attachment/tension during metaphase of mitosis and meiosis, are critical for preventing genome instability. Here we show that the DDR and SAC function together throughout the cell cycle to ensure genome integrity in C. elegans germ cells. Metaphase defects result in enrichment of SAC and DDR components to chromatin, and both SAC and DDR are required for metaphase delays. During persistent metaphase arrest following establishment of bi-oriented chromosomes, stability of the metaphase plate is compromised in the absence of DDR kinases ATR or CHK1 or SAC components, MAD1/MAD2, suggesting SAC functions in metaphase beyond its interactions with APC activator CDC20. In response to DNA damage, MAD2 and the histone variant CENPA become enriched at the nuclear periphery in a DDR-dependent manner. Further, depletion of either MAD1 or CENPA results in loss of peripherally associated damaged DNA. In contrast to a SAC-insensitive CDC20 mutant, germ cells deficient for SAC or CENPA cannot efficiently repair DNA damage, suggesting that SAC mediates DNA repair through CENPA interactions with the nuclear periphery. We also show that replication perturbations result in relocalization of MAD1/MAD2 in human cells, suggesting that the role of SAC in DNA repair is conserved.

  19. DNA damage checkpoint kinase ATM regulates germination and maintains genome stability in seeds.

    Science.gov (United States)

    Waterworth, Wanda M; Footitt, Steven; Bray, Clifford M; Finch-Savage, William E; West, Christopher E

    2016-08-23

    Genome integrity is crucial for cellular survival and the faithful transmission of genetic information. The eukaryotic cellular response to DNA damage is orchestrated by the DNA damage checkpoint kinases ATAXIA TELANGIECTASIA MUTATED (ATM) and ATM AND RAD3-RELATED (ATR). Here we identify important physiological roles for these sensor kinases in control of seed germination. We demonstrate that double-strand breaks (DSBs) are rate-limiting for germination. We identify that desiccation tolerant seeds exhibit a striking transcriptional DSB damage response during germination, indicative of high levels of genotoxic stress, which is induced following maturation drying and quiescence. Mutant atr and atm seeds are highly resistant to aging, establishing ATM and ATR as determinants of seed viability. In response to aging, ATM delays germination, whereas atm mutant seeds germinate with extensive chromosomal abnormalities. This identifies ATM as a major factor that controls germination in aged seeds, integrating progression through germination with surveillance of genome integrity. Mechanistically, ATM functions through control of DNA replication in imbibing seeds. ATM signaling is mediated by transcriptional control of the cell cycle inhibitor SIAMESE-RELATED 5, an essential factor required for the aging-induced delay to germination. In the soil seed bank, seeds exhibit increased transcript levels of ATM and ATR, with changes in dormancy and germination potential modulated by environmental signals, including temperature and soil moisture. Collectively, our findings reveal physiological functions for these sensor kinases in linking genome integrity to germination, thereby influencing seed quality, crucial for plant survival in the natural environment and sustainable crop production. PMID:27503884

  20. Immune checkpoint blockade with concurrent electrochemotherapy in advanced melanoma: a retrospective multicenter analysis.

    Science.gov (United States)

    Heppt, Markus V; Eigentler, Thomas K; Kähler, Katharina C; Herbst, Rudolf A; Göppner, Daniela; Gambichler, Thilo; Ulrich, Jens; Dippel, Edgar; Loquai, Carmen; Schell, Beatrice; Schilling, Bastian; Schäd, Susanne G; Schultz, Erwin S; Matheis, Fanny; Tietze, Julia K; Berking, Carola

    2016-08-01

    Growing evidence suggests that concurrent loco-regional and systemic treatment modalities may lead to synergistic anti-tumor effects in advanced melanoma. In this retrospective multicenter study, we evaluate the use of electrochemotherapy (ECT) combined with ipilimumab or PD-1 inhibition. We investigated patients with unresectable or metastatic melanoma who received the combination of ECT and immune checkpoint blockade for distant or cutaneous metastases within 4 weeks. Clinical and laboratory data were collected and analyzed with respect to safety and efficacy. A total of 33 patients from 13 centers were identified with a median follow-up time of 9 months. Twenty-eight patients received ipilimumab, while five patients were treated with a PD-1 inhibitor (pembrolizumab n = 3, nivolumab n = 2). The local overall response rate (ORR) was 66.7 %. The systemic ORR was 19.2 and 40.0 % in the ipilimumab and PD-1 cohort, respectively. The median duration of response was not reached in either group. The median time to disease progression was 2.5 months for the entire population with 2 months for ipilimumab and 5 months for PD-1 blockade. The median overall survival was not reached in patients with ipilimumab and 15 months in the PD-1 group. Severe systemic adverse events were detected in 25.0 % in the ipilimumab group. No treatment-related deaths were observed. This is the first reported evaluation of ECT and simultaneous PD-1 inhibition and the largest published dataset on ECT with concurrent ipilimumab. The local response was lower than reported for ECT only. Ipilimumab combined with ECT was feasible, tolerable and showed a high systemic response rate. PMID:27294607

  1. Multiple Defects of Cell Cycle Checkpoints in U937-ASPI3K, an U937 Cell Mutant Stably Expressing Anti-Sense ATM Gene cDNA

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    (Ataxia-telangiectasia mutated gene (ATM) functions in control of cell cycle checkpoints in responding to DNA damage and protects cells from undergoing apoptosis. Knock-out within tumor cells of endogenous ATM will achieve therapeutic benefits and nable a better understanding of the decisive mechanisms of cell death or survival in response to DNA damaging agents. ) In present paper, we sought to characterize the cell cycle checkpoint profiles in U937-ASPI3K, a U937 cell mutant that was previously established with endogenous ATM knock-out phenotype. Synchronized U937-ASPI3K was exposed to 137Cs irradiation, G1, S, G2/M cell cycle checkpoint profiles were evaluated by determining cell cycle kinetics, p53/p21 protein, cyclin dependent kinase 2 (CDK2) and p34CDC2 kinase activity in response to irradiation. U937-ASPI3K exhibited multiple defects in cell cycle checkpoints as defined by failing to arrest cells upon irradiation. The accumulation of cellular p53/p21 protein and inhibition of CDK kinase was also abolished in U937-ASPI3K. It was concluded that the stable expression of anti-sense PI3K cDNA fragment completely abolished multiple cell cycle checkpoints in U937-ASPI3K, and hence U937-ASPI3K with an AT-like phenotype could serves as a valuable model system for investigating the signal transduction pathway in responding to DNA damaging-based cancer therapy.

  2. Methylations of histone H3 lysine 9 and lysine 36 are functionally linked to DNA replication checkpoint control in fission yeast

    International Nuclear Information System (INIS)

    Recently, histone H4 lysine 20 and H3 lysine 79 methylations were functionally linked to DNA damage checkpoint. The crosstalk between histone methylation and the S-M checkpoint, however, has remained unclear. Here, we show that H3 lysine 9 (K9) and lysine 36 (K36) methylations catalyzed by two histone methyltransferases Clr4 and Set2 are involved in hydroxyurea (HU)-induced replication checkpoint. The clr4-set2 double mutants besides histone H3-K9 and K36 double mutants exhibited HU-sensitivity, a defective HU-induced S-M checkpoint, and a significant reduction of HU-induced phosphorylation of Cdc2. Intriguingly, the clr4-set2 double mutations impaired the HU-induced accumulation of a mitotic inhibitor Mik1. Double mutants in Alp13 and Swi6, which can specifically bind to H3-K36 and K9 methylations, exhibited phenotypes similar to those of the clr4-set2 mutants. Together, these findings suggest that methylations of histone H3-K9 and K36 by Clr4 and Set2 are functionally linked to DNA replication checkpoint via accumulation of Mik1

  3. Pch2 acts through Xrs2 and Tel1/ATM to modulate interhomolog bias and checkpoint function during meiosis.

    Directory of Open Access Journals (Sweden)

    Hsuan-Chung Ho

    2011-11-01

    Full Text Available Proper segregation of chromosomes during meiosis requires the formation and repair of double-strand breaks (DSBs to form crossovers. Repair is biased toward using the homolog as a substrate rather than the sister chromatid. Pch2 is a conserved member of the AAA(+-ATPase family of proteins and is implicated in a wide range of meiosis-specific processes including the recombination checkpoint, maturation of the chromosome axis, crossover control, and synapsis. We demonstrate a role for Pch2 in promoting and regulating interhomolog bias and the meiotic recombination checkpoint in response to unprocessed DSBs through the activation of axial proteins Hop1 and Mek1 in budding yeast. We show that Pch2 physically interacts with the putative BRCT repeats in the N-terminal region of Xrs2, a member of the MRX complex that acts at sites of unprocessed DSBs. Pch2, Xrs2, and the ATM ortholog Tel1 function in the same pathway leading to the phosphorylation of Hop1, independent of Rad17 and the ATR ortholog Mec1, which respond to the presence of single-stranded DNA. An N-terminal deletion of Xrs2 recapitulates the pch2Δ phenotypes for signaling unresected breaks. We propose that interaction with Xrs2 may enable Pch2 to remodel chromosome structure adjacent to the site of a DSB and thereby promote accessibility of Hop1 to the Tel1 kinase. In addition, Xrs2, like Pch2, is required for checkpoint-mediated delay conferred by the failure to synapse chromosomes.

  4. Mek1 stabilizes Hop1-Thr318 phosphorylation to promote interhomolog recombination and checkpoint responses during yeast meiosis.

    Science.gov (United States)

    Chuang, Chi-Ning; Cheng, Yun-Hsin; Wang, Ting-Fang

    2012-12-01

    Red1, Hop1 and Mek1 are three yeast meiosis-specific chromosomal proteins that uphold the interhomolog (IH) bias of meiotic recombination. Mek1 is also an effector protein kinase in a checkpoint that responds to aberrant DNA and/or axis structure. The activation of Mek1 requires Red1-dependent Hop1-Thr(T)318 phosphorylation, which is mediated by Mec1 and Tel1, the yeast homologs of the mammalian DNA damage sensor kinases ATR and ATM. As the ectopic expression of Mek1-glutathione S-transferase (GST) was shown to promote IH recombination in the absence of Mec1/Tel1-dependent checkpoint function, it was proposed that Mek1 might play dual roles during meiosis by directly phosphorylating targets that are involved in the recombination checkpoint. Here, we report that Mek1 has a positive feedback activity in the stabilization of Mec1/Tel1-mediated Hop1-T318 phosphorylation against the dephosphorylation mediated by protein phosphatase 4. Our results also reveal that GST-Mek1 or Mek1-GST further increases Hop1-T318 phosphorylation. This positive feedback function of Mek1 is independent of Mek1's kinase activity, but dependent on Mek1's forkhead-associated (FHA) domain and its arginine 51 residue. Arginine 51 directly mediates the interaction of Mek1-FHA and phosphorylated Hop1-T318. We suggest that the Hop1-Mek1 interaction is similar to the Rad53-Dun1 signaling pathway, which is mediated through the interaction of phosphorylated Rad53 and Dun1-FHA. PMID:23047948

  5. DNA-damage response network at the crossroads of cell-cycle checkpoints,cellular senescence and apoptosis

    Institute of Scientific and Technical Information of China (English)

    SCHMITT Estelle; PAQUET Claudie; BEAUCHEMIN Myriam; BERTRAND Richard

    2007-01-01

    Tissue homeostasis requires a carefully-orchestrated balance between cell proliferation,cellular senescence and cell death.Cells proliferate through a cell cycle that is tightly regulated by cyclin-dependent kinase activities.Cellular senescence is a safeguard program limiting the proliferative competence of cells in living organisms.Apoptosis eliminates unwanted cells by the coordinated activity of gene products that regulate and effect cell death.The intimate link between the cell cycle,cellular senescence,apoptosis regulation,cancer development and tumor responses to cancer treatment has become eminently apparent.Extensive research on tumor suppressor genes,oncogenes,the cell cycle and apoptosis regulatory genes has revealed how the DNA damage-sensing and -signaling pathways,referred to as the DNA-damage response network,are tied to cell proliferation,cell-cycle arrest,cellular senescence and apoptosis.DNA-damage responses are complex,involving "sensor" proteins that sense the damage,and transmit signals to "transducer" proteins,which,in turn,convey the signals to numerous "effector" proteins implicated in specific cellular pathways,including DNA repair mechanisms,cell-cycle checkpoints,cellular senescence and apoptosis.The Bcl-2 family of proteins stands among the most crucial regulators of apoptosis and performs vital functions in deciding whether a cell will live or die after cancer chemotherapy and irradiation.In addition,several studies have now revealed that members of the Bcl-2 family also interface with the cell cycle,DNA repair/recombination and cellular senescence,effects that are generally distinct from their function in apoptosis.In this review,we report progress in understanding the molecular networks that regulate cell-cycle checkpoints,cellular senescence and apoptosis after DNA damage,and discuss the influence of some Bcl-2 family members on cell-cycle checkpoint regulation.

  6. Structural insights into recognition of MDC1 by TopBP1 in DNA replication checkpoint control

    OpenAIRE

    Leung, Charles Chung Yun; Sun, Luxin; Gong, Zihua; Burkat, Michael; Edwards, Ross; Assmus, Mark; Chen, Junjie; Glover, J N Mark

    2013-01-01

    Activation of the DNA replication checkpoint by the ATR kinase requires protein interactions mediated by the ATR activating protein, TopBP1. Accumulation of TopBP1 at stalled replication forks requires the interaction of TopBP1 BRCT5 with the phosphorylated SDT repeats of the adaptor protein MDC1. Here we present the X-ray crystal structures of the tandem BRCT4/5 domains of TopBP1 free and in complex with a MDC1 consensus pSDpT phospho-peptide. TopBP1 BRCT4/5 adopts a variant BRCT-BRCT packin...

  7. PD-L1 Regulates a Critical Checkpoint for Autoimmune Myocarditis and Pneumonitis in MRL mice1

    OpenAIRE

    Lucas, Julie A.; Menke, Julia; Rabacal, Whitney A.; Schoen, Frederick J.; Sharpe, Arlene H.; Kelley, Vicki R.

    2008-01-01

    MRL/MpJ-Faslpr (MRL-Faslpr) mice develop a spontaneous T cell and macrophage (Mø) dependent autoimmune disease that shares features with human lupus. Interactions via the PD-1/PD-L1 pathway down-regulate immune responses and provide a negative regulatory checkpoint in mediating tolerance and autoimmune disease. Therefore, we tested the hypothesis that the PD-1/PD-L1 pathway suppresses lupus nephritis and the systemic illness in MRL-Faslpr mice. For this purpose, we compared kidney and systemi...

  8. Mlp1 Acts as a Mitotic Scaffold to Spatially Regulate Spindle Assembly Checkpoint Proteins in Aspergillus nidulans

    OpenAIRE

    De Souza, Colin P.; Hashmi, Shahr B.; Nayak, Tania; Oakley, Berl; Osmani, Stephen A.

    2009-01-01

    During open mitosis several nuclear pore complex (NPC) proteins have mitotic specific localizations and functions. We find that the Aspergillus nidulans Mlp1 NPC protein has previously unrealized mitotic roles involving spatial regulation of spindle assembly checkpoint (SAC) proteins. In interphase, An-Mlp1 tethers the An-Mad1 and An-Mad2 SAC proteins to NPCs. During a normal mitosis, An-Mlp1, An-Mad1, and An-Mad2 localize similarly on, and around, kinetochores until telophase when they trans...

  9. Effect of Spindle Checkpoint on Akt2-mediated Paclitaxel-resistance in A2780 Ovarian Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    周婷; 鲍引娣; 叶双梅; 翁丹卉; 陈刚; 卢运萍; 马丁; 王世宣

    2010-01-01

    Recent evidence has suggested that Akt2 plays an important role in the protection of cells from paclitaxel(PTX)-induced apoptosis and control of the cell cycle.In addition,some scholars suggested that the PTX sensitivity depends on a functional spindle assembly checkpoint.In the present study,we investigated the role of the Akt2/Bub1 cross-talking in apoptosis and cell cycle after exposure of the A2780 ovarian cancer cells to paclitaxel(PTX).Recombinant expression plasmid WT-Akt2 was transfected into A2780 ...

  10. X-ray structures of checkpoint kinase 2 in complex with inhibitors that target its gatekeeper-dependent hydrophobic pocket

    Energy Technology Data Exchange (ETDEWEB)

    Lountos, George T.; Jobson, Andrew G.; Tropea, Joseph E.; Self, Christopher R.; Zhang, Guangtao; Pommier, Yves; Shoemaker, Robert H.; Waugh, David S. (Provid); (NIH); (SAIC); (NCI)

    2012-09-17

    The serine/threonine checkpoint kinase 2 (Chk2) is an attractive molecular target for the development of small molecule inhibitors to treat cancer. Here, we report the rational design of Chk2 inhibitors that target the gatekeeper-dependent hydrophobic pocket located behind the adenine-binding region of the ATP-binding site. These compounds exhibit IC{sub 50} values in the low nanomolar range and are highly selective for Chk2 over Chk1. X-ray crystallography was used to determine the structures of the inhibitors in complex with the catalytic kinase domain of Chk2 to verify their modes of binding.

  11. Checkpoint inhibitors in cancer immunotherapy: Cross reactivity of a CTLA-4 antibody and IDO-inhibitor L-1MT in pigs

    DEFF Research Database (Denmark)

    Al-Shatrawi, Zina Adil; Frøsig, Thomas Mørch; Jungersen, Gregers

    Blockade of checkpoint inhibitors has recently shown very convincing results in the treatment of cancer. One key target is CTLA-4, which has been demonstrated to be a potent negative regulator of lymphocyte activation. The treatment with the FDA-approved fully human CTLA-4 monoclonal antibody...... a non-specific activation of porcine T cells. This will be further investigated to provide the basis for in vivo studies investigating checkpoint inhibitor blockade in combination with other cancer immunotherapies. Eventually our goal is to establish pigs as an alternative large animal model...... Ipilimumab increases anticancer T-cell reactivity and overall survival of metastatic cancer patients. Indole-amine 2,3-dioxygenase (IDO) is another checkpoint inhibitor which suppresses T-cell immunity by the depletion of tryptophan in the T-cell microenvironment, and also inhibition of IDO by L-1...

  12. Tetrandrine: A Potent Abrogator of G2 Checkpoint Function in Tumor Cells and Its Mechanism

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    (M phase). Endosomatic experiment showed that tetrandrine caused tumor growth delay in irradiated mice. Conclusion Tetrandrine boosts the cell killing activity of irradiation both in vitro and in vivo. Tetrandrine is a potent abrogator for G2 checkpoint control and can sensitize the cells to radiation.

  13. How oocytes try to get it right: spindle checkpoint control in meiosis.

    Science.gov (United States)

    Touati, Sandra A; Wassmann, Katja

    2016-06-01

    The generation of a viable, diploid organism depends on the formation of haploid gametes, oocytes, and spermatocytes, with the correct number of chromosomes. Halving the genome requires the execution of two consecutive specialized cell divisions named meiosis I and II. Unfortunately, and in contrast to male meiosis, chromosome segregation in oocytes is error prone, with human oocytes being extraordinarily "meiotically challenged". Aneuploid oocytes, that are with the wrong number of chromosomes, give rise to aneuploid embryos when fertilized. In humans, most aneuploidies are lethal and result in spontaneous abortions. However, some trisomies survive to birth or even adulthood, such as the well-known trisomy 21, which gives rise to Down syndrome (Nagaoka et al. in Nat Rev Genet 13:493-504, 2012). A staggering 20-25 % of oocytes ready to be fertilized are aneuploid in humans. If this were not bad enough, there is an additional increase in meiotic missegregations as women get closer to menopause. A woman above 40 has a risk of more than 30 % of getting pregnant with a trisomic child. Worse still, in industrialized western societies, child birth is delayed, with women getting their first child later in life than ever. This trend has led to an increase of trisomic pregnancies by 70 % in the last 30 years (Nagaoka et al. in Nat Rev Genet 13:493-504, 2012; Schmidt et al. in Hum Reprod Update 18:29-43, 2012). To understand why errors occur so frequently during the meiotic divisions in oocytes, we review here the molecular mechanisms at works to control chromosome segregation during meiosis. An important mitotic control mechanism, namely the spindle assembly checkpoint or SAC, has been adapted to the special requirements of the meiotic divisions, and this review will focus on our current knowledge of SAC control in mammalian oocytes. Knowledge on how chromosome segregation is controlled in mammalian oocytes may help to identify risk factors important for questions

  14. Structural and functional analysis of the Crb2–BRCT2 domain reveals distinct roles in checkpoint signaling and DNA damage repair

    OpenAIRE

    Kilkenny, Mairi L.; Doré, Andrew S.; Roe, S. Mark; Nestoras, Konstantinos; Ho, Jenny C. Y.; Watts, Felicity Z.; Pearl, Laurence H.

    2008-01-01

    Schizosaccharomyces pombe Crb2 is a checkpoint mediator required for the cellular response to DNA damage. Like human 53BP1 and Saccharomyces cerevisiae Rad9 it contains Tudor2 and BRCT2 domains. Crb2-Tudor2 domain interacts with methylated H4K20 and is required for recruitment to DNA dsDNA breaks. The BRCT2 domain is required for dimerization, but its precise role in DNA damage repair and checkpoint signaling is unclear. The crystal structure of the Crb2–BRCT2 domain, alone and in complex wit...

  15. Interactions of checkpoint-genes RAD9, RAD17, RAD24 and RAD53 determining radioresistance of Yeast Saccharomyces Cerevisiae

    International Nuclear Information System (INIS)

    The mechanisms of genetic control of progress through the division cell cycle (checkpoint-control) in yeast Saccharomyces cerevisiae have been studied intensively. To investigate the role of checkpoint-genes RAD9, RAD17, RAD24, RAD53 in cell radioresistance we have investigated cell sensitivity of double mutants to γ-ray. Double mutants involving various combinations with rad9Δ show epistatic interactions, i.e. the sensitivity of the double mutants to γ-ray was no greater than that of more sensitive of the two single mutants. This suggests that all these genes govern the same pathway. This group of genes was named RAD9-epistasis group. It is interesting to note that the genes RAD9 and RAD53 have positive effect but RAD17 and RAD24 have negative effect on radiosensitivity of yeast cells. Interactions between mutations may differ depending on the agent γ-ray or UV-light, for example mutations rad9Δ and rad24Δ show additive effect for γ-ray and epistatic effect for UV-light

  16. NDR1 modulates the UV-induced DNA-damage checkpoint and nucleotide excision repair

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jeong-Min; Choi, Ji Ye [Department of Biological Science, Dong-A University, Busan (Korea, Republic of); Yi, Joo Mi [Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan (Korea, Republic of); Chung, Jin Woong; Leem, Sun-Hee; Koh, Sang Seok [Department of Biological Science, Dong-A University, Busan (Korea, Republic of); Kang, Tae-Hong, E-mail: thkang@dau.ac.kr [Department of Biological Science, Dong-A University, Busan (Korea, Republic of)

    2015-06-05

    Nucleotide excision repair (NER) is the sole mechanism of UV-induced DNA lesion repair in mammals. A single round of NER requires multiple components including seven core NER factors, xeroderma pigmentosum A–G (XPA–XPG), and many auxiliary effector proteins including ATR serine/threonine kinase. The XPA protein helps to verify DNA damage and thus plays a rate-limiting role in NER. Hence, the regulation of XPA is important for the entire NER kinetic. We found that NDR1, a novel XPA-interacting protein, modulates NER by modulating the UV-induced DNA-damage checkpoint. In quiescent cells, NDR1 localized mainly in the cytoplasm. After UV irradiation, NDR1 accumulated in the nucleus. The siRNA knockdown of NDR1 delayed the repair of UV-induced cyclobutane pyrimidine dimers in both normal cells and cancer cells. It did not, however, alter the expression levels or the chromatin association levels of the core NER factors following UV irradiation. Instead, the NDR1-depleted cells displayed reduced activity of ATR for some set of its substrates including CHK1 and p53, suggesting that NDR1 modulates NER indirectly via the ATR pathway. - Highlights: • NDR1 is a novel XPA-interacting protein. • NDR1 accumulates in the nucleus in response to UV irradiation. • NDR1 modulates NER (nucleotide excision repair) by modulating the UV-induced DNA-damage checkpoint response.

  17. Stranglehold on the spindle assembly checkpoint: the human papillomavirus E2 protein provokes BUBR1-dependent aneuploidy.

    Science.gov (United States)

    Tan, Chye Ling; Teissier, Sébastien; Gunaratne, Jayantha; Quek, Ling Shih; Bellanger, Sophie

    2015-01-01

    The Human Papillomavirus (HPV) E2 protein, which inhibits the E6 and E7 viral oncogenes, is believed to have anti-oncogenic properties. Here, we challenge this view and show that HPV-18 E2 over-activates the Spindle Assembly Checkpoint (SAC) and induces DNA breaks in mitosis followed by aneuploidy. This phenotype is associated with interaction of E2 with the Mitotic Checkpoint Complex (MCC) proteins Cdc20, MAD2 and BUBR1. While BUBR1 silencing rescues the mitotic phenotype induced by E2, p53 silencing or presence of E6/E7 (inactivating p53 and increasing BUBR1 levels respectively) both amplify it. This work pinpoints E2 as a key protein in the initiation of HPV-induced cervical cancer and identifies the SAC as a target for oncogenic pathogens. Moreover, our results suggest a role of p53 in regulating the mitotic process itself and highlight SAC over-activation in a p53-negative context as a highly pathogenic event. PMID:25789401

  18. G1 checkpoint is compromised in mouse ESCs due to functional uncoupling of p53-p21Waf1 signaling.

    Science.gov (United States)

    Suvorova, Irina I; Grigorash, Bogdan B; Chuykin, Ilya A; Pospelova, Tatiana V; Pospelov, Valery A

    2016-01-01

    Mouse embryonic stem cells (mESCs) lack of G1 checkpoint despite that irradiation (IR) activates ATM/ATR-mediated DDR signaling pathway. The IR-induced p53 localizes in the nuclei and up-regulates p21/Waf1 transcription but that does not lead to accumulation of p21/Waf1 protein. The negative control of the p21Waf1 expression appears to occur at 2 levels of regulation. First, both p21/Waf1 gene transcription and the p21/Waf1 protein content increase in mESCs treated with histone-deacetylase inhibitors, implying its epigenetic regulation. Second, proteasome inhibitors cause the p21/Waf1 accumulation, indicating that the protein is a subject of proteasome-dependent degradation in ESСs. Then, the dynamics of IR-induced p21Waf1 protein show its accumulation at long-term time points (3 and 5 days) that coincides with an increase in the proportion of G1-phase cells, down-regulation of Oct4 and Nanog pluripotent gene transcription and activation of endoderm-specific genes sox17 and afp. In addition, nutlin-dependent stabilization of p53 in mESC was also accompanied by the accumulation of p21/Waf1 as well as restoration of G1 checkpoint and an onset of differentiation. Thus, the lack of functional p21/Waf1 is indispensable for maintaining self-renewal and pluripotency of mESCs. PMID:26636245

  19. NDR1 modulates the UV-induced DNA-damage checkpoint and nucleotide excision repair

    International Nuclear Information System (INIS)

    Nucleotide excision repair (NER) is the sole mechanism of UV-induced DNA lesion repair in mammals. A single round of NER requires multiple components including seven core NER factors, xeroderma pigmentosum A–G (XPA–XPG), and many auxiliary effector proteins including ATR serine/threonine kinase. The XPA protein helps to verify DNA damage and thus plays a rate-limiting role in NER. Hence, the regulation of XPA is important for the entire NER kinetic. We found that NDR1, a novel XPA-interacting protein, modulates NER by modulating the UV-induced DNA-damage checkpoint. In quiescent cells, NDR1 localized mainly in the cytoplasm. After UV irradiation, NDR1 accumulated in the nucleus. The siRNA knockdown of NDR1 delayed the repair of UV-induced cyclobutane pyrimidine dimers in both normal cells and cancer cells. It did not, however, alter the expression levels or the chromatin association levels of the core NER factors following UV irradiation. Instead, the NDR1-depleted cells displayed reduced activity of ATR for some set of its substrates including CHK1 and p53, suggesting that NDR1 modulates NER indirectly via the ATR pathway. - Highlights: • NDR1 is a novel XPA-interacting protein. • NDR1 accumulates in the nucleus in response to UV irradiation. • NDR1 modulates NER (nucleotide excision repair) by modulating the UV-induced DNA-damage checkpoint response

  20. In silico analysis of deleterious single nucleotide polymorphisms in human BUB1 mitotic checkpoint serine/threonine kinase B gene.

    Science.gov (United States)

    Akhoundi, Fatemeh; Parvaneh, Nikpour; Modjtaba, Emadi-Baygi

    2016-09-01

    One of the major challenges in the analysis of human genetic variation is to distinguish mutations that are functionally neutral from those that contribute to disease. BubR1 is a key protein mediating spindle-checkpoint activation that plays a role in the inhibition of the anaphase-promoting complex/cyclosome (APC/C), delaying the onset of anaphase and ensuring proper chromosome segregation. Owing to the importance of BUB1B gene in mitotic checkpoint a functional analysis using different in silico approaches was undertaken to explore the possible associations between genetic mutations and phenotypic variation. In this work we found that 3 nsSNPs I82N, P334L and R814H have a functional effect on protein function and stability. A literature search revealed that R814H was already implicated in human diseases. Additionally, 2 SNPs in the 5' UTR region was predicted to exhibit a pattern change in the internal ribosome entry site (IRES), and eight MicroRNA binding sites were found to be highly affected due to 3' UTR SNPs. These in silico predictions will provide useful information in selecting the target SNPs that are likely to have functional impact on the BUB1B gene. PMID:27331020

  1. Ermenilerin Kontrol Noktası: İskenderun Limanı Alexandretta Port - Checkpoint For Armenians

    Directory of Open Access Journals (Sweden)

    Naim ÜRKMEZ

    2013-07-01

    Full Text Available The period of Yavuz Sultan Selim and the Ottoman Empire’s entering Alexandretta until the first half of the 19th century had identity of a very small village. Right to the end of the century, the industry revolution’s impact and trading goods’ large amount of transport means gained importance starting especially in the city of Aleppo and then in one of South Anatolia’s natural ports as of the day the preliminary plan came out. After the 1890’s the increase of Armenian activities in Anatolia and the outbreak of revolts in many other places parallel the Ottoman government’s large forces’ involvement to prevent and be able to keep Armenian rebels under control, taking various measures. Abdul Hamid II started under this negative initiative; the Sultan tried to resist in various ways. From these measures in any of the provinces of Aleppo, Mamuretülaziz, Adana, Kayseri, Bitlis, Van, Diyarbekir, or Erzurum, Armenians living in the diaspora and connecting supplier port, as in Alexandretta, have a checkpoint facility. Sultan Abdul Hamid II’s direction of the facility is seen at the checkpoint from the dock outside rising or Anatolia geography’s various residential areas incoming Armenian travelers often reserved monthly lists. In addition to this, England in the grip of Cyprus was able to check Armenians’ entries into Anatolia set by a weapons and bombs training field. It is understood that this matter is elaborated on further in books prepared from lists. These books will come to light with the acceleration of the classification of records. Yavuz Sultan Selim döneminde Osmanlı hâkimiyetine giren İskenderun, 19. yüzyılın ilk yarısına kadar çok küçük bir köy hüviyetinde kalmıştır. Yüzyılın sonuna doğru sanayi devriminin etkisi ve ticari emtianın çokça taşınması vesilesi ile önem kazanmaya başlayan şehir özellikle Halep ve ardı ile Güney Anadolu’nun doğal bir limanı olarak gün geçtikçe ön plana

  2. Structure of a Blinkin-BUBR1 complex reveals an interaction crucial for kinetochore-mitotic checkpoint regulation via an unanticipated binding Site

    DEFF Research Database (Denmark)

    Bolanos-Garcia, Victor M; Lischetti, Tiziana; Matak-Vinković, Dijana;

    2011-01-01

    The maintenance of genomic stability relies on the spindle assembly checkpoint (SAC), which ensures accurate chromosome segregation by delaying the onset of anaphase until all chromosomes are properly bioriented and attached to the mitotic spindle. BUB1 and BUBR1 kinases are central for this proc...

  3. Abrogation of Chk1-mediated S/G2 checkpoint by UCN-01 enhances ara-C-induced cytotoxicity in human colon cancer cells

    Institute of Scientific and Technical Information of China (English)

    Rong-guang SHAO; Chun-Xia CAO; Yves POMMIER

    2004-01-01

    AIM: To investigate whether 7-hydroxystaurosporine (UCN-01) affects cell cycle progression in arabinosylcytosine (ara-C) treated human colon carcinoma HT-29 cells. METHODS: Cytotoxicity, DNA synthesis, cell cycle distribution,protein level, and kinase activity were determined by clonogenic assay, flow cytometry, DNA synthesis assay,immunoblotting, and kinase assays, respectively. RESULTS: UCN-01 abrogated an S/G2-phase checkpoint in HT29 cells treated with ara-C. When UCN-01 was added after treatment with ara-C, the rate of recovery of DNA synthesis was enhanced and colony-forming ability diminished. Thus, premature recovery of DNA synthesis was associated with increased cytotoxicity. Measurements of cyclin A and B protein levels, Cdk2 and Cdc2 kinase activities, Cdc25C phosphorylation, and Chkl kinase activity were consistent with UCN-01-induced abrogation of the S/G2-phase checkpoint in ara-C treated cells. CONCLUSION: The abrogation of the S/G2 checkpoint may be due to inhibition of Chkl kinase by UCN-01. The enhanced cytotoxicity produced when UCN-01 was combined with ara-C suggested a rationale for the use of this drug combination for tumors that might be susceptible to cell cycle checkpoint abrogation.

  4. Xenopus Cds1 Is Regulated by DNA-Dependent Protein Kinase and ATR during the Cell Cycle Checkpoint Response to Double-Stranded DNA Ends

    Science.gov (United States)

    McSherry, Troy D.; Mueller, Paul R.

    2004-01-01

    The checkpoint kinase Cds1 (Chk2) plays a key role in cell cycle checkpoint responses with functions in cell cycle arrest, DNA repair, and induction of apoptosis. Proper regulation of Cds1 is essential for appropriate cellular responses to checkpoint-inducing insults. While the kinase ATM has been shown to be important in the regulation of human Cds1 (hCds1), here we report that the kinases ATR and DNA-dependent protein kinase (DNA-PK) play more significant roles in the regulation of Xenopus Cds1 (XCds1). Under normal cell cycle conditions, nonactivated XCds1 constitutively associates with a Xenopus ATR complex. The association of XCds1 with this complex does not require a functional forkhead activation domain but does require a putative SH3 binding region that is found in XCds1. In response to double-stranded DNA ends, the amino terminus of XCds1 is rapidly phosphorylated in a sequential pattern. First DNA-PK phosphorylates serine 39, a site not previously recognized as important in Cds1 regulation. Xenopus ATM, ATR, and/or DNA-PK then phosphorylate three consensus serine/glutamine sites. Together, these phosphorylations have the dual function of inducing dissociation from the ATR complex and independently promoting the full activation of XCds1. Thus, the checkpoint-mediated activation of XCds1 requires phosphorylation by multiple phosphoinositide 3-kinase-related kinases, protein-protein dissociation, and autophosphorylation. PMID:15509799

  5. The Level of Europium-154 Contaminating Samarium-153-EDTMP Activates the Radiation Alarm System at the US Homeland Security Checkpoints

    Directory of Open Access Journals (Sweden)

    Mohammed Najeeb Al Hallak

    2009-08-01

    Full Text Available 153Sm-EDTMP is a radiopharmaceutical composed of EDTMP (ethylenediamine-tetramethylenephosphonate and Samarium-153 [1]. 153Sm-EDTMP has an affinity for skeletal tissue and concentrates in areas with increased bone turnover; thus, it is successfully used in relieving pain related to diffuse bone metastases [1]. The manufacturing process of 153Sm-EDTMP leads to contamination with 154Eu (Europium-154 [2]. A previous study only alluded to the retention of 154Eu in the bones after receiving treatment with 153Sm-EDTMP [2]. Activation of the alarm at security checkpoints after 153Sm-EDTMP therapy has not been previously reported. Two out of 15 patients who received 153Sm-EDTMP at Roger Maris Cancer Center (Fargo, N. Dak., USA activated the radiation activity sensors while passing through checkpoints; one at a US airport and the other while crossing theAmerican-Canadian border. We assume that the 154Eu which remained in the patients’ bones activated the sensors. Methods: In order to investigate this hypothesis, we obtained the consent from 3 of our 15 patients who received 153Sm-EDTMP within the previous 4 months to 2 years, including the patient who had activated the radiation alarm at the airport. The patients were scanned with a handheld detector and a gamma camera for energies from 511 keV to 1.3 MeV. Results: All three patients exhibited identical spectral images, and further analysis showed that the observed spectra are the result of 154Eu emissions. Conclusion: Depending on the detection thresholds and windows used by local and federal authorities, the remaining activity of 154Eu retained in patients who received 153Sm-EDTMP could be sufficient enough to increase the count rates above background levels and activate the sensors. At Roger Maris Cancer Center, patients are now informed of the potential consequences of 153Sm-EDTMP therapy prior to initiating treatment. In addition, patients treated with 153Sm-EDTMP at Roger Maris Cancer Center

  6. Centromere-tethered Mps1 pombe homolog (Mph1) kinase is a sufficient marker for recruitment of the spindle checkpoint protein Bub1, but not Mad1.

    Science.gov (United States)

    Ito, Daisuke; Saito, Yu; Matsumoto, Tomohiro

    2012-01-01

    The spindle checkpoint delays the onset of anaphase until all of the chromosomes properly achieve bipolar attachment to the spindle. It has been shown that unattached kinetochores are the site that emits a signal for activation of the checkpoint. Although the components of the checkpoint such as Bub1, Mad1 and Mad2 selectively accumulate at unattached kinetochores, the answer to how they recognize unattached kinetochores has remained elusive. Mps1 pombe homolog (Mph1) kinase has been shown to function upstream of most of the components of the checkpoint and thus it is thought to recognize unattached kinetochores by itself and recruit other components. In this study we have expressed a fusion protein of Mph1 and Ndc80 (a kinetochore protein of the outer plate) and shown that the fusion protein arrests cell cycle progression in a spindle-checkpoint\\x{2013}dependent manner in fission yeast. When expression of Mad2 is turned off, the cells grow normally with Mph1 constitutively localized at centromeres/kinetochores. Under this condition, Bub1 can be found with Mph1 throughout the cell cycle, indicating that localization of Mph1 at centromeres/kinetochores is sufficient to recruit Bub1. In contrast, Mad1 is found to transiently localize at kinetochores, which are presumably unattached to the spindle, but soon it dissociates from kinetochores. We propose that Mph1 is a sufficient marker for recruitment of Bub1. Mad1, in contrast, requires an additional condition/component for stable association with kinetochores. PMID:22184248

  7. Rad4 mainly functions in Chk1-mediated DNA damage checkpoint pathway as a scaffold protein in the fission yeast Schizosaccharomyces pombe.

    Directory of Open Access Journals (Sweden)

    Ming Yue

    Full Text Available Rad4/Cut5 is a scaffold protein in the Chk1-mediated DNA damage checkpoint in S. pombe. However, whether it contains a robust ATR-activation domain (AAD required for checkpoint signaling like its orthologs TopBP1 in humans and Dpb11 in budding yeast has been incompletely clear. To identify the putative AAD in Rad4, we carried out an extensive genetic screen looking for novel mutants with an enhanced sensitivity to replication stress or DNA damage in which the function of the AAD can be eliminated by the mutations. Two new mutations near the N-terminus were identified that caused significantly higher sensitivities to DNA damage or chronic replication stress than all previously reported mutants, suggesting that most of the checkpoint function of the protein is eliminated. However, these mutations did not affect the activation of Rad3 (ATR in humans yet eliminated the scaffolding function of the protein required for the activation of Chk1. Several mutations were also identified in or near the recently reported AAD in the C-terminus of Rad4. However, all mutations in the C-terminus only slightly sensitized the cells to DNA damage. Interestingly, a mutant lacking the whole C-terminus was found resistant to DNA damage and replication stress almost like the wild type cells. Consistent with the resistance, all known Rad3 dependent phosphorylations of checkpoint proteins remained intact in the C-terminal deletion mutant, indicating that unlike that in Dpb11, the C-terminus of Rad4 does not contain a robust AAD. These results, together with those from the biochemical studies, show that Rad4 mainly functions as a scaffold protein in the Chk1, not the Cds1(CHK2 in humans, checkpoint pathway. It plays a minor role or is functionally redundant with an unknown factor in Rad3 activation.

  8. Cell-cycle dependent expression of a translocation-mediated fusion oncogene mediates checkpoint adaptation in rhabdomyosarcoma.

    Science.gov (United States)

    Kikuchi, Ken; Hettmer, Simone; Aslam, M Imran; Michalek, Joel E; Laub, Wolfram; Wilky, Breelyn A; Loeb, David M; Rubin, Brian P; Wagers, Amy J; Keller, Charles

    2014-01-01

    Rhabdomyosarcoma is the most commonly occurring soft-tissue sarcoma in childhood. Most rhabdomyosarcoma falls into one of two biologically distinct subgroups represented by alveolar or embryonal histology. The alveolar subtype harbors a translocation-mediated PAX3:FOXO1A fusion gene and has an extremely poor prognosis. However, tumor cells have heterogeneous expression for the fusion gene. Using a conditional genetic mouse model as well as human tumor cell lines, we show that that Pax3:Foxo1a expression is enriched in G2 and triggers a transcriptional program conducive to checkpoint adaptation under stress conditions such as irradiation in vitro and in vivo. Pax3:Foxo1a also tolerizes tumor cells to clinically-established chemotherapy agents and emerging molecularly-targeted agents. Thus, the surprisingly dynamic regulation of the Pax3:Foxo1a locus is a paradigm that has important implications for the way in which oncogenes are modeled in cancer cells. PMID:24453992

  9. Cell-cycle dependent expression of a translocation-mediated fusion oncogene mediates checkpoint adaptation in rhabdomyosarcoma.

    Directory of Open Access Journals (Sweden)

    Ken Kikuchi

    2014-01-01

    Full Text Available Rhabdomyosarcoma is the most commonly occurring soft-tissue sarcoma in childhood. Most rhabdomyosarcoma falls into one of two biologically distinct subgroups represented by alveolar or embryonal histology. The alveolar subtype harbors a translocation-mediated PAX3:FOXO1A fusion gene and has an extremely poor prognosis. However, tumor cells have heterogeneous expression for the fusion gene. Using a conditional genetic mouse model as well as human tumor cell lines, we show that that Pax3:Foxo1a expression is enriched in G2 and triggers a transcriptional program conducive to checkpoint adaptation under stress conditions such as irradiation in vitro and in vivo. Pax3:Foxo1a also tolerizes tumor cells to clinically-established chemotherapy agents and emerging molecularly-targeted agents. Thus, the surprisingly dynamic regulation of the Pax3:Foxo1a locus is a paradigm that has important implications for the way in which oncogenes are modeled in cancer cells.

  10. Human papillomavirus type 16 E7 oncoprotein engages but does not abrogate the mitotic spindle assembly checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Yueyang [Division of Infectious Diseases, Brigham and Women' s Hospital and Biological and Biomedical Sciences Program, Harvard Medical School, Boston, MA 02115 (United States); Munger, Karl, E-mail: kmunger@rics.bwh.harvard.edu [Division of Infectious Diseases, Brigham and Women' s Hospital and Biological and Biomedical Sciences Program, Harvard Medical School, Boston, MA 02115 (United States)

    2012-10-10

    The mitotic spindle assembly checkpoint (SAC) ensures faithful chromosome segregation during mitosis by censoring kinetochore-microtubule interactions. It is frequently rendered dysfunctional during carcinogenesis causing chromosome missegregation and genomic instability. There are conflicting reports whether the HPV16 E7 oncoprotein drives chromosomal instability by abolishing the SAC. Here we report that degradation of mitotic cyclins is impaired in cells with HPV16 E7 expression. RNAi-mediated depletion of Mad2 or BubR1 indicated the involvement of the SAC, suggesting that HPV16 E7 expression causes sustained SAC engagement. Mutational analyses revealed that HPV16 E7 sequences that are necessary for retinoblastoma tumor suppressor protein binding as well as sequences previously implicated in binding the nuclear and mitotic apparatus (NuMA) protein and in delocalizing dynein from the mitotic spindle contribute to SAC engagement. Importantly, however, HPV16 E7 does not markedly compromise the SAC response to microtubule poisons.

  11. Fault-Tolerance through Message-logging and Check-pointing: Disaster Recovery for CORBA-based Distributed Bank Servers

    CERN Document Server

    Vassev, Emil; Kuang, Heng

    2009-01-01

    This report presents results of our endeavor towards developing a failure-recovery variant of a CORBA-based bank server that provides fault tolerance features through message logging and checkpoint logging. In this group of projects, three components were developed to satisfy the requirements: 1) a message-logging protocol for the branch servers of the distributed banking system to log required information; 2) a recovery module that restarts the bank server using the message log to help the restarted bank server process subsequent requests for various operations; 3) a monitor module that periodically checks whether the bank server is down and helps the recovery module restart the bank server if the latter has crashed.

  12. RPA mediates recombination repair during replication stress and is displaced from DNA by checkpoint signalling in human cells

    DEFF Research Database (Denmark)

    Sleeth, Kate M; Sørensen, Claus Storgaard; Issaeva, Natalia;

    2007-01-01

    The replication protein A (RPA) is involved in most, if not all, nuclear metabolism involving single-stranded DNA. Here, we show that RPA is involved in genome maintenance at stalled replication forks by the homologous recombination repair system in humans. Depletion of the RPA protein inhibited...... the formation of RAD51 nuclear foci after hydroxyurea-induced replication stalling leading to persistent unrepaired DNA double-strand breaks (DSBs). We demonstrate a direct role of RPA in homology directed recombination repair. We find that RPA is dispensable for checkpoint kinase 1 (Chk1) activation...... and that RPA directly binds RAD52 upon replication stress, suggesting a direct role in recombination repair. In addition we show that inhibition of Chk1 with UCN-01 decreases dissociation of RPA from the chromatin and inhibits association of RAD51 and RAD52 with DNA. Altogether, our data suggest a...

  13. Spindle assembly checkpoint protein expression correlates with cellular proliferation and shorter time to recurrence in ovarian cancer.

    LENUS (Irish Health Repository)

    McGrogan, Barbara

    2014-07-01

    Ovarian carcinoma (OC) is the most lethal of the gynecological malignancies, often presenting at an advanced stage. Treatment is hampered by high levels of drug resistance. The taxanes are microtubule stabilizing agents, used as first-line agents in the treatment of OC that exert their apoptotic effects through the spindle assembly checkpoint. BUB1-related protein kinase (BUBR1) and mitotic arrest deficient 2 (MAD2), essential spindle assembly checkpoint components, play a key role in response to taxanes. BUBR1, MAD2, and Ki-67 were assessed on an OC tissue microarray platform representing 72 OC tumors of varying histologic subtypes. Sixty-one of these patients received paclitaxel and platinum agents combined; 11 received platinum alone. Overall survival was available for all 72 patients, whereas recurrence-free survival (RFS) was available for 66 patients. Increased BUBR1 expression was seen in serous carcinomas, compared with other histologies (P = .03). Increased BUBR1 was significantly associated with tumors of advanced stage (P = .05). Increased MAD2 and BUBR1 expression also correlated with increased cellular proliferation (P < .0002 and P = .02, respectively). Reduced MAD2 nuclear intensity was associated with a shorter RFS (P = .03), in ovarian tumors of differing histologic subtype (n = 66). In this subgroup, for those women who received paclitaxel and platinum agents combined (n = 57), reduced MAD2 intensity also identified women with a shorter RFS (P < .007). For the entire cohort of patients, irrespective of histologic subtype or treatment, MAD2 nuclear intensity retained independent significance in a multivariate model, with tumors showing reduced nuclear MAD2 intensity identifying patients with a poorer RFS (P = .05).

  14. Targeted therapies and immune checkpoint inhibitors in the treatment of metastatic melanoma patients: a guide and update for pathologists.

    Science.gov (United States)

    Kakavand, Hojabr; Wilmott, James S; Long, Georgina V; Scolyer, Richard A

    2016-02-01

    The previously dismal prospects for patients with advanced stage metastatic melanoma have greatly improved in recent years. Enhanced understanding of both the pathogenesis of melanoma and its molecular drivers, as well as the importance and regulation of anti-tumour immune responses, have provided new therapeutic opportunities for melanoma patients. There are two major distinct categories of systemic treatments with activity for patients with metastatic melanoma: (1) targeted therapies, which act to inhibit the oncogenes that drive the aberrant growth and dissemination of the tumour; and (2) immune checkpoint inhibitor therapies, which act to enhance anti-tumour immune responses by blocking negative regulators of immunity. Pathologists play a critical and expanding role in the selection of the most appropriate treatment for individual metastatic melanoma patients in the modern era of personalised/precision medicine. The molecular pathology testing of melanoma tumour tissue for the presence of targetable oncogenic mutations is already part of routine practice in many institutions. In addition, other potential oncogenic therapeutic targets continue to be identified and pathology testing techniques must readily adapt to this rapidly changing field. Recent research findings suggest that pathological assessment of tumour associated immune cells and immunosuppressive ligand expression of the tumour are likely to be important in identifying patients most likely to benefit from immune checkpoint inhibitors. Similarly, pathological and molecular observations of on-treatment tumour tissue biopsies taken from patients on targeted therapies have provided new insights into the mechanisms of action of targeted molecular therapies, have contributed to the identification of resistance mechanisms to these novel therapies and may be of higher value for selecting patients most likely to benefit from therapies. These data have already provided a rational biological basis for the

  15. Immune Reactivation by Cell-Free Fetal DNA in Healthy Pregnancies Re-Purposed to Target Tumors: Novel Checkpoint Inhibition in Cancer Therapeutics

    OpenAIRE

    Enninga, Elizabeth Ann L.; Nevala, Wendy K.; Holtan, Shernan G.; Svetomir N. Markovic

    2015-01-01

    The role of the immune system in cancer progression has become increasingly evident over the past decade. Chronic inflammation in the promotion of tumorigenesis is well established, and cancer-associated tolerance/immune evasion has long been appreciated. Recent developments of immunotherapies targeting cancer-associated inflammation and immune tolerance, such as cancer vaccines, cell therapies, neutralizing antibodies, and immune checkpoint inhibitors, have shown promising clinical results. ...

  16. A plant cyclin B2 is degraded early in mitosis and its ectopic expression shortens G2-phase and alleviates the DNA-damage checkpoint

    Czech Academy of Sciences Publication Activity Database

    Weingartner, M.; Pelayo, H. R.; Binarová, Pavla; Zwerger, K.; Melikant, B.; Torre, C.; Heberle-Bors, E.; Bogre, L.

    2003-01-01

    Roč. 116, č. 3 (2003), s. 487-498. ISSN 0021-9533 R&D Projects: GA MŠk LN00A081 Grant ostatní: GA Ministerio de Ciencia y Tecnologia (ES) BMC2001-2195; Biotechnology and Biological Science Research Council(AU) 111/P133340 Institutional research plan: CEZ:AV0Z5020903 Keywords : cyclin B * mitosis * checkpoint Subject RIV: EE - Microbiology, Virology Impact factor: 7.250, year: 2003

  17. Immune Checkpoint Blockade in Cancer Treatment: A Double-Edged Sword Cross-Targeting the Host as an “Innocent Bystander”

    Directory of Open Access Journals (Sweden)

    Lucia Gelao

    2014-03-01

    Full Text Available Targeted immune checkpoint blockade augments anti-tumor immunity and induces durable responses in patients with melanoma and other solid tumors. It also induces specific “immune-related adverse events” (irAEs. IrAEs mainly include gastrointestinal, dermatological, hepatic and endocrinological toxicities. Off-target effects that arise appear to account for much of the toxicity of the immune checkpoint blockade. These unique “innocent bystander” effects are likely a direct result of breaking immune tolerance upon immune check point blockade and require specific treatment guidelines that include symptomatic therapies or systemic corticosteroids. What do we need going forward to limit immune checkpoint blockade-induced toxicity? Most importantly, we need a better understanding of the roles played by these agents in normal tissues, so that we can begin to predict potentially problematic side effects on the basis of their selectivity profile. Second, we need to focus on the predictive factors of the response and toxicity of the host rather than serially focusing on individual agents. Third, rigorous biomarker-driven clinical trials are needed to further elucidate the mechanisms of both the benefit and toxicity. We will summarize the double-edged sword effect of immunotherapeutics in cancer treatment.

  18. ATM/ATR-related checkpoint signals mediate arsenite-induced G{sub 2}/M arrest in primary aortic endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Tsou, Tsui-Chun; Tsai, Feng-Yuan; Yeh, Szu-Ching; Chang, Louis W. [National Health Research Institutes, Division of Environmental Health and Occupational Medicine, Miaoli County (Taiwan)

    2006-12-15

    Epidemiological studies have demonstrated a high association of inorganic arsenic exposure with vascular disease. Our recent in vitro studies have linked this vascular damage to vascular endothelial dysfunction induced by arsenic exposure. However, cell-cycle arrest induced by arsenic and its involvement in vascular dysfunction remain to be clarified. In this study, we employed primary porcine aortic endothelial cells to investigate regulatory mechanisms of G{sub 2}/M phase arrest induced by arsenite. Our study revealed that lower concentrations of arsenite (1 and 3 {mu}M) increased cell proliferation, whereas higher concentrations of arsenite (10, 20, and 30 {mu}M) inhibited cell proliferation together with correlated increases in G{sub 2}/M phase arrest. We found that this arsenite-induced G{sub 2}/M phase arrest was accompanied by accumulation and/or phosphorylation of checkpoint-related molecules, including p53, Cdc25B, Cdc25C, and securin. Inhibition of activations of these checkpoint-related molecules by caffeine significantly attenuated the 30-{mu}M arsenite-induced G{sub 2}/M phase arrest by 93%. Our data suggest that the DNA damage responsive kinases ATM (ataxia-telangiectasia mutated) and ATR (ATM and Rad3-related) play critical roles in arsenite-induced G{sub 2}/M phase arrest in aortic endothelial cells possibly via regulation of checkpoint-related signaling molecules including p53, Cdc25B, Cdc25C, and securin. (orig.)

  19. Loss of yeast peroxiredoxin Tsa1p induces genome instability through activation of the DNA damage checkpoint and elevation of dNTP levels.

    Directory of Open Access Journals (Sweden)

    Hei-Man Vincent Tang

    2009-10-01

    Full Text Available Peroxiredoxins are a family of antioxidant enzymes critically involved in cellular defense and signaling. Particularly, yeast peroxiredoxin Tsa1p is thought to play a role in the maintenance of genome integrity, but the underlying mechanism is not understood. In this study, we took a genetic approach to investigate the cause of genome instability in tsa1Delta cells. Strong genetic interactions of TSA1 with DNA damage checkpoint components DUN1, SML1, and CRT1 were found when mutant cells were analyzed for either sensitivity to DNA damage or rate of spontaneous base substitutions. An elevation in intracellular dNTP production was observed in tsa1Delta cells. This was associated with constitutive activation of the DNA damage checkpoint as indicated by phosphorylation of Rad9/Rad53p, reduced steady-state amount of Sml1p, and induction of RNR and HUG1 genes. In addition, defects in the DNA damage checkpoint did not modulate intracellular level of reactive oxygen species, but suppressed the mutator phenotype of tsa1Delta cells. On the contrary, overexpression of RNR1 exacerbated this phenotype by increasing dNTP levels. Taken together, our findings uncover a new role of TSA1 in preventing the overproduction of dNTPs, which is a root cause of genome instability.

  20. Identification of a mutation causing a defective spindle assembly checkpoint in high ethyl caproate-producing sake yeast strain K1801.

    Science.gov (United States)

    Goshima, Tetsuya; Nakamura, Ryo; Kume, Kazunori; Okada, Hiroki; Ichikawa, Eri; Tamura, Hiroyasu; Hasuda, Hirokazu; Inahashi, Masaaki; Okazaki, Naoto; Akao, Takeshi; Shimoi, Hitoshi; Mizunuma, Masaki; Ohya, Yoshikazu; Hirata, Dai

    2016-08-01

    In high-quality sake brewing, the cerulenin-resistant sake yeast K1801 with high ethyl caproate-producing ability has been used widely; however, K1801 has a defective spindle assembly checkpoint (SAC). To identify the mutation causing this defect, we first searched for sake yeasts with a SAC-defect like K1801 and found that K13 had such a defect. Then, we searched for a common SNP in only K1801 and K13 by examining 15 checkpoint-related genes in 23 sake yeasts, and found 1 mutation, R48P of Cdc55, the PP2A regulatory B subunit that is important for the SAC. Furthermore, we confirmed that the Cdc55-R48P mutation was responsible for the SAC-defect in K1801 by molecular genetic analyses. Morphological analysis indicated that this mutation caused a high cell morphological variation. But this mutation did not affect the excellent brewing properties of K1801. Thus, this mutation is a target for breeding of a new risk-free K1801 with normal checkpoint integrity. PMID:27191586

  1. Inactivating the spindle checkpoint kinase Bub1 during embryonic development results in a global shutdown of proliferation

    Directory of Open Access Journals (Sweden)

    Taylor Stephen S

    2009-09-01

    Full Text Available Abstract Background Bub1 is a component of the spindle assembly checkpoint, a surveillance mechanism that maintains chromosome stability during M-phase. Bub1 is essential during the early stages of embryogenesis, with homozygous BUB1-null mice dying shortly after day E3.5. Bub1 is also required later during embryogenesis; inactivation of BUB1 on day E10.5 appears to rapidly block all further development. However, the mechanism(s responsible for this phenotype remain unclear. Findings Here we show that inactivating BUB1 on day E10.5 stalls embryogenesis within 48 hours. This is accompanied by a global shutdown of proliferation, widespread apoptosis and haemorrhaging. Conclusion Our results suggest that Bub1 is required throughout the developing embryo for cellular proliferation. Therefore, Bub1 has been shown to be essential in all scenarios analyzed thus far in mice: proliferation of cultured fibroblasts, spermatogenesis, oogenesis and both early and late embryonic development. This likely reflects the fact that Bub1 has dual functions during mitosis, being required for both SAC function and chromosome alignment.

  2. Toxic effect of silica nanoparticles on endothelial cells through DNA damage response via Chk1-dependent G2/M checkpoint.

    Directory of Open Access Journals (Sweden)

    Junchao Duan

    Full Text Available Silica nanoparticles have become promising carriers for drug delivery or gene therapy. Endothelial cells could be directly exposed to silica nanoparticles by intravenous administration. However, the underlying toxic effect mechanisms of silica nanoparticles on endothelial cells are still poorly understood. In order to clarify the cytotoxicity of endothelial cells induced by silica nanoparticles and its mechanisms, cellular morphology, cell viability and lactate dehydrogenase (LDH release were observed in human umbilical vein endothelial cells (HUVECs as assessing cytotoxicity, resulted in a dose- and time- dependent manner. Silica nanoparticles-induced reactive oxygen species (ROS generation caused oxidative damage followed by the production of malondialdehyde (MDA as well as the inhibition of superoxide dismutase (SOD and glutathione peroxidase (GSH-Px. Both necrosis and apoptosis were increased significantly after 24 h exposure. The mitochondrial membrane potential (MMP decreased obviously in a dose-dependent manner. The degree of DNA damage including the percentage of tail DNA, tail length and Olive tail moment (OTM were markedly aggravated. Silica nanoparticles also induced G2/M arrest through the upregulation of Chk1 and the downregulation of Cdc25C, cyclin B1/Cdc2. In summary, our data indicated that the toxic effect mechanisms of silica nanoparticles on endothelial cells was through DNA damage response (DDR via Chk1-dependent G2/M checkpoint signaling pathway, suggesting that exposure to silica nanoparticles could be a potential hazards for the development of cardiovascular diseases.

  3. The energy sensor AMPK regulates Hedgehog signaling in human cells through a unique Gli1 metabolic checkpoint

    Science.gov (United States)

    Di Magno, Laura; Basile, Alessio; Coni, Sonia; Manni, Simona; Sdruscia, Giulia; D'Amico, Davide; Antonucci, Laura; Infante, Paola; De Smaele, Enrico; Cucchi, Danilo; Ferretti, Elisabetta; Di Marcotullio, Lucia; Screpanti, Isabella; Canettieri, Gianluca

    2016-01-01

    Hedgehog signaling controls proliferation of cerebellar granule cell precursors (GCPs) and its aberrant activation is a leading cause of Medulloblastoma, the most frequent pediatric brain tumor. We show here that the energy sensor AMPK inhibits Hh signaling by phosphorylating a single residue of human Gli1 that is not conserved in other species. Studies with selective agonists and genetic deletion have revealed that AMPK activation inhibits canonical Hh signaling in human, but not in mouse cells. Indeed we show that AMPK phosphorylates Gli1 at the unique residue Ser408, which is conserved only in primates but not in other species. Once phosphorylated, Gli1 is targeted for proteasomal degradation. Notably, we show that selective AMPK activation inhibits Gli1-driven proliferation and that this effect is linked to Ser408 phosphorylation, which represents a key metabolic checkpoint for Hh signaling. Collectively, this data unveil a novel mechanism of inhibition of Gli1 function, which is exclusive for human cells and may be exploited for the treatment of Medulloblastoma or other Gli1 driven tumors. PMID:26843621

  4. Differential contribution of three immune checkpoint (VISTA, CTLA-4, PD-1) pathways to antitumor responses against squamous cell carcinoma.

    Science.gov (United States)

    Kondo, Yuta; Ohno, Tatsukuni; Nishii, Naoto; Harada, Kiyoshi; Yagita, Hideo; Azuma, Miyuki

    2016-06-01

    V domain-containing Ig suppressor of T-cell activation (VISTA)/PD-1H is a novel immune checkpoint molecule for regulating T-cell activation. We examined the effects of anti-VISTA mAb monotherapy and combination therapy with CTLA-4 or PD-1 blockade in a squamous cell carcinoma (SCCVII) model. VISTA monotherapy did not show clear tumor growth regression, but efficiently induced CD8(+) T cell activation by converting resting and exhausted cells into functional effector cells. VISTA monotherapy did not inhibit recruitment of regulatory T cells (Tregs) in the tumor microenvironment (TME). As an additional treatment to VISTA, CTLA-4 blockade, but not PD-1 blockade, elicited further tumor regression. The CTLA-4 and VISTA combination efficiently inhibited Treg recruitment and increased the ratios of both CD8 T/Treg and CD4 conventional T (Tcon)/Treg in the TME, whereas the PD-1 and VISTA combination dramatically increased tumor-recruiting CD8(+) T cells, but markedly reduced the Tcon/Treg ratio. Our results demonstrate that VISTA blockade efficiently converts CD8(+) T cells into functional effector T cells, but is not sufficient to regress tumor growth due to weak Treg suppression in the TME. Our results suggest that combined CTLA-4 and VISTA blockade is more efficacious than combined PD-1 and VISTA blockade for tumors like head and neck squamous cell carcinoma in which Treg-mediated immune regulation is dominant. PMID:27208845

  5. Human Zwint-1 Specifies Localization of Zeste White 10 to Kinetochores and Is Essential for Mitotic Checkpoint Signaling

    Institute of Scientific and Technical Information of China (English)

    HongmeiWang; XiaoyuHu; XiaDing; ZhenDou; ZhihongYank; AndrewW.Shaw; MaikunTcng; DonW.Cleveland; MichaelL.Goldberg; LiwenNiu; XucbiaoYao

    2005-01-01

    Chromosome segregation in mitosis is orchestrated by dynamic interaction between spindle microtubules and the kinetochore, a multiprotein complex assembled onto centromeric DNA of the chromosome. Here we show that Zwint-1 is required and is sufficient for kinetochore localization of Zestc White 10 (ZW10) in HeLa cells. Zwint-1 specifies the kinetochore association of ZW10 by interacting with its N-terminal domain. Suppression of synthesis of Zwint-1 by small interfering RNA abolishes the localization of ZW10 to the kinetochore, demonstrating the requirement of Zwint-1 for ZWl0 kinetochore localization. In addition, dcplction of Zwint-1 affects no mitotic arrest but causes aberrant premature chromo. some segregation. These Zwint-l-suppressed cells dis. play chromosome bridge phenotype with sister chromatids inter-connected. Moreover, Zwint-1 is required for stable association of CENP.F and dynamitin but not BUB1 with the kinetochore. Finally, our studies showthat Zwint-1 is a new component of the mitotic check. point, as cells lacking Zwint-1 fail to arrest in mitosis when exposed to microtubule inhibitors, yielding inter. phase cells with multinuclei. As ZWl0 and Zwint.1 are absent from yeast, we reasoned that metazoans evolved an elaborate spindle checkpoint machinery to ensure faithful chromosome segregation in mitosis.

  6. Taxifolin enhances andrographolide-induced mitotic arrest and apoptosis in human prostate cancer cells via spindle assembly checkpoint activation.

    Directory of Open Access Journals (Sweden)

    Zhong Rong Zhang

    Full Text Available Andrographolide (Andro suppresses proliferation and triggers apoptosis in many types of cancer cells. Taxifolin (Taxi has been proposed to prevent cancer development similar to other dietary flavonoids. In the present study, the cytotoxic and apoptotic effects of the addition of Andro alone and Andro and Taxi together on human prostate carcinoma DU145 cells were assessed. Andro inhibited prostate cancer cell proliferation by mitotic arrest and activation of the intrinsic apoptotic pathway. Although the effect of Taxi alone on DU145 cell proliferation was not significant, the combined use of Taxi with Andro significantly potentiated the anti-proliferative effect of increased mitotic arrest and apoptosis by enhancing the cleavage of poly(ADP-ribose polymerase, and caspases-7 and -9. Andro together with Taxi enhanced microtubule polymerization in vitro, and they induced the formation of twisted and elongated spindles in the cancer cells, thus leading to mitotic arrest. In addition, we showed that depletion of MAD2, a component in the spindle assembly checkpoint (SAC, alleviated the mitotic block induced by the two compounds, suggesting that they trigger mitotic arrest by SAC activation. This study suggests that the anti-cancer activity of Andro can be significantly enhanced in combination with Taxi by disrupting microtubule dynamics and activating the SAC.

  7. Modulation of human checkpoint kinase Chk1 by the regulatory beta-subunit of protein kinase CK2

    DEFF Research Database (Denmark)

    Guerra, Barbara; Issinger, Olaf-Georg; Wang, Jean Y J

    2003-01-01

    Protein kinase CK2 is a serine/threonine protein kinase involved in various aspects of cellular regulation. The regulatory beta-subunit of CK2 exerts a central role not only in mediating formation of tetrameric CK2 complexes but also as a docking partner for several protein kinases. In this study......, CK2beta is found to interact with the human cell cycle checkpoint kinase Chk1. The Chk1-interacting region of CK2beta is localized at the C-terminus and the complex between CK2beta and Chk1 is devoid of the catalytic CK2alpha-subunit. The interaction between CK2beta and Chk1 leads to an increase in...... the Cdc25C phosphorylation activity of Chk1. The screening of several cell lines has revealed that the association between CK2beta and Chk1 also occurs in vivo at a different degree. Collectively, these studies confirm the implication of the regulatory beta-subunit of protein kinase CK2 in cell cycle...

  8. Ataxia telangiectasia-mutated-Rad3-related DNA damage checkpoint signaling pathway triggered by hepatitis B virus infection

    Institute of Scientific and Technical Information of China (English)

    Fan Zhao; Qing-Jun Ma; Hui Zhong; Ning-Bo Hou; Xiao-Li Yang; Xiang He; Yu Liu; Yan-Hong Zhang; Cong-Wen Wei; Ting Song; Li Li

    2008-01-01

    AIM: To explore whether acute cellular DNA damage response is induced upon hepatitis B virus (HBV) infection and the effects of the HBV infection. METHODS: We incubated HL7702 hepatocytes with HBV-positive serum, mimicking a natural HBV infection process. We used immunoblotting to evaluate protein expression levels in HBV-infected cells or in non-infected cells; immunofluorescence to show ATR foci ands Chk1 phosphorylation loci formation; flow cytometry to analyze the cell cycle and apoptosis; ultraviolet (UV) radiation and ionizing radiation (IR)-treated cells to mimic DNA damage; and Trypan blue staining to count the viable cells.RESULTS: We found that HBV infection induced an increased steady state of ATR protein and increased phosphorylation of multiple downstream targets including Chkl, p53 and H2AX. In contrast to ATR and its target, the phosphorylated form of ATM at Ser-1981 and its downstream substrate Chk2 phosphorylation at Thr-68 did not visibly increase upon infection. However, the level of Mre11 and p21 were reduced beginning at 0.5 h after HBV-positive serum addition. Also, HBV infection led to transient cell cycle arrest in the S and the G2 phases without accompanying increased apoptosis. Research on cell survival changes upon radiation following HBV infection showed that survival of UV-treated host cells was greatly increased by HBV infection, owing to the reduced apoptosis. Meanwhile, survival of IR-treated host cells was reduced by HBV infection. CONCLUSION: HBV infection activates ATR DNA damage response to replication stress and abrogates the checkpoint signaling controlled by DNA damage response.

  9. Knockdown of checkpoint kinase 1 is associated with the increased radiosensitivity of glioblastoma stem-like cells

    International Nuclear Information System (INIS)

    Glioblastoma multiforme is an aggressive brain tumor with a poor prognosis. The glioblastoma stem-like cells (GSCs) represent a rare fraction of human glioblastoma cells with the capacity for multi-lineage differentiation, self-renewal and exact recapitulation of the original tumor. Interestingly, GSCs are more radioresistant compared with other tumor cells. In addition, the remarkable radioresistance of GSCs has been known to promote radiotherapy failure and therefore is associated with a significantly higher risk of a local tumor recurrence. Moreover, the hyperactive cell cycle checkpoint kinase (Chk) 1 and 2 play a pivotal role in the DNA damage response including radiation and chemical therapy. Based on aforementioned, we hypothesized that knockdown of Chk1 or Chk2 might confer radiosensitivity on GSCs and thereby increases the efficiency of radiotherapy. In this study, we knocked down the expression of Chk1 or Chk2 in human GSCs using lentivirus-delivered short hairpin RNA (shRNA) to examine its effect on the radiosensitivity. After radiation, the apoptosis rate and the cell cycle of GSCs were measured with Flow Cytometry. Compared with control GSCs (apoptosis, 7.82±0.38%; G2/M arrest, 60.20±1.28%), Chk1 knockdown in GSCs increased the apoptosis rate (37.87±0.32%) and decreased the degree of the G2/M arrest (22.37±2.01%). In contrast, the radiosensitivity was not enhanced by Chk2 knockdown in GSCs. These results suggest that depletion of Chk1 may improve the radio-sensitivity of GSCs via inducing cell apoptosis. In summary, the therapy targeting Chk1 gene in the GSCs may be a novel way to treat glioblastoma. (author)

  10. Human CAR T cells with cell-intrinsic PD-1 checkpoint blockade resist tumor-mediated inhibition.

    Science.gov (United States)

    Cherkassky, Leonid; Morello, Aurore; Villena-Vargas, Jonathan; Feng, Yang; Dimitrov, Dimiter S; Jones, David R; Sadelain, Michel; Adusumilli, Prasad S

    2016-08-01

    Following immune attack, solid tumors upregulate coinhibitory ligands that bind to inhibitory receptors on T cells. This adaptive resistance compromises the efficacy of chimeric antigen receptor (CAR) T cell therapies, which redirect T cells to solid tumors. Here, we investigated whether programmed death-1-mediated (PD-1-mediated) T cell exhaustion affects mesothelin-targeted CAR T cells and explored cell-intrinsic strategies to overcome inhibition of CAR T cells. Using an orthotopic mouse model of pleural mesothelioma, we determined that relatively high doses of both CD28- and 4-1BB-based second-generation CAR T cells achieved tumor eradication. CAR-mediated CD28 and 4-1BB costimulation resulted in similar levels of T cell persistence in animals treated with low T cell doses; however, PD-1 upregulation within the tumor microenvironment inhibited T cell function. At lower doses, 4-1BB CAR T cells retained their cytotoxic and cytokine secretion functions longer than CD28 CAR T cells. The prolonged function of 4-1BB CAR T cells correlated with improved survival. PD-1/PD-1 ligand [PD-L1] pathway interference, through PD-1 antibody checkpoint blockade, cell-intrinsic PD-1 shRNA blockade, or a PD-1 dominant negative receptor, restored the effector function of CD28 CAR T cells. These findings provide mechanistic insights into human CAR T cell exhaustion in solid tumors and suggest that PD-1/PD-L1 blockade may be an effective strategy for improving the potency of CAR T cell therapies. PMID:27454297

  11. Piperine causes G1 phase cell cycle arrest and apoptosis in melanoma cells through checkpoint kinase-1 activation.

    Directory of Open Access Journals (Sweden)

    Neel M Fofaria

    Full Text Available In this study, we determined the cytotoxic effects of piperine, a major constituent of black and long pepper in melanoma cells. Piperine treatment inhibited the growth of SK MEL 28 and B16 F0 cells in a dose and time-dependent manner. The growth inhibitory effects of piperine were mediated by cell cycle arrest of both the cell lines in G1 phase. The G1 arrest by piperine correlated with the down-regulation of cyclin D1 and induction of p21. Furthermore, this growth arrest by piperine treatment was associated with DNA damage as indicated by phosphorylation of H2AX at Ser139, activation of ataxia telangiectasia and rad3-related protein (ATR and checkpoint kinase 1 (Chk1. Pretreatment with AZD 7762, a Chk1 inhibitor not only abrogated the activation of Chk1 but also piperine mediated G1 arrest. Similarly, transfection of cells with Chk1 siRNA completely protected the cells from G1 arrest induced by piperine. Piperine treatment caused down-regulation of E2F1 and phosphorylation of retinoblastoma protein (Rb. Apoptosis induced by piperine was associated with down-regulation of XIAP, Bid (full length and cleavage of Caspase-3 and PARP. Furthermore, our results showed that piperine treatment generated ROS in melanoma cells. Blocking ROS by tiron protected the cells from piperine mediated cell cycle arrest and apoptosis. These results suggest that piperine mediated ROS played a critical role in inducing DNA damage and activation of Chk1 leading to G1 cell cycle arrest and apoptosis.

  12. Exposure to methyl tert-butyl ether, benzene, and total hydrocarbons at the Singapore-Malaysia causeway immigration checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Tan, C.; Ong, H.Y.; Kok, P.W. [and others

    1996-12-31

    The primary aim of this study was to determine the extent and levels of exposure to volatile organic compounds (VOCs) from automobile emissions in a group of immigration officers at a busy cross-border checkpoint. A majority (80%) of the workers monitored were exposed to benzene at levels between 0.01 and 0.5 ppm, with only 1.2% exceeding the current Occupational Safety and Health Administration occupational exposure limit of 1 ppm. The geometric mean (GM) concentrations of 8-hr time-weighted average exposure were 0.03 ppm, 0.9 ppm, and 2.46 ppm for methyl-tert-butyl ether (MTBE), benzene, and total hydrocarbons (THC), respectively. The highest time-weighted average concentrations measured were 1.05 ppm for MTBE, 2.01 ppm for benzene, and 34 ppm for THC. It was found that motorbikes emitted a more significant amount of pollutants compared with motor cars. On average, officers at the motorcycle booths were exposed to four to five times higher levels of VOCs (GMs of 0.07 ppm, 0.23 ppm, and 4.7 ppm for MTBE, benzene, and THC) than their counterparts at the motor car booths (GMs of 0.01 ppm, 0.05 ppm, and 1.5 ppm). The airborne concentrations of all three pollutants correlated with the flow of vehicle traffic. Close correlations were also noted for the concentrations in ambient air for the three pollutants measured. Benzene and MTBE had a correlation coefficient of 0.97. The overall findings showed that the concentrations of various VOCs were closely related to the traffic density, suggesting that they were from a common source, such as exhaust emissions from the vehicles. The results also indicated that although benzene, MTBE, and THC are known to be volatile, a significant amount could still be detected in the ambient environment, thus contributing to our exposure to these compounds. 4 refs., 6 figs.

  13. Differential genetic interactions between Sgs1, DNA-damage checkpoint components and DNA repair factors in the maintenance of chromosome stability

    Directory of Open Access Journals (Sweden)

    Doerfler Lillian

    2011-10-01

    Full Text Available Abstract Background Genome instability is associated with human cancers and chromosome breakage syndromes, including Bloom's syndrome, caused by inactivation of BLM helicase. Numerous mutations that lead to genome instability are known, yet how they interact genetically is poorly understood. Results We show that spontaneous translocations that arise by nonallelic homologous recombination in DNA-damage-checkpoint-defective yeast lacking the BLM-related Sgs1 helicase (sgs1Δ mec3Δ are inhibited if cells lack Mec1/ATR kinase. Tel1/ATM, in contrast, acts as a suppressor independently of Mec3 and Sgs1. Translocations are also inhibited in cells lacking Dun1 kinase, but not in cells defective in a parallel checkpoint branch defined by Chk1 kinase. While we had previously shown that RAD51 deletion did not inhibit translocation formation, RAD59 deletion led to inhibition comparable to the rad52Δ mutation. A candidate screen of other DNA metabolic factors identified Exo1 as a strong suppressor of chromosomal rearrangements in the sgs1Δ mutant, becoming even more important for chromosomal stability upon MEC3 deletion. We determined that the C-terminal third of Exo1, harboring mismatch repair protein binding sites and phosphorylation sites, is dispensable for Exo1's roles in chromosomal rearrangement suppression, mutation avoidance and resistance to DNA-damaging agents. Conclusions Our findings suggest that translocations between related genes can form by Rad59-dependent, Rad51-independent homologous recombination, which is independently suppressed by Sgs1, Tel1, Mec3 and Exo1 but promoted by Dun1 and the telomerase-inhibitor Mec1. We propose a model for the functional interaction between mitotic recombination and the DNA-damage checkpoint in the suppression of chromosomal rearrangements in sgs1Δ cells.

  14. Chk1 regulates the S phase checkpoint by coupling the physiological turnover and ionizing radiation-induced accelerated proteolysis of Cdc25A

    DEFF Research Database (Denmark)

    Sørensen, Claus Storgaard; Syljuåsen, Randi G; Falck, Jacob;

    2003-01-01

    Chk1 kinase coordinates cell cycle progression and preserves genome integrity. Here, we show that chemical or genetic ablation of human Chk1 triggered supraphysiological accumulation of the S phase-promoting Cdc25A phosphatase, prevented ionizing radiation (IR)-induced degradation of Cdc25A, and...... by a combined action of Chk1 and Chk2 kinases. Finally, phosphorylation of Chk1 by ATM was required to fully accelerate the IR-induced degradation of Cdc25A. Our results provide evidence that the mammalian S phase checkpoint functions via amplification of physiologically operating, Chk1-dependent...

  15. Mechanism-based screen for G1/S checkpoint activators identifies a selective activator of EIF2AK3/PERK signalling.

    Directory of Open Access Journals (Sweden)

    Simon R Stockwell

    Full Text Available Human cancers often contain genetic alterations that disable G1/S checkpoint control and loss of this checkpoint is thought to critically contribute to cancer generation by permitting inappropriate proliferation and distorting fate-driven cell cycle exit. The identification of cell permeable small molecules that activate the G1/S checkpoint may therefore represent a broadly applicable and clinically effective strategy for the treatment of cancer. Here we describe the identification of several novel small molecules that trigger G1/S checkpoint activation and characterise the mechanism of action for one, CCT020312, in detail. Transcriptional profiling by cDNA microarray combined with reverse genetics revealed phosphorylation of the eukaryotic initiation factor 2-alpha (EIF2A through the eukaryotic translation initiation factor 2-alpha kinase 3 (EIF2AK3/PERK as the mechanism of action of this compound. While EIF2AK3/PERK activation classically follows endoplasmic reticulum (ER stress signalling that sets off a range of different cellular responses, CCT020312 does not trigger these other cellular responses but instead selectively elicits EIF2AK3/PERK signalling. Phosphorylation of EIF2A by EIF2A kinases is a known means to block protein translation and hence restriction point transit in G1, but further supports apoptosis in specific contexts. Significantly, EIF2AK3/PERK signalling has previously been linked to the resistance of cancer cells to multiple anticancer chemotherapeutic agents, including drugs that target the ubiquitin/proteasome pathway and taxanes. Consistent with such findings CCT020312 sensitizes cancer cells with defective taxane-induced EIF2A phosphorylation to paclitaxel treatment. Our work therefore identifies CCT020312 as a novel small molecule chemical tool for the selective activation of EIF2A-mediated translation control with utility for proof-of-concept applications in EIF2A-centered therapeutic approaches, and as a chemical

  16. miR-424(322) reverses chemoresistance via T-cell immune response activation by blocking the PD-L1 immune checkpoint

    OpenAIRE

    Xu, Shaohua; Tao, Zhen; Hai, Bo; Liang, Huagen; SHI, YING; Tao WANG; Song, Wen; Chen, Yong; Ouyang, Jun; Chen, Jinhong; Kong, Fanfei; Dong, Yishan; Jiang, Shi-Wen; LI, WEIYONG; Ping WANG

    2016-01-01

    Immune checkpoint blockade of the inhibitory immune receptors PD-L1, PD-1 and CTLA-4 has emerged as a successful treatment strategy for several advanced cancers. Here we demonstrate that miR-424(322) regulates the PD-L1/PD-1 and CD80/CTLA-4 pathways in chemoresistant ovarian cancer. miR-424(322) is inversely correlated with PD-L1, PD-1, CD80 and CTLA-4 expression. High levels of miR-424(322) in the tumours are positively correlated with the progression-free survival of ovarian cancer patients...

  17. Two S-phase checkpoint systems, one involving the function of both BIME and Tyr15 phosphorylation of p34cdc2, inhibit NIMA and prevent premature mitosis.

    OpenAIRE

    Ye, X S; Fincher, R R; Tang, A.; O'Donnell, K; Osmani, S A

    1996-01-01

    We demonstrate that there are at least two S-phase checkpoint mechanisms controlling mitosis in Aspergillus. The first responds to the rate of DNA replication and inhibits mitosis via tyrosine phosphorylation of p34cdc2. Cells unable to tyrosine phosphorylate p34cdc2 are therefore viable but are unable to tolerate low levels of hydroxyurea and prematurely enter lethal mitosis when S-phase is slowed. However, if the NIMA mitosis-promoting kinase is inactivated then non-tyrosine-phosphorylated ...

  18. Hypomorphic bimA(APC3) alleles cause errors in chromosome metabolism that activate the DNA damage checkpoint blocking cytokinesis in Aspergillus nidulans.

    OpenAIRE

    Wolkow, T D; Mirabito, P.M.; Venkatram, S; Hamer, J. E.

    2000-01-01

    The Aspergillus nidulans sepI(+) gene has been implicated in the coordination of septation with nuclear division and cell growth. We find that the temperature-sensitive (ts) sepI1 mutation represents a novel allele of bimA(APC3), which encodes a conserved component of the anaphase-promoting complex/cyclosome (APC/C). We have characterized the septation, nuclear division, cell-cycle checkpoint defects, and DNA sequence alterations of sepI1 (renamed bimA10) and two other ts lethal bimA(APC3) al...

  19. In Silico Exploration of 1,7-Diazacarbazole Analogs as Checkpoint Kinase 1 Inhibitors by Using 3D QSAR, Molecular Docking Study, and Molecular Dynamics Simulations

    OpenAIRE

    Xiaodong Gao; Liping Han; Yujie Ren

    2016-01-01

    Checkpoint kinase 1 (Chk1) is an important serine/threonine kinase with a self-protection function. The combination of Chk1 inhibitors and anti-cancer drugs can enhance the selectivity of tumor therapy. In this work, a set of 1,7-diazacarbazole analogs were identified as potent Chk1 inhibitors through a series of computer-aided drug design processes, including three-dimensional quantitative structure–activity relationship (3D-QSAR) modeling, molecular docking, and molecular dynamics simulatio...

  20. Phosphorylation of the regulatory beta-subunit of protein kinase CK2 by checkpoint kinase Chk1: identification of the in vitro CK2beta phosphorylation site

    DEFF Research Database (Denmark)

    Kristensen, Lars P; Larsen, Martin Røssel; Højrup, Peter; Issinger, Olaf-Georg; Guerra, Barbara

    The regulatory beta-subunit of protein kinase CK2 mediates the formation of the CK2 tetrameric form and it has functions independent of CK2 catalytic subunit through interaction with several intracellular proteins. Recently, we have shown that CK2beta associates with the human checkpoint kinase Chk......1. In this study, we show that Chk1 specifically phosphorylates in vitro the regulatory beta-subunit of CK2. Chymotryptic peptides and mutational analyses have revealed that CK2beta is phosphorylated at Thr213. Formation of a stable complex between CK2beta and Chk1 is not affected by the...

  1. miR-424(322) reverses chemoresistance via T-cell immune response activation by blocking the PD-L1 immune checkpoint

    Science.gov (United States)

    Xu, Shaohua; Tao, Zhen; Hai, Bo; Liang, Huagen; Shi, Ying; Wang, Tao; Song, Wen; Chen, Yong; OuYang, Jun; Chen, Jinhong; Kong, Fanfei; Dong, Yishan; Jiang, Shi-Wen; Li, Weiyong; Wang, Ping; Yuan, Zhiyong; Wan, Xiaoping; Wang, Chenguang; Li, Wencheng; Zhang, Xiaoping; Chen, Ke

    2016-01-01

    Immune checkpoint blockade of the inhibitory immune receptors PD-L1, PD-1 and CTLA-4 has emerged as a successful treatment strategy for several advanced cancers. Here we demonstrate that miR-424(322) regulates the PD-L1/PD-1 and CD80/CTLA-4 pathways in chemoresistant ovarian cancer. miR-424(322) is inversely correlated with PD-L1, PD-1, CD80 and CTLA-4 expression. High levels of miR-424(322) in the tumours are positively correlated with the progression-free survival of ovarian cancer patients. Mechanistic investigations demonstrated that miR-424(322) inhibited PD-L1 and CD80 expression through direct binding to the 3′-untranslated region. Restoration of miR-424(322) expression reverses chemoresistance, which is accompanied by blockage of the PD-L1 immune checkpoint. The synergistic effect of chemotherapy and immunotherapy is associated with the proliferation of functional cytotoxic CD8+ T cells and the inhibition of myeloid-derived suppressive cells and regulatory T cells. Collectively, our data suggest a biological and functional interaction between PD-L1 and chemoresistance through the microRNA regulatory cascade. PMID:27147225

  2. RAG-mediated DNA double-strand breaks activate a cell type-specific checkpoint to inhibit pre-B cell receptor signals.

    Science.gov (United States)

    Bednarski, Jeffrey J; Pandey, Ruchi; Schulte, Emily; White, Lynn S; Chen, Bo-Ruei; Sandoval, Gabriel J; Kohyama, Masako; Haldar, Malay; Nickless, Andrew; Trott, Amanda; Cheng, Genhong; Murphy, Kenneth M; Bassing, Craig H; Payton, Jacqueline E; Sleckman, Barry P

    2016-02-01

    DNA double-strand breaks (DSBs) activate a canonical DNA damage response, including highly conserved cell cycle checkpoint pathways that prevent cells with DSBs from progressing through the cell cycle. In developing B cells, pre-B cell receptor (pre-BCR) signals initiate immunoglobulin light (Igl) chain gene assembly, leading to RAG-mediated DNA DSBs. The pre-BCR also promotes cell cycle entry, which could cause aberrant DSB repair and genome instability in pre-B cells. Here, we show that RAG DSBs inhibit pre-BCR signals through the ATM- and NF-κB2-dependent induction of SPIC, a hematopoietic-specific transcriptional repressor. SPIC inhibits expression of the SYK tyrosine kinase and BLNK adaptor, resulting in suppression of pre-BCR signaling. This regulatory circuit prevents the pre-BCR from inducing additional Igl chain gene rearrangements and driving pre-B cells with RAG DSBs into cycle. We propose that pre-B cells toggle between pre-BCR signals and a RAG DSB-dependent checkpoint to maintain genome stability while iteratively assembling Igl chain genes. PMID:26834154

  3. Kaposi sarcoma herpes virus latency associated nuclear antigen protein release the G2/M cell cycle blocks by modulating ATM/ATR mediated checkpoint pathway.

    Directory of Open Access Journals (Sweden)

    Amit Kumar

    Full Text Available The Kaposi's sarcoma-associated herpesvirus infects the human population and maintains latency stage of viral life cycle in a variety of cell types including cells of epithelial, mesenchymal and endothelial origin. The establishment of latent infection by KSHV requires the expression of an unique repertoire of genes among which latency associated nuclear antigen (LANA plays a critical role in the replication of the viral genome. LANA regulates the transcription of a number of viral and cellular genes essential for the survival of the virus in the host cell. The present study demonstrates the disruption of the host G2/M cell cycle checkpoint regulation as an associated function of LANA. DNA profile of LANA expressing human B-cells demonstrated the ability of this nuclear antigen in relieving the drug (Nocodazole induced G2/M checkpoint arrest. Caffeine suppressed nocodazole induced G2/M arrest indicating involvement of the ATM/ATR. Notably, we have also shown the direct interaction of LANA with Chk2, the ATM/ATR signalling effector and is responsible for the release of the G2/M cell cycle block.

  4. miR-424(322) reverses chemoresistance via T-cell immune response activation by blocking the PD-L1 immune checkpoint.

    Science.gov (United States)

    Xu, Shaohua; Tao, Zhen; Hai, Bo; Liang, Huagen; Shi, Ying; Wang, Tao; Song, Wen; Chen, Yong; OuYang, Jun; Chen, Jinhong; Kong, Fanfei; Dong, Yishan; Jiang, Shi-Wen; Li, Weiyong; Wang, Ping; Yuan, Zhiyong; Wan, Xiaoping; Wang, Chenguang; Li, Wencheng; Zhang, Xiaoping; Chen, Ke

    2016-01-01

    Immune checkpoint blockade of the inhibitory immune receptors PD-L1, PD-1 and CTLA-4 has emerged as a successful treatment strategy for several advanced cancers. Here we demonstrate that miR-424(322) regulates the PD-L1/PD-1 and CD80/CTLA-4 pathways in chemoresistant ovarian cancer. miR-424(322) is inversely correlated with PD-L1, PD-1, CD80 and CTLA-4 expression. High levels of miR-424(322) in the tumours are positively correlated with the progression-free survival of ovarian cancer patients. Mechanistic investigations demonstrated that miR-424(322) inhibited PD-L1 and CD80 expression through direct binding to the 3'-untranslated region. Restoration of miR-424(322) expression reverses chemoresistance, which is accompanied by blockage of the PD-L1 immune checkpoint. The synergistic effect of chemotherapy and immunotherapy is associated with the proliferation of functional cytotoxic CD8+ T cells and the inhibition of myeloid-derived suppressive cells and regulatory T cells. Collectively, our data suggest a biological and functional interaction between PD-L1 and chemoresistance through the microRNA regulatory cascade. PMID:27147225

  5. 面向异构并行计算系统的流水线式压缩检查点%Pipelined Compressed Checkpointing for Heterogeneous Systems

    Institute of Scientific and Technical Information of China (English)

    刘勇鹏; 王锋; 卢凯; 刘勇燕

    2012-01-01

    Checkpointing is an effective technique to improve the reliability of large scale parallel computing systems.Data compression is a promising technique to reduce the size of data to be saved in the files in the storage subsystem and the amount of data to go through the communication subsystem. However, compression causes a huge amount of time overhead. The time overhead is the main technical barrier of its practical usability. In this paper, we propose a parallel compressed checkpointing technique to reduce the time overhead of compression in heterogenous architectures. It integrates a number of optimization techniques, which include transmitting checkpointing data between host and GPU in buffered pipelines, aggregating file write operations, employing a pipelined parallel compression algorithm, and delegating compression operations to GPU,etc. The paper reports an implementation of the technique in the TH-1 system and the evaluation experiments with the system. The experiment data show that the technique is efficient and practically useable.%在大规模并行计算系统中,并行检查点触发大量结点同时保存计算状态,造成巨大文件存储空间开销,以及对通信和存储系统的巨大访问压力.数据压缩可以缩小检查点文件尺寸,从而降低存储空间开销以及对通信和存储系统的访问压力.但是,它也带来额外的压缩计算开销.本文针对异构并行计算系统,提出流水线式并行压缩检查点技术,采用一系列优化技术来降低压缩引入的计算延时,包括:流水线式双重写缓存队列、文件写操作的合并、GPU加速的流水压缩算法和GPU资源的多进程调度,等等.本文介绍了该技术在天河一号系统中的实现,并对所实现的检查点系统进行综合评测.实验数据表明该方法在大规模异构并行计算系统中是可行、高效、实用的.

  6. Genetic analysis of the spindle checkpoint genes san-1, mdf-2, bub-3 and the CENP-F homologues hcp-1 and hcp-2 in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Moore Landon L

    2008-02-01

    Full Text Available Abstract Background The spindle checkpoint delays the onset of anaphase until all sister chromatids are aligned properly at the metaphase plate. To investigate the role san-1, the MAD3 homologue, has in Caenorhabditis elegans embryos we used RNA interference (RNAi to identify genes synthetic lethal with the viable san-1(ok1580 deletion mutant. Results The san-1(ok1580 animal has low penetrating phenotypes including an increased incidence of males, larvae arrest, slow growth, protruding vulva, and defects in vulva morphogenesis. We found that the viability of san-1(ok1580 embryos is significantly reduced when HCP-1 (CENP-F homologue, MDF-1 (MAD-1 homologue, MDF-2 (MAD-2 homologue or BUB-3 (predicted BUB-3 homologue are reduced by RNAi. Interestingly, the viability of san-1(ok1580 embryos is not significantly reduced when the paralog of HCP-1, HCP-2, is reduced. The phenotype of san-1(ok1580;hcp-1(RNAi embryos includes embryonic and larval lethality, abnormal organ development, and an increase in abnormal chromosome segregation (aberrant mitotic nuclei, anaphase bridging. Several of the san-1(ok1580;hcp-1(RNAi animals displayed abnormal kinetochore (detected by MPM-2 and microtubule structure. The survival of mdf-2(RNAi;hcp-1(RNAi embryos but not bub-3(RNAi;hcp-1(RNAi embryos was also compromised. Finally, we found that san-1(ok1580 and bub-3(RNAi, but not hcp-1(RNAi embryos, were sensitive to anoxia, suggesting that like SAN-1, BUB-3 has a functional role as a spindle checkpoint protein. Conclusion Together, these data suggest that in the C. elegans embryo, HCP-1 interacts with a subset of the spindle checkpoint pathway. Furthermore, the fact that san-1(ok1580;hcp-1(RNAi animals had a severe viability defect whereas in the san-1(ok1580;hcp-2(RNAi and san-1(ok1580;hcp-2(ok1757 animals the viability defect was not as severe suggesting that hcp-1 and hcp-2 are not completely redundant.

  7. Loss of p53-regulatory protein IFI16 induces NBS1 leading to activation of p53-mediated checkpoint by phosphorylation of p53 SER37.

    Science.gov (United States)

    Tawara, Hideyuki; Fujiuchi, Nobuko; Sironi, Juan; Martin, Sarah; Aglipay, Jason; Ouchi, Mutsuko; Taga, Makoto; Chen, Phang-Lang; Ouchi, Toru

    2008-01-01

    Our previous results that IFI16 is involved in p53 transcription activity under conditions of ionizing radiation (IR), and that the protein is frequently lost in human breast cancer cell lines and breast adenocarcinoma tissues suggesting that IFI16 plays a crucial role in controlling cell growth. Here, we show that loss of IFI16 by RNA interference in cell culture causes elevated phosphorylation of p53 Ser37 and accumulated NBS1 (nibrin) and p21WAF1, leading to growth retardation. Consistent with these observations, doxycyclin-induced NBS1 caused accumulation of p21WAF1 and increased phosphorylation of p53 Ser37, leading to cell cycle arrest in G1 phase. Wortmannin treatment was found to decrease p53 Ser37 phosphorylation in NBS-induced cells. These results suggest that loss of IFI16 activates p53 checkpoint through NBS1-DNA-PKcs pathway. PMID:17981542

  8. The regulatory beta-subunit of protein kinase CK2 accelerates the degradation of CDC25A phosphatase through the checkpoint kinase Chk1

    DEFF Research Database (Denmark)

    Kreutzer, Jan Nicolas; Guerra, Barbara

    2007-01-01

    Human CDC25 phosphatases play an important role in cell cycle regulation by removing inhibitory phosphate groups on cyclin-CDKs. Chk1 has been shown to phosphorylate CDC25 family members down-regulating their phosphatase activity through distinct mechanisms. The kinase activity of Chk1 is evident...... cell cycle progression is shown to enhance CDC25A degradation, and this occurs in a manner similar to that by which CDC25A is down-regulated upon activation of cellular checkpoint responses. By using RNA interference to specifically deplete cells of Chk1, we demonstrate that Chk1 mediates the down-regulation...... cell cycle regulation and indicate the mechanism by which CDC25A turnover might be regulated by Chk1 in the absence of DNA damage....

  9. Inhibition of clathrin by pitstop 2 activates the spindle assembly checkpoint and induces cell death in dividing HeLa cancer cells

    Directory of Open Access Journals (Sweden)

    Smith Charlotte M

    2013-01-01

    Full Text Available Abstract Background During metaphase clathrin stabilises the mitotic spindle kinetochore(K-fibres. Many anti-mitotic compounds target microtubule dynamics. Pitstop 2™ is the first small molecule inhibitor of clathrin terminal domain and inhibits clathrin-mediated endocytosis. We investigated its effects on a second function for clathrin in mitosis. Results Pitstop 2 did not impair clathrin recruitment to the spindle but disrupted its function once stationed there. Pitstop 2 trapped HeLa cells in metaphase through loss of mitotic spindle integrity and activation of the spindle assembly checkpoint, phenocopying clathrin depletion and aurora A kinase inhibition. Conclusions Pitstop 2 is therefore a new tool for investigating clathrin spindle dynamics. Pitstop 2 reduced viability in dividing HeLa cells, without affecting dividing non-cancerous NIH3T3 cells, suggesting that clathrin is a possible novel anti-mitotic drug target.

  10. 面向大规模MPI程序的应用级checkpointing技术%Application-Level Checkpointing for Large-Scale MPI Programs

    Institute of Scientific and Technical Information of China (English)

    王攀峰; 杜云飞; 周海芳; 杨学军

    2009-01-01

    应用级checkpointing是一种在大规模科学计算领域中备受关注的容错技术.但是应用级checkpointing技术要求用户决定哪些是需要保存的关键数据,这增加了用户的负担.介绍一个基于MPI并行程序活跃变量分析的源到源的预编译工具ALEC,它可用于辅助应用级checkpointing.在一个512处理器的Cluster系统上,对经过ALEC编译的5个Fortran/MPI应用进行了性能评测.结果表明,ALEC能够有效减小checkpoint的大小和应用级checkpointing保存和恢复的开销.

  11. Oxidative stress-induced DNA damage of mouse zygotes triggers G2/M checkpoint and phosphorylates Cdc25 and Cdc2.

    Science.gov (United States)

    Zhang, Yuting; Qian, Diting; Li, Zhiling; Huang, Yue; Wu, Que; Ru, Gaizhen; Chen, Man; Wang, Bin

    2016-07-01

    In vitro fertilized (IVF) embryos show both cell cycle and developmental arrest. We previously showed oxidative damage activates the ATM → Chk1 → Cdc25B/Cdc25C cascade to mediate G2/M cell cycle arrest for repair of hydrogen peroxide (H2O2)-induced oxidative damage in sperm. However, the mechanisms underlying the developmental delay of zygotes are unknown. To develop a model of oxidative-damaged zygotes, we treated mouse zygotes with different concentrations of H2O2 (0, 0.01, 0.02, 0.03, 0.04, 0.05 mM), and evaluated in vitro zygote development, BrdU incorporation to detect the duration of S phase. We also examined reactive oxygen species level and used immunofluorescence to detect activation of γH2AX, Cdc2, and Cdc25. Oxidatively damaged zygotes showed a delay in G2/M phase and produced a higher level of ROS. At the same time, γH2AX was detected in oxidatively damaged zygotes as well as phospho-Cdc25B (Ser323), phospho-Cdc25C (Ser216), and phospho-Cdc2 (Tyr15). Our study indicates that oxidative stress-induced DNA damage of mouse zygotes triggers the cell cycle checkpoint, which results in G2/M cell cycle arrest, and that phospho-Cdc25B (Ser323), phospho-Cdc25C (Ser216), and phospho-Cdc2 (Tyr15) participate in activating the G2/M checkpoint. PMID:27117522

  12. The involvement of cell cycle checkpoint-mutations in the mutagenesis induced in Drosophila by a longer wavelength light band of solar UV.

    Science.gov (United States)

    Toyoshima, Megumi; Takinami, Syogo; Hieda, Kotaro; Fursawa, Yoshiya; Negishi, Tomoe

    2002-03-01

    Solar ultraviolet radiation is considered to be injurious rather than necessary for most organisms living on the earth. It is reported that the risk of skin cancer in humans increases by the depletion of the ozone layer. We have examined the genotoxicity of solar ultraviolet, especially the longer wavelength light, using Drosophila. Recently, we have demonstrated that light of wavelength up to 340 nm is mutagenic on Drosophila larvae. Using an excision repair-deficient Drosophila strain (mus201), we have obtained results suggesting that the lesion caused in larvae by the 320 nm-light irradiation may be similar to the damage induced by irradiation at 310 nm, and that light of 330 and 340 nm may induce damage different from that induced by 310 and 320 nm-light. To examine the difference in DNA damage induced by light of a particular wavelength, we performed monochromatic irradiation on larvae of two Drosophila strains; one excision repair-deficient (mei-9) and another postreplication repair-deficient (mei-41). 310 and 320 nm-light was more mutagenic in the mei-9 strain than in mei-41, whereas 330 and 340 nm-light was more mutagenic in mei-41 than in mei-9. It is demonstrated that the mei-41 gene is a homologue of the human atm gene which is responsible for a cell cycle checkpoint. This result suggests that 310-320 nm-light induces DNA damage that is subject to nucleotide excision repair (NER) and that 330-360 nm-light causes damage to be recognized by the cell cycle checkpoint but it is not repairable by NER. PMID:12659514

  13. When the genome plays dice: circumvention of the spindle assembly checkpoint and near-random chromosome segregation in multipolar cancer cell mitoses.

    Directory of Open Access Journals (Sweden)

    David Gisselsson

    Full Text Available BACKGROUND: Normal cell division is coordinated by a bipolar mitotic spindle, ensuring symmetrical segregation of chromosomes. Cancer cells, however, occasionally divide into three or more directions. Such multipolar mitoses have been proposed to generate genetic diversity and thereby contribute to clonal evolution. However, this notion has been little validated experimentally. PRINCIPAL FINDINGS: Chromosome segregation and DNA content in daughter cells from multipolar mitoses were assessed by multiphoton cross sectioning and fluorescence in situ hybridization in cancer cells and non-neoplastic transformed cells. The DNA distribution resulting from multipolar cell division was found to be highly variable, with frequent nullisomies in the daughter cells. Time-lapse imaging of H2B/GFP-labelled multipolar mitoses revealed that the time from the initiation of metaphase to the beginning of anaphase was prolonged and that the metaphase plates often switched polarity several times before metaphase-anaphase transition. The multipolar metaphase-anaphase transition was accompanied by a normal reduction of cellular cyclin B levels, but typically occurred before completion of the normal separase activity cycle. Centromeric AURKB and MAD2 foci were observed frequently to remain on the centromeres of multipolar ana-telophase chromosomes, indicating that multipolar mitoses were able to circumvent the spindle assembly checkpoint with some sister chromatids remaining unseparated after anaphase. Accordingly, scoring the distribution of individual chromosomes in multipolar daughter nuclei revealed a high frequency of nondisjunction events, resulting in a near-binomial allotment of sister chromatids to the daughter cells. CONCLUSION: The capability of multipolar mitoses to circumvent the spindle assembly checkpoint system typically results in a near-random distribution of chromosomes to daughter cells. Spindle multipolarity could thus be a highly efficient

  14. Induction of chromosome instability and stomach cancer by altering the expression pattern of mitotic checkpoint genes in mice exposed to areca-nut

    International Nuclear Information System (INIS)

    There are strong indications for a causal association between areca-nut consumption and cancers. In Meghalaya, India, the variety of areca-nut is used as raw and unprocessed form whose chemical composition and pharmacological actions have been reported. Yet we know little on the initial pathway involved in areca-nut associated carcinogenesis since it is difficult to assess its effects on genetic alterations without interference of other compounding factors. Therefore, present study was undertaken in mice to verify the ability of raw areca-nut (RAN) to induce cancer and to monitor the expression of certain genes involved in carcinogenesis. This study was not intended to isolate any active ingredients from the RAN and to look its action. Three groups of mice (n = 25 in each) were taken and used at different time-points for different experimental analysis. The other three groups of mice (n = 15 in each) were considered for tumor induction studies. In each set, two groups were administered RAN-extract ad libitum in drinking water with or without lime. The expression of certain genes was assessed by conventional RT-PCR and immunoblotting. The mice were given the whole RAN-extract with and without lime in order to mimic the human consumption style of RAN. Histological preparation of stomach tissue revealed that RAN induced stomach cancer. A gradual increase in the frequency of precocious anaphase and aneuploid cells was observed in the bone marrow cells with a greater increment following RAN + lime administeration. Levels of p53, Bax, Securin and p65 in esophageal and stomach cells were elevated during early days of RAN exposure while those of different mitotic checkpoint proteins were downregulated. Apoptotic cell death was detected in non-cancerous stomach cells but not in tumor cells which showed overexpression of Bax and absence of PARP. Present study suggested (a) RAN induces stomach cancer, however, presence of lime promoted higher cell transformation and thereby

  15. Radiation resistance due to high expression of miR-21 and G2/M checkpoint arrest in breast cancer cells

    International Nuclear Information System (INIS)

    There is evidence that the extent of the G2/M arrest following irradiation is correlated with tumour cell survival and hence therapeutic success. We studied the regulation of cellular response to radiation treatment by miR-21-mediated modulation of cell cycle progression in breast cancer cells and analysed miR-21 expression in breast cancer tissue samples with long-term follow up. The miR-21 expression levels were quantified (qRT-PCR) in a panel of 86 cases of invasive breast carcinomas in relation to metastasis free survival. The cellular radiosensitivity of human breast cancer cells after irradiation was determined comparing two cell lines (T47D and MDA-MB-361) by cell proliferation and colony forming assays. The influence of miR-21 overexpression or downregulation on cell cycle progression and G2/M checkpoint arrest after irradiation was assessed by flow cytometric analysis. The expression of miR-21 was transiently increased 8 hours after irradiation in the radioresistant T47D cells and significantly changed with lower extent in radiosensitive MDA-MB-361 cells. Anti-miR-21 treated breast cancer cells failed to exhibit the DNA damage-G2 checkpoint increase after irradiation. Apoptotic activity was significantly enhanced from 7% to 27% in T47D cells and from 18% to 30% in MDA-MB-361 cells 24 hours after 5 Gy irradiation. Additionally, we characterized expression of miR-21 in invasive breast carcinomas. In comparison to non-cancerous adjacent breast tissue, tumours samples had increased miR-21 expression that inversely correlated with the distant metastases-free survival of patients (p = 0.029). Our data indicate that miR-21 expression in breast cancer cells contributes to radiation resistance by compromising cell cycle progression. These data point to the potential of combining radiotherapy with an anti-miR-21 as a potent G2/M check point inhibitor in overcoming radiation resistance of tumours

  16. Targeting immune checkpoints in unresectable metastatic cutaneous melanoma: a systematic review and meta-analysis of anti-CTLA-4 and anti-PD-1 agents trials.

    Science.gov (United States)

    Yun, Seongseok; Vincelette, Nicole D; Green, Myke R; Wahner Hendrickson, Andrea E; Abraham, Ivo

    2016-07-01

    Anti-cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) and anti-programmed cell death-1 (PD-1) inhibitors have been shown to significantly improve survival in patients with metastatic cutaneous melanoma. However, there was some heterogeneity as well as some variation in the degree of benefit across studies. We reviewed randomized trials and performed a meta-analysis to determine the efficacy and safety of immune checkpoint inhibitors in comparison with conventional regimens. Eligible studies were limited to randomized controlled trials comparing anti-CTLA-4 or anti-PD-1 inhibitors to chemotherapy or vaccination treatment in adult patients with unresectable cutaneous metastatic melanoma. Progression-free survival (PFS) rate at 6 months was 28.5% versus 17.7% (RR: 0.84, 95% CI: 0.76-0.93), overall survival (OS) rate at 1 year was 51.2% versus 38.8% (RR: 0.72, 95% CI: 0.59-0.88), and overall response rate (ORR) at 6 months was 29.6% versus 17.7% (RR: 0.85, 95% CI: 0.76-0.95) favoring immune check point inhibitors over chemotherapies or vaccination. Immune check point inhibitors were associated with more frequent immune-related adverse events at 13.7% versus 2.4% of treated patients (RR: 6.74, 95% CI: 4.65-9.75). Subgroup analyses demonstrated significant PFS (RR: 0.92 vs. 0.74, P < 0.00001) and ORR (RR: 0.95 vs. 0.76, P = 0.0004) improvement with anti-PD-1 treatment compared to anti-CTLA-4 when each of them was compared to control treatments. Collectively, these results demonstrate that immune checkpoint inhibitors have superior outcomes compared to conventional chemotherapies or vaccination, and support the results of recent randomized trials that showed superior outcomes with anti-PD-1 agents over ipilimumab in unresectable metastatic cutaneous melanoma patients. PMID:27167347

  17. Radiation resistance due to high expression of miR-21 and G2/M checkpoint arrest in breast cancer cells

    Directory of Open Access Journals (Sweden)

    Anastasov Nataša

    2012-12-01

    Full Text Available Abstract Background There is evidence that the extent of the G2/M arrest following irradiation is correlated with tumour cell survival and hence therapeutic success. We studied the regulation of cellular response to radiation treatment by miR-21-mediated modulation of cell cycle progression in breast cancer cells and analysed miR-21 expression in breast cancer tissue samples with long-term follow up. Methods The miR-21 expression levels were quantified (qRT-PCR in a panel of 86 cases of invasive breast carcinomas in relation to metastasis free survival. The cellular radiosensitivity of human breast cancer cells after irradiation was determined comparing two cell lines (T47D and MDA-MB-361 by cell proliferation and colony forming assays. The influence of miR-21 overexpression or downregulation on cell cycle progression and G2/M checkpoint arrest after irradiation was assessed by flow cytometric analysis. Results The expression of miR-21 was transiently increased 8 hours after irradiation in the radioresistant T47D cells and significantly changed with lower extent in radiosensitive MDA-MB-361 cells. Anti-miR-21 treated breast cancer cells failed to exhibit the DNA damage-G2 checkpoint increase after irradiation. Apoptotic activity was significantly enhanced from 7% to 27% in T47D cells and from 18% to 30% in MDA-MB-361 cells 24 hours after 5 Gy irradiation. Additionally, we characterized expression of miR-21 in invasive breast carcinomas. In comparison to non-cancerous adjacent breast tissue, tumours samples had increased miR-21 expression that inversely correlated with the distant metastases-free survival of patients (p = 0.029. Conclusions Our data indicate that miR-21 expression in breast cancer cells contributes to radiation resistance by compromising cell cycle progression. These data point to the potential of combining radiotherapy with an anti-miR-21 as a potent G2/M check point inhibitor in overcoming radiation resistance of tumours.

  18. 一种基于移动计算的非阻塞协同检查点算法%A Non-blocking Coordinated Checkpoint Algorithm for Mobile Computing Environment

    Institute of Scientific and Technical Information of China (English)

    姚建盛; 刘艳玲

    2011-01-01

    传统分布式网络中协同检查点算法有许多优点,但移动计算环境有很多新问题,如缺少可靠存储、低无线网络带宽、高移动性、断开连接和能量限制等,使得协同检查点算法不能很好地适用于移动计算环境.为此提出一种适合移动计算环境的非阻塞的协同检查点算法,算法中检查点的发起和同步由移动支持站完成,只需记录部分中途消息就能实现一致可恢复.算法对移动主机是透明的,低开销的.%Although there are many advantages for coordinated checkpoint, mobile computing environment raises many new issues such as lack of stable storage, low bandwidth of wireless channel, high mobility, frequent disconnection and limited energy. These make the coordinated checkpoint algorithms unsuitable for mobile computing.This paper proposes a non-blocking coordinated checkpoint algorithm, the startup and synchronization of the coordinated checkpoint are completed by mobile support station. The algorithm is transparent for mobile host, and low-overheard and consistent rollback-recovery only save part of in-transit message.

  19. Mismatch repair-dependent G2 checkpoint induced by low doses of SN1 type methylating agents requires the ATR kinase.

    Science.gov (United States)

    Stojic, Lovorka; Mojas, Nina; Cejka, Petr; Di Pietro, Massimiliano; Ferrari, Stefano; Marra, Giancarlo; Jiricny, Josef

    2004-06-01

    S(N)1-type alkylating agents represent an important class of chemotherapeutics, but the molecular mechanisms underlying their cytotoxicity are unknown. Thus, although these substances modify predominantly purine nitrogen atoms, their toxicity appears to result from the processing of O(6)-methylguanine ((6Me)G)-containing mispairs by the mismatch repair (MMR) system, because cells with defective MMR are highly resistant to killing by these agents. In an attempt to understand the role of the MMR system in the molecular transactions underlying the toxicity of alkylating agents, we studied the response of human MMR-proficient and MMR-deficient cells to low concentrations of the prototypic methylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). We now show that MNNG treatment induced a cell cycle arrest that was absolutely dependent on functional MMR. Unusually, the cells arrested only in the second G(2) phase after treatment. Downstream targets of both ATM (Ataxia telangiectasia mutated) and ATR (ATM and Rad3-related) kinases were modified, but only the ablation of ATR, or the inhibition of CHK1, attenuated the arrest. The checkpoint activation was accompanied by the formation of nuclear foci containing the signaling and repair proteins ATR, the S(*)/T(*)Q substrate, gamma-H2AX, and replication protein A (RPA). The persistence of these foci implied that they may represent sites of irreparable damage. PMID:15175264

  20. Immune re-activation by cell-free fetal DNA in healthy pregnancies re-purposed to target tumors: novel check-point inhibition in cancer therapeutics

    Directory of Open Access Journals (Sweden)

    Elizabeth Ann Lieser Enninga

    2015-08-01

    Full Text Available The role of the immune system in cancer progression has become increasingly evident over the past decade. Chronic inflammation in the promotion of tumorigenesis is well established, and cancer-associated tolerance/immune evasion has long been appreciated. Recent developments of immunotherapies targeting cancer-associated inflammation and immune tolerance such as cancer vaccines, cell therapies, neutralizing antibodies, and immune checkpoint inhibitors, have shown promising clinical results. However, despite significant therapeutic advances, most patients diagnosed with metastatic cancer still succumb to their malignancy. Treatments are often toxic, and the financial burden of novel therapies is significant. Thus, new methods for utilizing similar biological systems to compare complex biological processes can give us new hypotheses for combating cancer. One such approach is comparing trophoblastic growth and regulation to tumor invasion and immune escape. Novel concepts regarding immune activation in pregnancy, especially reactivation of the immune system at labor through toll like receptor engagement by fetal derived DNA, may be applicable to cancer immunotherapy. This review summarizes mechanisms of inflammation in cancer, current immunotherapies used in the clinic, and suggestions for looking beyond oncology for novel methods to reverse cancer-associated tolerance and immunologic exhaustion utilizing mechanisms encountered in normal human pregnancy.

  1. Tumor genotype and immune microenvironment in POLE-ultramutated and MSI-hypermutated Endometrial Cancers: New candidates for checkpoint blockade immunotherapy?

    Science.gov (United States)

    Gargiulo, Piera; Della Pepa, Chiara; Berardi, Simona; Califano, Daniela; Scala, Stefania; Buonaguro, Luigi; Ciliberto, Gennaro; Brauchli, Peter; Pignata, Sandro

    2016-07-01

    Endometrial Cancer (EC) is still a challenge for gynecological oncologists because the treatment of the advanced disease remains an unmet need for patients. The Cancer Genome Atlas Research Network (TCGA) recently provided a comprehensive genomic and transcriptomic analysis of EC, offering a new classification of the disease, based on genetic features, which defines four subgroups of cancer rather than the two traditionally recognized. In the molecular classification two types of EC, the polymerase epsilon (POLE)-ultramutated and the microsatellite instability (MSI)-hypermutated, seem to present an enhanced immune microenvironment and a high mutation burden. The blockade of the immune checkpoints is an innovative approach that has largely demonstrated to be effective in solid malignancies, such as lung, renal and melanoma; it acts by reducing the cancer-induced immune-suppression through inhibition of the PD-1/PD-L1 (Programmed Death and PD-Ligand) axis. All available evidence supporting an over-expression of the PD-1/PD-L1 pathway in EC has been reviewed. In particular in the POLE and MSI ECs an up-regulation of this pathway was found, aiming to suggest a rationale for testing the PD-1/PD-L1 immunotherapy in these cancer subgroups. PMID:27362548

  2. In Silico Exploration of 1,7-Diazacarbazole Analogs as Checkpoint Kinase 1 Inhibitors by Using 3D QSAR, Molecular Docking Study, and Molecular Dynamics Simulations.

    Science.gov (United States)

    Gao, Xiaodong; Han, Liping; Ren, Yujie

    2016-01-01

    Checkpoint kinase 1 (Chk1) is an important serine/threonine kinase with a self-protection function. The combination of Chk1 inhibitors and anti-cancer drugs can enhance the selectivity of tumor therapy. In this work, a set of 1,7-diazacarbazole analogs were identified as potent Chk1 inhibitors through a series of computer-aided drug design processes, including three-dimensional quantitative structure-activity relationship (3D-QSAR) modeling, molecular docking, and molecular dynamics simulations. The optimal QSAR models showed significant cross-validated correlation q² values (0.531, 0.726), fitted correlation r² coefficients (higher than 0.90), and standard error of prediction (less than 0.250). These results suggested that the developed models possess good predictive ability. Moreover, molecular docking and molecular dynamics simulations were applied to highlight the important interactions between the ligand and the Chk1 receptor protein. This study shows that hydrogen bonding and electrostatic forces are key interactions that confer bioactivity. PMID:27164065

  3. Immune Reactivation by Cell-Free Fetal DNA in Healthy Pregnancies Re-Purposed to Target Tumors: Novel Checkpoint Inhibition in Cancer Therapeutics

    Science.gov (United States)

    Enninga, Elizabeth Ann L.; Nevala, Wendy K.; Holtan, Shernan G.; Markovic, Svetomir N.

    2015-01-01

    The role of the immune system in cancer progression has become increasingly evident over the past decade. Chronic inflammation in the promotion of tumorigenesis is well established, and cancer-associated tolerance/immune evasion has long been appreciated. Recent developments of immunotherapies targeting cancer-associated inflammation and immune tolerance, such as cancer vaccines, cell therapies, neutralizing antibodies, and immune checkpoint inhibitors, have shown promising clinical results. However, despite significant therapeutic advances, most patients diagnosed with metastatic cancer still succumb to their malignancy. Treatments are often toxic, and the financial burden of novel therapies is significant. Thus, new methods for utilizing similar biological systems to compare complex biological processes can give us new hypotheses for combating cancer. One such approach is comparing trophoblastic growth and regulation to tumor invasion and immune escape. Novel concepts regarding immune activation in pregnancy, especially reactivation of the immune system at labor through toll like receptor engagement by fetal derived DNA, may be applicable to cancer immunotherapy. This review summarizes mechanisms of inflammation in cancer, current immunotherapies used in the clinic, and suggestions for looking beyond oncology for novel methods to reverse cancer-associated tolerance and immunologic exhaustion utilizing mechanisms encountered in normal human pregnancy. PMID:26379664

  4. In Silico Exploration of 1,7-Diazacarbazole Analogs as Checkpoint Kinase 1 Inhibitors by Using 3D QSAR, Molecular Docking Study, and Molecular Dynamics Simulations

    Directory of Open Access Journals (Sweden)

    Xiaodong Gao

    2016-05-01

    Full Text Available Checkpoint kinase 1 (Chk1 is an important serine/threonine kinase with a self-protection function. The combination of Chk1 inhibitors and anti-cancer drugs can enhance the selectivity of tumor therapy. In this work, a set of 1,7-diazacarbazole analogs were identified as potent Chk1 inhibitors through a series of computer-aided drug design processes, including three-dimensional quantitative structure–activity relationship (3D-QSAR modeling, molecular docking, and molecular dynamics simulations. The optimal QSAR models showed significant cross-validated correlation q2 values (0.531, 0.726, fitted correlation r2 coefficients (higher than 0.90, and standard error of prediction (less than 0.250. These results suggested that the developed models possess good predictive ability. Moreover, molecular docking and molecular dynamics simulations were applied to highlight the important interactions between the ligand and the Chk1 receptor protein. This study shows that hydrogen bonding and electrostatic forces are key interactions that confer bioactivity.

  5. Large-Scale Profiling of Kinase Dependencies in Cancer Cell Lines

    Science.gov (United States)

    Campbell, James; Ryan, Colm J.; Brough, Rachel; Bajrami, Ilirjana; Pemberton, Helen N.; Chong, Irene Y.; Costa-Cabral, Sara; Frankum, Jessica; Gulati, Aditi; Holme, Harriet; Miller, Rowan; Postel-Vinay, Sophie; Rafiq, Rumana; Wei, Wenbin; Williamson, Chris T.; Quigley, David A.; Tym, Joe; Al-Lazikani, Bissan; Fenton, Timothy; Natrajan, Rachael; Strauss, Sandra J.; Ashworth, Alan; Lord, Christopher J.

    2016-01-01

    Summary One approach to identifying cancer-specific vulnerabilities and therapeutic targets is to profile genetic dependencies in cancer cell lines. Here, we describe data from a series of siRNA screens that identify the kinase genetic dependencies in 117 cancer cell lines from ten cancer types. By integrating the siRNA screen data with molecular profiling data, including exome sequencing data, we show how vulnerabilities/genetic dependencies that are associated with mutations in specific cancer driver genes can be identified. By integrating additional data sets into this analysis, including protein-protein interaction data, we also demonstrate that the genetic dependencies associated with many cancer driver genes form dense connections on functional interaction networks. We demonstrate the utility of this resource by using it to predict the drug sensitivity of genetically or histologically defined subsets of tumor cell lines, including an increased sensitivity of osteosarcoma cell lines to FGFR inhibitors and SMAD4 mutant tumor cells to mitotic inhibitors. PMID:26947069

  6. Leptin stimulates pituitary prolactin release through an extracellular signal-regulated kinase-dependent pathway

    DEFF Research Database (Denmark)

    Tipsmark, Christian K; Strom, Christina N; Bailey, Sean T;

    2008-01-01

    Leptin was initially identified as a regulator of appetite and weight control centers in the hypothalamus, but appears to be involved in a number of physiological processes. This study was carried out to examine the possible role of leptin in regulating prolactin (PRL) release using the teleost...... pituitary model system. This advantageous system allows isolation of a nearly pure population of lactotropes in their natural, in situ aggregated state. The rostral pars distalis were dissected from tilapia pituitaries and exposed to varying concentrations of leptin (0, 1, 10, 100 nM) for 1 h. Release of...... PRL was stimulated by leptin in a potent and concentration-dependent manner. A time-course experiment showed that the strongest response in PRL release with leptin occurs within the first hour (approximately sixfold), and stimulation was sustained after 16 h (approximately twofold). Many of the...

  7. Acanthamoeba castellanii Induces Host Cell Death via a Phosphatidylinositol 3-Kinase-Dependent Mechanism

    Science.gov (United States)

    Sissons, James; Kim, Kwang Sik; Stins, Monique; Jayasekera, Samantha; Alsam, Selwa; Khan, Naveed Ahmed

    2005-01-01

    Granulomatous amoebic encephalitis due to Acanthamoeba castellanii is a serious human infection with fatal consequences, but it is not clear how the circulating amoebae interact with the blood-brain barrier and transmigrate into the central nervous system. We studied the effects of an Acanthamoeba encephalitis isolate belonging to the T1 genotype on human brain microvascular endothelial cells, which constitute the blood-brain barrier. Using an apoptosis-specific enzyme-linked immunosorbent assay, we showed that Acanthamoeba induces programmed cell death in brain microvascular endothelial cells. Next, we observed that Acanthamoeba specifically activates phosphatidylinositol 3-kinase. Acanthamoeba-mediated brain endothelial cell death was abolished using LY294002, a phosphatidylinositol 3-kinase inhibitor. These results were further confirmed using brain microvascular endothelial cells expressing dominant negative forms of phosphatidylinositol 3-kinase. This is the first demonstration that Acanthamoeba-mediated brain microvascular endothelial cell death is dependent on phosphatidylinositol 3-kinase. PMID:15845472

  8. Acanthamoeba castellanii induces host cell death via a phosphatidylinositol 3-kinase-dependent mechanism

    OpenAIRE

    Sissons, James; Kim, Kwang Sik; Stins, Monique; Jayasekera, Samantha; Alsam, Selwa; Khan, Naveed Ahmed

    2005-01-01

    Granulomatous amoebic encephalitis due to Acanthamoeba castellanii is a serious human infection with fatal consequences, but it is not clear how the circulating amoebae interact with the blood-brain barrier and transmigrate into the central nervous system. We studied the effects of an Acanthamoeba encephalitis isolate belonging to the T1 genotype on human brain microvascular endothelial cells, which constitute the blood-brain barrier. Using an apoptosis-specific enzyme-linked immunosorbent as...

  9. Large-Scale Profiling of Kinase Dependencies in Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    James Campbell

    2016-03-01

    Full Text Available One approach to identifying cancer-specific vulnerabilities and therapeutic targets is to profile genetic dependencies in cancer cell lines. Here, we describe data from a series of siRNA screens that identify the kinase genetic dependencies in 117 cancer cell lines from ten cancer types. By integrating the siRNA screen data with molecular profiling data, including exome sequencing data, we show how vulnerabilities/genetic dependencies that are associated with mutations in specific cancer driver genes can be identified. By integrating additional data sets into this analysis, including protein-protein interaction data, we also demonstrate that the genetic dependencies associated with many cancer driver genes form dense connections on functional interaction networks. We demonstrate the utility of this resource by using it to predict the drug sensitivity of genetically or histologically defined subsets of tumor cell lines, including an increased sensitivity of osteosarcoma cell lines to FGFR inhibitors and SMAD4 mutant tumor cells to mitotic inhibitors.

  10. Leptin augments coronary vasoconstriction and smooth muscle proliferation via a Rho-kinase-dependent pathway.

    Science.gov (United States)

    Noblet, Jillian N; Goodwill, Adam G; Sassoon, Daniel J; Kiel, Alexander M; Tune, Johnathan D

    2016-05-01

    Leptin has been implicated as a key upstream mediator of pathways associated with coronary vascular dysfunction and disease. The purpose of this investigation was to test the hypothesis that leptin modifies the coronary artery proteome and promotes increases in coronary smooth muscle contraction and proliferation via influences on Rho kinase signaling. Global proteomic assessment of coronary arteries from lean swine cultured with obese concentrations of leptin (30 ng/mL) for 3 days revealed significant alterations in the coronary artery proteome (68 proteins) and identified an association between leptin treatment and calcium signaling/contraction (four proteins) and cellular growth and proliferation (35 proteins). Isometric tension studies demonstrated that both acute (30 min) and chronic (3 days, serum-free media) exposure to obese concentrations of leptin potentiated depolarization-induced contraction of coronary arteries. Inhibition of Rho kinase significantly reduced leptin-mediated increases in coronary artery contractions. The effects of leptin on the functional expression of Rho kinase were time-dependent, as acute treatment increased Rho kinase activity while chronic (3 day) exposure was associated with increases in Rho kinase protein abundance. Proliferation assays following chronic leptin administration (8 day, serum-containing media) demonstrated that leptin augmented coronary vascular smooth muscle proliferation and increased Rho kinase activity. Inhibition of Rho kinase significantly reduced these effects of leptin. Taken together, these findings demonstrate that leptin promotes increases in coronary vasoconstriction and smooth muscle proliferation and indicate that these phenotypic effects are associated with alterations in the coronary artery proteome and dynamic effects on the Rho kinase pathway. PMID:26975316

  11. A MAP kinase dependent feedback mechanism controls Rho1 GTPase and actin distribution in yeast.

    Directory of Open Access Journals (Sweden)

    Shuguang Guo

    Full Text Available In the yeast Saccharomyces cerevisiae the guanosine triphosphatase (GTPase Rho1 controls actin polarization and cell wall expansion. When cells are exposed to various environmental stresses that perturb the cell wall, Rho1 activates Pkc1, a mammalian Protein Kinase C homologue, and Mpk1, a mitogen activated protein kinase (MAPK, resulting in actin depolarization and cell wall remodeling. In this study, we demonstrate a novel feedback loop in this Rho1-mediated Pkc1-MAPK pathway that involves regulation of Rom2, the guanine nucleotide exchange factor of Rho1, by Mpk1, the end kinase of the pathway. This previously unrecognized Mpk1-dependent feedback is a critical step in regulating Rho1 function. Activation of this feedback mechanism is responsible for redistribution of Rom2 and cell wall synthesis activity from the bud to cell periphery under stress conditions. It is also required for terminating Rho1 activity toward the Pkc1-MAPK pathway and for repolarizing actin cytoskeleton and restoring growth after the stressed cells become adapted.

  12. Metformin is an AMP kinase-dependent growth inhibitor for breast cancer cells.

    Science.gov (United States)

    Zakikhani, Mahvash; Dowling, Ryan; Fantus, I George; Sonenberg, Nahum; Pollak, Michael

    2006-11-01

    Recent population studies provide clues that the use of metformin may be associated with reduced incidence and improved prognosis of certain cancers. This drug is widely used in the treatment of type 2 diabetes, where it is often referred to as an "insulin sensitizer" because it not only lowers blood glucose but also reduces the hyperinsulinemia associated with insulin resistance. As insulin and insulin-like growth factors stimulate proliferation of many normal and transformed cell types, agents that facilitate signaling through these receptors would be expected to enhance proliferation. We show here that metformin acts as a growth inhibitor rather than an insulin sensitizer for epithelial cells. Breast cancer cells can be protected against metformin-induced growth inhibition by small interfering RNA against AMP kinase. This shows that AMP kinase pathway activation by metformin, recently shown to be necessary for metformin inhibition of gluconeogenesis in hepatocytes, is also involved in metformin-induced growth inhibition of epithelial cells. The growth inhibition was associated with decreased mammalian target of rapamycin and S6 kinase activation and a general decrease in mRNA translation. These results provide evidence for a mechanism that may contribute to the antineoplastic effects of metformin suggested by recent population studies and justify further work to explore potential roles for activators of AMP kinase in cancer prevention and treatment. PMID:17062558

  13. Discovery of checkpoint kinase inhibitor (S)-5-(3-fluorophenyl)-N-(piperidin-3-yl)-3-ureidothiophene-2-carboxamide (AZD7762) by structure-based design and optimization of thiophenecarboxamide ureas.

    Science.gov (United States)

    Oza, Vibha; Ashwell, Susan; Almeida, Lynsie; Brassil, Patrick; Breed, Jason; Deng, Chun; Gero, Thomas; Grondine, Michael; Horn, Candice; Ioannidis, Stephanos; Liu, Dongfang; Lyne, Paul; Newcombe, Nicholas; Pass, Martin; Read, Jon; Ready, Shannon; Rowsell, Siân; Su, Mei; Toader, Dorin; Vasbinder, Melissa; Yu, Dingwei; Yu, Yan; Xue, Yafeng; Zabludoff, Sonya; Janetka, James

    2012-06-14

    Checkpoint kinases CHK1 and CHK2 are activated in response to DNA damage that results in cell cycle arrest, allowing sufficient time for DNA repair. Agents that lead to abrogation of such checkpoints have potential to increase the efficacy of such compounds as chemo- and radiotherapies. Thiophenecarboxamide ureas (TCUs) were identified as inhibitors of CHK1 by high throughput screening. A structure-based approach is described using crystal structures of JNK1 and CHK1 in complex with 1 and 2 and of the CHK1-3b complex. The ribose binding pocket of CHK1 was targeted to generate inhibitors with excellent cellular potency and selectivity over CDK1and IKKβ, key features lacking from the initial compounds. Optimization of 3b resulted in the identification of a regioisomeric 3-TCU lead 12a. Optimization of 12a led to the discovery of the clinical candidate 4 (AZD7762), which strongly potentiates the efficacy of a variety of DNA-damaging agents in preclinical models. PMID:22551018

  14. Role of the human papillomavirus E2 protein at cell cycle checkpoints%人乳头瘤病毒E2蛋白在细胞周期检测点中的作用研究进展

    Institute of Scientific and Technical Information of China (English)

    李耀林; 唐双阳; 万艳平

    2012-01-01

    The human papillomavirus (HPV) E2 protein is a transcription-inhibiting factor and tumor suppressor of E6 or E7. Recent studies found that E2 protein interacts with the mitotic checkpoint during HPV-induced cell transformation. The protein affects the activity of Cdc20, Skp2, and APC/C, which are involved in the spindle assembly checkpoint, and is related to a cell's genetic stability. Due to the role the E2 protein plays in encouraging cancer to develop, it may become a new target for the prevention and treatment of cancer caused by high-risk HPV.%人乳头瘤病毒(HPV) E2蛋白一直被认为是E6/E7的转录抑制因子与肿瘤抑制因子.近年研究发现,在HPV所致细胞转化过程中,E2蛋白与细胞有丝分裂检测点相互作用,影响Cdc20、Skp2和APC/C等活性,涉及纺锤体组装检测点,关系到细胞基因的稳定性.由于E2蛋白可能在HPV致癌中具有推动作用,因而有望成为防治高危型HPVs所致肿瘤的一个新靶点.

  15. GRID COMPUTING AND CHECKPOINT APPROACH

    OpenAIRE

    Pankaj gupta

    2011-01-01

    Grid computing is a means of allocating the computational power of alarge number of computers to complex difficult computation or problem. Grid computing is a distributed computing paradigm thatdiffers from traditional distributed computing in that it is aimed toward large scale systems that even span organizational boundaries. In this paper we investigate the different techniques of fault tolerance which are used in many real time distributed systems. The main focus is on types of fault occu...

  16. Biological and molecular mechanisms of sulfur mustard analogue-induced toxicity in JB6 and HaCaT cells: possible role of ataxia telangiectasia-mutated/ataxia telangiectasia-Rad3-related cell cycle checkpoint pathway.

    Science.gov (United States)

    Tewari-Singh, Neera; Gu, Mallikarjuna; Agarwal, Chapla; White, Carl W; Agarwal, Rajesh

    2010-06-21

    Effective medical treatment and preventive measures for chemical warfare agent sulfur mustard (HD)-caused incapacitating skin toxicity are lacking, because of limited knowledge of its mechanism of action. The proliferating basal epidermal cells are primary major sites of attack during HD-caused skin injury. Therefore, employing mouse JB6 and human HaCaT epidermal cells, here, we investigated the molecular mechanism of HD analogue 2-chloroethyl ethyl sulfide (CEES)-induced skin cytotoxicity. As compared to the control, up to 1 mM CEES treatment of these cells for 2, 4, and 24 h caused dose-dependent decreases in cell viability and proliferation as measured by DNA synthesis, together with S and G2-M phase arrest in cell cycle progression. Mechanistic studies showed phosphorylation of DNA damage sensors and checkpoint kinases, ataxia telangiectasia-mutated (ATM) at ser1981 and ataxia telangiectasia-Rad3-related (ATR) at ser428 within 30 min of CEES exposure, and modulation of S and G2-M phase-associated cell cycle regulatory proteins, which are downstream targets of ATM and ATR kinases. Hoechst-propidium iodide staining demonstrated that CEES-induced cell death was both necrotic and apoptotic in nature, and the latter was induced at 4 and 24 h of CEES treatment in HaCaT and JB6 cells, respectively. An increase in caspase-3 activity and both caspase-3 and poly(ADP-ribose)polymerase (PARP) cleavage coinciding with CEES-caused apoptosis in both cell lines suggested the involvement of the caspase pathway. Together, our findings suggest a DNA-damaging effect of CEES that activates ATM/ATR cell cycle checkpoint signaling as well as caspase-PARP pathways, leading to cell cycle arrest and apoptosis/necrosis in both JB6 and HaCaT cells. The identified molecular targets, quantitative biomarkers, and epidermal cell models in this study have the potential and usefulness in rapid development of effective prophylactic and therapeutic interventions against HD-induced skin toxicity

  17. Cellular Inhibition of Checkpoint Kinase 2 (Chk2) and Potentiation of Camptothecins and Radiation by the Novel Chk2 Inhibitor PV1019 [7-Nitro-1H-indole-2-carboxylic acid {4-[1-(guanidinohydrazone)-ethyl]-phenyl}-amide

    Energy Technology Data Exchange (ETDEWEB)

    Jobson, Andrew G.; Lountos, George T.; Lorenzi, Philip L.; Llamas, Jenny; Connelly, John; Cerna, David; Tropea, Joseph E.; Onda, Akikazu; Zoppoli, Gabriele; Kondapaka, Sudhir; Zhang, Guangtao; Caplen, Natasha J.; Cardellina, II, John H.; Yoo, Stephen S.; Monks, Anne; Self, Christopher; Waugh, David S.; Shoemaker, Robert H.; Pommier, Yves; (NIH)

    2010-04-05

    Chk2 is a checkpoint kinase involved in the ataxia telangiectasia mutated pathway, which is activated by genomic instability and DNA damage, leading to either cell death (apoptosis) or cell cycle arrest. Chk2 provides an unexplored therapeutic target against cancer cells. We recently reported 4,4'-diacetyldiphenylurea-bis(guanylhydrazone) (NSC 109555) as a novel chemotype Chk2 inhibitor. We have now synthesized a derivative of NSC 109555, PV1019 (NSC 744039) [7-nitro-1H-indole-2-carboxylic acid {l_brace}4-[1-(guanidinohydrazone)-ethyl]-phenyl{r_brace}-amide], which is a selective submicromolar inhibitor of Chk2 in vitro. The cocrystal structure of PV1019 bound in the ATP binding pocket of Chk2 confirmed enzymatic/biochemical observations that PV1019 acts as a competitive inhibitor of Chk2 with respect to ATP. PV1019 was found to inhibit Chk2 in cells. It inhibits Chk2 autophosphorylation (which represents the cellular kinase activation of Chk2), Cdc25C phosphorylation, and HDMX degradation in response to DNA damage. PV1019 also protects normal mouse thymocytes against ionizing radiation-induced apoptosis, and it shows synergistic antiproliferative activity with topotecan, camptothecin, and radiation in human tumor cell lines. We also show that PV1019 and Chk2 small interfering RNAs can exert antiproliferative activity themselves in the cancer cells with high Chk2 expression in the NCI-60 screen. These data indicate that PV1019 is a potent and selective inhibitor of Chk2 with chemotherapeutic and radiosensitization potential.

  18. Phosphatidylinositol-3-kinase-dependent phosphorylation of SLP-76 by the lymphoma-associated ITK-SYK fusion-protein

    International Nuclear Information System (INIS)

    Recurrent chromosomal translocations have long been implicated in various types of lymphomas and other malignancies. Novel recurrent t(5;9)(q33;q22) has been recently discovered in un-specified peripheral T-cell lymphoma. To elucidate the role of this translocation, the corresponding fusion construct encoding the N-terminal portion of the ITK kinase and the C-terminal catalytic region of the SYK kinase was generated. We herein show that the ITK-SYK fusion-protein is constitutively active. Moreover, we demonstrate that ITK-SYK is phosphorylated on key tyrosine residues and is capable of potently phosphorylating the related adapter proteins BLNK and SLP-76. In transiently transfected cells, SYK was phosphorylated at Y352 but not detectably at the activation-loop tyrosines Y525/Y526. In contrast, ITK-SYK was phosphorylated both at Y212 and the activation-loop tyrosines Y385/Y386, corresponding to Y352 and Y525/Y526 in SYK, respectively. In resting primary lymphocytes, ITK-SYK predominantly localizes to the cell surface. In addition, we demonstrate that following stimulation, the ITK-SYK fusion-protein in cell lines translocates to the cell membrane and, moreover, that this phenomenon as well as SLP-76 phosphorylation are blocked upon phosphatidylinositol-3-kinase (PI3-kinase) inhibition.

  19. Spindle-F Is the Central Mediator of Ik2 Kinase-Dependent Dendrite Pruning in Drosophila Sensory Neurons.

    Science.gov (United States)

    Lin, Tzu; Pan, Po-Yuan; Lai, Yu-Ting; Chiang, Kai-Wen; Hsieh, Hsin-Lun; Wu, Yi-Ping; Ke, Jian-Ming; Lee, Myong-Chol; Liao, Shih-Sian; Shih, Hsueh-Tzu; Tang, Chiou-Yang; Yang, Shi-Bing; Cheng, Hsu-Chen; Wu, June-Tai; Jan, Yuh-Nung; Lee, Hsiu-Hsiang

    2015-11-01

    During development, certain Drosophila sensory neurons undergo dendrite pruning that selectively eliminates their dendrites but leaves the axons intact. How these neurons regulate pruning activity in the dendrites remains unknown. Here, we identify a coiled-coil protein Spindle-F (Spn-F) that is required for dendrite pruning in Drosophila sensory neurons. Spn-F acts downstream of IKK-related kinase Ik2 in the same pathway for dendrite pruning. Spn-F exhibits a punctate pattern in larval neurons, whereas these Spn-F puncta become redistributed in pupal neurons, a step that is essential for dendrite pruning. The redistribution of Spn-F from puncta in pupal neurons requires the phosphorylation of Spn-F by Ik2 kinase to decrease Spn-F self-association, and depends on the function of microtubule motor dynein complex. Spn-F is a key component to link Ik2 kinase to dynein motor complex, and the formation of Ik2/Spn-F/dynein complex is critical for Spn-F redistribution and for dendrite pruning. Our findings reveal a novel regulatory mechanism for dendrite pruning achieved by temporal activation of Ik2 kinase and dynein-mediated redistribution of Ik2/Spn-F complex in neurons. PMID:26540204

  20. Differential Rho-kinase dependency of full and partial muscarinic receptor agonists in airway smooth muscle contraction

    NARCIS (Netherlands)

    Schaafsma, D; Boterman, M; de Jong, AM; Hovens, Iris; Penninks, JM; Nelemans, SA; Meurs, H; Zaagsma, J

    2006-01-01

    1 In airway smooth muscle (ASM), full and partial muscarinic receptor agonists have been described to have large differences in their ability to induce signal transduction, including Ca2+-mobilization. Despite these differences, partial agonists are capable of inducing a submaximal to maximal ASM co

  1. Stanniocalcin-1 inhibits renal ischemia/reperfusion injury via an AMP-activated protein kinase-dependent pathway

    Science.gov (United States)

    AKI is associated with increased morbidity, mortality, and cost of care, and therapeutic options remain limited. Reactive oxygen species are critical for the genesis of ischemic AKI. Stanniocalcin-1 (STC1) suppresses superoxide generation through induction of uncoupling proteins (UCPs), and transgen...

  2. TNF causes changes in glomerular endothelial permeability and morphology through a Rho and myosin light chain kinase-dependent mechanism.

    Science.gov (United States)

    Xu, Chang; Wu, Xiaoyan; Hack, Bradley K; Bao, Lihua; Cunningham, Patrick N

    2015-12-01

    A key function of the endothelium is to serve as a regulated barrier between tissue compartments. We have previously shown that tumor necrosis factor (TNF) plays a crucial role in lipopolysaccharide (LPS)-induced acute kidney injury, in part by causing injury to the renal endothelium through its receptor TNFR1. Here, we report that TNF increased permeability to albumin in primary culture mouse renal endothelial cells, as well as human glomerular endothelial cells. This process occurred in association with changes in the actin cytoskeleton and was associated with gaps between previously confluent cells in culture and decreases in the tight junction protein occludin. This process was dependent on myosin light chain activation, as seen by its prevention with Rho-associated kinase and myosin light chain kinase (MLCK) inhibitors. Surprisingly, permeability was not blocked by inhibition of apoptosis with caspase inhibitors. Additionally, we found that the renal glycocalyx, which plays an important role in barrier function, was also degraded by TNF in a Rho and MLCK dependent fashion. TNF treatment caused a decrease in the size of endothelial fenestrae, dependent on Rho and MLCK, although the relevance of this to changes in permeability is uncertain. In summary, TNF-induced barrier dysfunction in renal endothelial cells is crucially dependent upon the Rho/MLCK signaling pathway. PMID:26634902

  3. Salinomycin Activates AMP-Activated Protein Kinase-Dependent Autophagy in Cultured Osteoblastoma Cells: A Negative Regulator against Cell Apoptosis

    OpenAIRE

    Zhu, Lun-qing; Zhen, Yun-Fang; Zhang, Ya; Guo, Zhi-Xiong; Dai, Jin; Wang, Xiao-Dong

    2013-01-01

    Background The malignant osteoblastoma has poor prognosis, thus the search for novel and more efficient chemo-agents against this disease is urgent. Salinomycin induces broad anti-cancer effects both in vivo and in vitro, however, its role in osteoblastoma is still not clear. Key Findings Salinomycin induced both apoptosis and autophagy in cultured U2OS and MG-63 osteoblastoma cells. Inhibition of autophagy by 3-methyladenine (3-MA), or by RNA interference (RNAi) of light chain 3B (LC3B), enh...

  4. PI3-kinase-dependent activation of apoptotic machinery oc-curs on commitment of epidermal keratinocytes to terminal differentiation

    Institute of Scientific and Technical Information of China (English)

    Sam M Janes; Tyler A Ofstad; Douglas H Campbell; Ayad Eddaoudi; Gary Warnes; Derek Davies; Fiona M Watt

    2009-01-01

    We have investigated the earliest events in commitment of human epidermal keratinocytes to terminal differen-tiation. Phosphorylated Akt and caspase activation were detected in cells exiting the basal layer of the epidermis. Activation of Akt by retroviral transduction of primary cultures of human keratinocytes resulted in an increase in abortive clones founded by transit amplifying cells, while inhibition of the upstream kinase, Pl3-kinase, inhibited suspension-induced terminal differentiation. Caspase inhibition also blocked differentiation, the primary mediator being caspase 8. Caspase activation was initiated by 2 h in suspension, preceding the onset of expression of the termi-nal differentiation marker involucrin by several hours. Incubation of suspended cells with fibronectin or inhibition of PI3-kinase prevented caspase induction. At 2 h in suspension, keratinocytes that had become committed to terminal differentiation had increased side scatter, were 7-aminoactinomycin D (7-AAD) positive and annexin V negative; they exhibited loss of mitochondrial membrane potential and increased cardiolipin oxidation, but with no increase in reac-tive oxygen species. These properties indicate that the onset of terminal differentiation, while regulated by Pl3-kinase and caspases, is not a classical apoptotic process.

  5. Phosphoinositide 3-kinase dependent inhibition as a broad basis for opponent coding in Mammalian olfactory receptor neurons.

    Directory of Open Access Journals (Sweden)

    Kirill Ukhanov

    Full Text Available Phosphoinositide 3-kinase (PI3K signaling has been implicated in mediating inhibitory odorant input to mammalian olfactory receptor neurons (ORNs. To better understand the breadth of such inhibition in odor coding, we screened a panel of odorants representing different chemical classes, as well as odorants known to occur in a natural odor object (tomato, for their ability to rapidly activate PI3K-dependent inhibitory signaling. Odorants were screened on dissociated native rat ORNs before and after pre-incubation with the PI3K-isoform specific blockers AS252424 and TGX221. Many different odorants increased their excitatory strength for particular ORNs following PI3K blockade in a manner consistent with activating PI3K-dependent inhibitory signaling in those cells. The PI3K-dependent inhibitory odorants overlapped with conventional excitatory odorants, but did not share the same bias, indicating partial partitioning of the odor space. Finding that PI3K-dependent inhibition can be activated by a wide range of otherwise conventional excitatory odorants strongly implies PI3K-dependent inhibition provides a broad basis for opponent coding in mammalian ORNs.

  6. LIGHT May Improve Immune Checkpoint Blockade Response.

    Science.gov (United States)

    2016-06-01

    A new study suggests that insufficient T-cell infiltration may explain why a majority of patients do not respond to immunotherapy. Combining PD-L1 inhibitors with antibody-guided LIGHT, a protein that recruits tumor-infiltrating lymphocytes, increased antitumor response in mice, and may have the potential to improve patient response rates to immunotherapy. PMID:27080334

  7. G2 Checkpoint Responses in Arabidopsis

    Energy Technology Data Exchange (ETDEWEB)

    Britt, Anne

    2013-03-18

    This project focused on the mechanism and biological significance of the G2 arrest response to replication stress in plants. We employed both forward and reverse genetic approaches to identify genes required for this response. A total of 3 different postdocs, 5 undergraduates, and 2 graduate students participated in the project. We identified several genes required for damage response in plants, including homologs of genes previously identified in animals (ATM and ATR), novel, a plant-specific genes (SOG1) and a gene known in animals but previously thought to be missing from the Arabidopsis genome (ATRIP). We characterized the transcriptome of gamma-irradiated plants, and found that plants, unlike animals, express a robust transcriptional response to damage, involving genes that regulate the cell cycle and DNA metabolism. This response requires both ATM and the transcription factor SOG1. We found that both ATM and ATR play a role in meiosis in plants. We also found that plants have a cell-type-specific programmed cell death response to ionizing radiation and UV light, and that this response requires ATR, ATM, and SOG1. These results were published in a series of 5 papers.

  8. Delta-like 1/fetal antigen-1 (Dlk1/FA1) is a novel regulator of chondrogenic cell differentiation via inhibition of the Akt kinase-dependent pathway

    DEFF Research Database (Denmark)

    Chen, Li; Qanie, Diyako; Jafari, Abbas; Taipaleenmaki, Hanna; Jensen, Charlotte H; Säämänen, Anna-Marja; Sanz, Maria Luisa Nueda; Laborda, Jorge; Abdallah, Basem M; Kassem, Moustapha

    2011-01-01

    Delta-like 1 (Dlk1, also known as fetal antigen-1, FA1) is a member of Notch/Delta family that inhibits adipocyte and osteoblast differentiation; however, its role in chondrogenesis is still not clear. Thus, we overexpressed Dlk1/FA1 in mouse embryonic ATDC5 cells and tested its effects on...

  9. Mitogen-activated protein kinase-dependent apoptosis in norcan-tharidin-treated A375-S2 cells is proceeded by the activation of protein kinase C

    Institute of Scientific and Technical Information of China (English)

    AN Wei-wei; WANG Min-wei; Tashiro Shin-ichi; Onodera Satoshi; Ikejima Takashi

    2005-01-01

    Background We have reported that norcantharidin (NCTD) induces human melanoma A375-S2 cell apoptosis and that the activation of caspase and the mitochondrial pathway are involved in the apoptotic process. This study aimed at investigating the roles of mitogen-activated protein kinase (MAPK) and protein kinase C (PKC) in A375-S2 cell apoptosis induced by NCTD. Methods We assessed the effects of NCTD on cell growth inhibition using the 3-(4,5-dimethylthiazol-2-yl)-2,5-dipheyltetrazolium bromide (MTT) assay, DNA fragmentation (DNA agarose gel electrophoresis), and MAPK protein levels (Western blot analysis) in A375-S2 cells. Photomicroscopic data were also collected.Results The NCTD inhibitory effect on A375-S2 cells was partially reversed by MAPK and PKC inhibitors. The expression of phosphorylated JNK and p38 also increased after the treatment with NCTD, and inhibitors of c-Jun NH2-terminal kinase (JNK) and p38 (SP600125 and SB203580, respectively) had significant inhibitory effects on the upregulation of phosphorylated JNK and p38 expression. Simultaneously, the PKC inhibitor staurosporine blocked the upregulation of phosphorylated JNK and phosphorylated p38, but had little effect on extracellular signal-regulated kinase (ERK) expression. Conclusion These results suggest that the activation of JNK and p38 MAPK promotes the process of NCTD-induced A375-S2 cell apoptosis and that PKC plays an important regulation role in the activation of MAPKs.

  10. Sip4, a Snf1 kinase-dependent transcriptional activator, binds to the carbon source-responsive element of gluconeogenic genes.

    OpenAIRE

    Vincent, O.; Carlson, M

    1998-01-01

    The carbon source-responsive element (CSRE) mediates transcriptional activation of the gluconeogenic genes during growth of the yeast Saccharomyces cerevisiae on non-fermentable carbon sources. Previous studies have suggested that the Cat8 protein activates the expression of CSRE-binding factors. We show here that one of these factors is Sip4, a glucose-regulated C6 zinc cluster activator which was identified by its interaction with the Snf1 protein kinase. We present genetic evidence that Si...

  11. P-Selectin Cross-Links PSGL-1 and Enhances Neutrophil Adhesion to Fibrinogen and ICAM-1 in a Src Kinase-Dependent, but GPCR-Independent Mechanism

    OpenAIRE

    Xu, Tao; Zhang, Lei; Geng, Zhen H; Wang, Hai-Bo; Wang, Jin-Tao; Chen, Ming; Geng, Jian-Guo

    2007-01-01

    Endothelial and platelet P-selectin (CD62P) and leukocyte integrin αMβ2 (CD11bCD18, Mac-1) are cell adhesion molecules essential for host defense and innate immunity. Upon inflammatory challenges, P-selectin binds to PSGL-1 (P-selectin glycoprotein ligand-1, CD162) to mediate neutrophil rolling, during which integrins become activated by extracellular stimuli for their firm adhesion in a G-protein coupled receptor (GPCR)-dependent mechanism. Here we show that cross-linking of PSGL-1 by dimeri...

  12. C-peptide increases Na,K-ATPase expression via PKC- and MAP kinase-dependent activation of transcription factor ZEB in human renal tubular cells

    DEFF Research Database (Denmark)

    Galuska, Dana; Pirkmajer, Sergej; Barres, Romain;

    2011-01-01

    Replacement of proinsulin C-peptide in type 1 diabetes ameliorates nerve and kidney dysfunction, conditions which are associated with a decrease in Na,K-ATPase activity. We determined the molecular mechanism by which long term exposure to C-peptide stimulates Na,K-ATPase expression and activity in...

  13. C-peptide increases Na,K-ATPase expression via PKC- and MAP kinase-dependent activation of transcription factor ZEB in human renal tubular cells.

    Directory of Open Access Journals (Sweden)

    Dana Galuska

    Full Text Available BACKGROUND: Replacement of proinsulin C-peptide in type 1 diabetes ameliorates nerve and kidney dysfunction, conditions which are associated with a decrease in Na,K-ATPase activity. We determined the molecular mechanism by which long term exposure to C-peptide stimulates Na,K-ATPase expression and activity in primary human renal tubular cells (HRTC in control and hyperglycemic conditions. METHODOLOGY/PRINCIPAL FINDINGS: HRTC were cultured from the outer cortex obtained from patients undergoing elective nephrectomy. Ouabain-sensitive rubidium ((86Rb(+ uptake and Na,K-ATPase activity were determined. Abundance of Na,K-ATPase was determined by Western blotting in intact cells or isolated basolateral membranes (BLM. DNA binding activity was determined by electrical mobility shift assay (EMSA. Culturing of HRTCs for 5 days with 1 nM, but not 10 nM of human C-peptide leads to increase in Na,K-ATPase α(1-subunit protein expression, accompanied with increase in (86Rb(+ uptake, both in normal- and hyperglycemic conditions. Na,K-ATPase α(1-subunit expression and Na,K-ATPase activity were reduced in BLM isolated from cells cultured in presence of high glucose. Exposure to1 nM, but not 10 nM of C-peptide increased PKCε phosphorylation as well as phosphorylation and abundance of nuclear ERK1/2 regardless of glucose concentration. Exposure to 1 nM of C-peptide increased DNA binding activity of transcription factor ZEB (AREB6, concomitant with Na,K-ATPase α(1-subunit mRNA expression. Effects of 1 nM C-peptide on Na,K-ATPase α(1-subunit expression and/or ZEB DNA binding activity in HRTC were abolished by incubation with PKC or MEK1/2 inhibitors and ZEB siRNA silencing. CONCLUSIONS/SIGNIFICANCE: Despite activation of ERK1/2 and PKC by hyperglycemia, a distinct pool of PKCs and ERK1/2 is involved in regulation of Na,K-ATPase expression and activity by C-peptide. Most likely C-peptide stimulates sodium pump expression via activation of ZEB, a transcription factor that has not been previously implicated in C-peptide-mediated signaling. Importantly, only physiological concentrations of C-peptide elicit this effect.

  14. Constitutive excluslion of Csk from Hck-positive membrane microdomains permits Src kinase-dependent proliferation of Theileria-transformed B lymphocytes

    Czech Academy of Sciences Publication Activity Database

    Baumgartner, M.; Angelisová, Pavla; Setterblad, N.; Mooney, N.; Werling, D.; Hořejší, Václav; Langsley, G.

    2003-01-01

    Roč. 101, č. 5 (2003), s. 1874-1881. ISSN 0006-4971 R&D Projects: GA MŠk LN00A026 Institutional research plan: CEZ:AV0Z5052915 Keywords : Theileriosis * transformation * Csk Subject RIV: EC - Immunology Impact factor: 10.120, year: 2003

  15. Interleukin 17A promotes pneumococcal clearance by recruiting neutrophils and inducing apoptosis through a p38 mitogen-activated protein kinase-dependent mechanism in acute otitis media.

    Science.gov (United States)

    Wang, Wei; Zhou, Aie; Zhang, Xuemei; Xiang, Yun; Huang, Yifei; Wang, Lei; Zhang, Shuai; Liu, Yusi; Yin, Yibing; He, Yujuan

    2014-06-01

    Streptococcus pneumoniae is a Gram-positive and human-restricted pathogen colonizing the nasopharynx with an absence of clinical symptoms as well as a major pathogen causing otitis media (OM), one of the most common childhood infections. Upon bacterial infection, neutrophils are rapidly activated and recruited to the infected site, acting as the frontline defender against emerging microbial pathogens via different ways. Evidence shows that interleukin 17A (IL-17A), a neutrophil-inducing factor, plays important roles in the immune responses in several diseases. However, its function in response to S. pneumoniae OM remains unclear. In this study, the function of IL-17A in response to S. pneumoniae OM was examined using an in vivo model. We developed a model of acute OM (AOM) in C57BL/6 mice and found that neutrophils were the dominant immune cells that infiltrated to the middle ear cavity (MEC) and contributed to bacterial clearance. Using IL-17A knockout (KO) mice, we found that IL-17A boosted neutrophil recruitment to the MEC and afterwards induced apoptosis, which was identified to be conducive to bacterial clearance. In addition, our observation suggested that the p38 mitogen-activated protein kinase (MAPK) signaling pathway was involved in the recruitment and apoptosis of neutrophils mediated by IL-17A. These data support the conclusion that IL-17A contributes to the host immune response against S. pneumoniae by promoting neutrophil recruitment and apoptosis through the p38 MAPK signaling pathway. PMID:24664502

  16. STI571 reduces TRAIL-induced apoptosis in colon cancer cells: c-Abl activation by the death receptor leads to stress kinase-dependent cell death

    Directory of Open Access Journals (Sweden)

    Huang Duen-Yi

    2012-03-01

    Full Text Available Abstract Background In an effort to achieve better cancer therapies, we elucidated the combination cancer therapy of STI571 (an inhibitor of Bcr-Abl and clinically used for chronic myelogenous leukemia and TNF-related apoptosis-inducing ligand (TRAIL, a developing antitumor agent in leukemia, colon, and prostate cancer cells. Methods Colon cancer (HCT116, SW480, prostate cancer (PC3, LNCaP and leukemia (K562 cells were treated with STI571 and TRAIL. Cell viability was determined by MTT assay and sub-G1 appearance. Protein expression and kinase phosphorylation were determined by Western blotting. c-Abl and p73 activities were inhibited by target-specific small interfering (siRNA. In vitro kinase assay of c-Abl was conducted using CRK as a substrate. Results We found that STI571 exerts opposite effects on the antitumor activity of TRAIL. It enhanced cytotoxicity in TRAIL-treated K562 leukemia cells and reduced TRAIL-induced apoptosis in HCT116 and SW480 colon cancer cells, while having no effect on PC3 and LNCaP cells. In colon and prostate cancer cells, TRAIL caused c-Abl cleavage to the active form via a caspase pathway. Interestingly, JNK and p38 MAPK inhibitors effectively blocked TRAIL-induced toxicity in the colon, but not in prostate cancer cells. Next, we found that STI571 could attenuate TRAIL-induced c-Abl, JNK and p38 activation in HCT116 cells. In addition, siRNA targeting knockdown of c-Abl and p73 also reduced TRAIL-induced cytotoxicity, rendering HCT116 cells less responsive to stress kinase activation, and masking the cytoprotective effect of STI571. Conclusions All together we demonstrate a novel mediator role of p73 in activating the stress kinases p38 and JNK in the classical apoptotic pathway of TRAIL. TRAIL via caspase-dependent action can sequentially activate c-Abl, p73, and stress kinases, which contribute to apoptosis in colon cancer cells. Through the inhibition of c-Abl-mediated apoptotic p73 signaling, STI571 reduces the antitumor activity of TRAIL in colon cancer cells. Our results raise additional concerns when developing combination cancer therapy with TRAIL and STI571 in the future.

  17. Insulin promotes Rip11 accumulation at the plasma membrane by inhibiting a dynamin- and PI3-kinase-dependent, but Akt-independent, internalisation event

    OpenAIRE

    Boal, Frédéric; Hodgson, Lorna R.; Reed, Sam E; Yarwood, Sophie E; Just, Victoria J; Stephens, David J; McCaffrey, Mary W.; Tavaré, Jeremy M.

    2016-01-01

    Rip11 is a Rab11 effector protein that has been shown to be important in controlling the trafficking of several intracellular cargoes, including the fatty acid transporter FAT/CD36, V-ATPase and the glucose transporter GLUT4. We have previously demonstrated that Rip11 translocates to the plasma membrane in response to insulin and here we examine the basis of this regulated phenomenon in more detail. We show that Rip11 rapidly recycles between the cell interior and surface, and that the abilit...

  18. Insulin promotes Rip11 accumulation at the plasma membrane by inhibiting a dynamin- and PI3-kinase-dependent, but Akt-independent, internalisation event.

    Science.gov (United States)

    Boal, Frédéric; Hodgson, Lorna R; Reed, Sam E; Yarwood, Sophie E; Just, Victoria J; Stephens, David J; McCaffrey, Mary W; Tavaré, Jeremy M

    2016-01-01

    Rip11 is a Rab11 effector protein that has been shown to be important in controlling the trafficking of several intracellular cargoes, including the fatty acid transporter FAT/CD36, V-ATPase and the glucose transporter GLUT4. We have previously demonstrated that Rip11 translocates to the plasma membrane in response to insulin and here we examine the basis of this regulated phenomenon in more detail. We show that Rip11 rapidly recycles between the cell interior and surface, and that the ability of insulin to increase the appearance of Rip11 at the cell surface involves an inhibition of Rip11 internalisation from the plasma membrane. By contrast the hormone has no effect on the rate of Rip11 translocation towards the plasma membrane. The ability of insulin to inhibit Rip11 internalisation requires dynamin and class I PI3-kinases, but is independent of the activation of the protein kinase Akt; characteristics which are very similar to the mechanism by which insulin inhibits GLUT4 endocytosis. PMID:26515129

  19. Substance P induces rapid and transient membrane blebbing in U373MG cells in a p21-activated kinase-dependent manner.

    Directory of Open Access Journals (Sweden)

    John Meshki

    Full Text Available U373MG astrocytoma cells endogenously express the full-length neurokinin 1 receptor (NK1R. Substance P (SP, the natural ligand for NK1R, triggers rapid and transient membrane blebbing and we report that these morphological changes have different dynamics and intracellular signaling as compared to the changes that we have previously described in HEK293-NK1R cells. In both cell lines, the SP-induced morphological changes are Gq-independent, and they require the Rho, Rho-associated coiled-coil kinase (ROCK signaling pathway. Using confocal microscopy we have demonstrated that tubulin is phosphorylated subsequent to cell stimulation with SP and that tubulin accumulates inside the blebs. Colchicine, a tubulin polymerization inhibitor, blocked SP-induced blebbing in U373MG but not in HEK293-NK1R cells. Although p21-activated kinase (PAK is expressed in both cell lines, SP induced rapid phosphorylation of PAK in U373MG, but failed to phosphorylate PAK in HEK293-NK1R cells. The cell-permeable Rho inhibitor C3 transferase inhibited SP-induced PAK phosphorylation, but the ROCK inhibitor Y27632 had no effect on PAK phosphorylation, suggesting that Rho activates PAK in a ROCK-independent manner. Our study demonstrates that SP triggers rapid changes in cell morphology mediated by distinct intracellular signaling mechanisms in U373MG versus HEK293-NK1R cells.

  20. Cyclic AMP (cAMP)-mediated stimulation of adipocyte differentiation requires the synergistic action of Epac- and cAMP-dependent protein kinase-dependent processes

    DEFF Research Database (Denmark)

    Petersen, Rasmus Koefoed; Madsen, Lise; Pedersen, Lone Møller;

    2008-01-01

    Cyclic AMP (cAMP)-dependent processes are pivotal during the early stages of adipocyte differentiation. We show that exchange protein directly activated by cAMP (Epac), which functions as a guanine nucleotide exchange factor for the Ras-like GTPases Rap1 and Rap2, was required for c...

  1. High glucose enhances transient receptor potential channel canonical type 6-dependent calcium influx in human platelets via phosphatidylinositol 3-kinase-dependent pathway

    DEFF Research Database (Denmark)

    Liu, Daoyan; Maier, Alexandra; Scholze, Alexandra; Rauch, Ursula; Boltzen, Ulrike; Zhao, Zhigang; Zhu, Zhiming; Tepel, Martin

    2008-01-01

    Transient receptor potential canonical type 6 (TRPC6) channels mediating 1-oleoyl-2-acetyl-sn-glycerol (OAG)-induced calcium entry have been identified on human platelets. In the present study we tested the hypothesis that hyperglycemia increases the expression of TRPC6 channels....

  2. Multi-walled carbon nanotubes induce COX-2 and iNOS expression via MAP Kinase-dependent and -independent mechanisms in mouse RAW264.7 macrophages

    Directory of Open Access Journals (Sweden)

    Lee Jong

    2012-05-01

    Full Text Available Abstract Background Carbon nanotubes (CNTs are engineered graphene cylinders with numerous applications in engineering, electronics and medicine. However, CNTs cause inflammation and fibrosis in the rodent lung, suggesting a potential human health risk. We hypothesized that multi-walled CNTs (MWCNTs induce two key inflammatory enzymes in macrophages, cyclooxygenase-2 (COX-2 and inducible nitric oxide synthase (iNOS, through activation of extracellular signal-regulated kinases (ERK1,2. Methods RAW264.7 macrophages were exposed to MWCNTs or carbon black nanoparticles (CBNPs over a range of doses and time course. Uptake and subcellular localization of MWCNTs was visualized by transmission electron microscopy (TEM. Protein levels of COX-2, iNOS, and ERK1,2 (total ERK and phosphorylated ERK were measured by Western blot analysis. Prostaglandin-E2 (PGE2 and nitric oxide (NO levels in cell supernatants were measured by ELISA and Greiss assay, respectively. Results MWCNTs, but not CBNPs, induced COX-2 and iNOS in a time- and dose-dependent manner. COX-2 and iNOS induction by MWCNTs correlated with increased PGE2 and NO production, respectively. MWCNTs caused ERK1,2 activation and inhibition of ERK1,2 (U0126 blocked MWCNT induction of COX-2 and PGE2 production, but did not reduce the induction of iNOS. Inhibition of iNOS (L-NAME did not affect ERK1,2 activation, nor did L-NAME significantly decrease COX-2 induction by MWCNT. Nickel nanoparticles (NiNPs, which are present in MWCNTs as a residual catalyst, also induced COX-2 via ERK-1,2. However, a comparison of COX-2 induction by MWCNTs containing 4.5 and 1.8% Ni did not show a significant difference in ability to induce COX-2, indicating that characteristics of MWCNTs in addition to Ni content contribute to COX-2 induction. Conclusion This study identifies COX-2 and subsequent PGE2 production, along with iNOS induction and NO production, as inflammatory mediators involved in the macrophage response to MWCNTs. Furthermore, our work demonstrates that COX-2 induction by MWCNTs in RAW264.7 macrophages is ERK1,2-dependent, while iNOS induction by MWCNTs is ERK1,2-independent. Our data also suggest contributory physicochemical factors other than residual Ni catalyst play a role in COX-2 induction to MWCNT.

  3. Novel immune check-point regulators in tolerance maintenance

    Directory of Open Access Journals (Sweden)

    Yanxia eGuo

    2015-08-01

    Full Text Available The great success of anti-cytotoxic lymphocyte antigen 4 (CTLA4 and anti-programmed cell death protein 1 (PD1 in cancer treatment has encouraged more effort in harnessing the immune response through immunomodulatory molecules in various diseases. The immunoglobulin (Ig super family comprises the majority of immunomodulatory molecules. Discovery of novel Ig super family members has brought novel insights into the function of different immune cells in tolerance maintenance. In this review, we discuss the function of newly-identified B7 family molecules B7-H4 and V-domain Ig Suppressor of T cell Activation (VISTA, and the butyrophilin/butyrophilin-like (BTN/BTNL family members. We discuss the current stages of immunomodulatory molecules in clinical trials of organ transplantation. The potential of engaging the novel Ig superfamily members in tolerance maintenance is also discussed. We conclude with the challenges remaining to manipulate these molecules in the immune response.

  4. A Metabolic Immune Checkpoint: Adenosine in Tumor Microenvironment

    Science.gov (United States)

    Ohta, Akio

    2016-01-01

    Within tumors, some areas are less oxygenated than others. Since their home ground is under chronic hypoxia, tumor cells adapt to this condition by activating aerobic glycolysis; however, this hypoxic environment is very harsh for incoming immune cells. Deprivation of oxygen limits availability of energy sources and induces accumulation of extracellular adenosine in tumors. Extracellular adenosine, upon binding with adenosine receptors on the surface of various immune cells, suppresses pro-inflammatory activities. In addition, signaling through adenosine receptors upregulates a number of anti-inflammatory molecules and immunoregulatory cells, leading to the establishment of a long-lasting immunosuppressive environment. Thus, due to hypoxia and adenosine, tumors can discourage antitumor immune responses no matter how the response was induced, whether it was spontaneous or artificially introduced with a therapeutic intention. Preclinical studies have shown the significance of adenosine in tumor survival strategy by demonstrating tumor regression after inactivation of adenosine receptors, inhibition of adenosine-producing enzymes, or reversal of tissue hypoxia. These promising results indicate a potential use of the inhibitors of the hypoxia–adenosine pathway for cancer immunotherapy. PMID:27066002

  5. Developmental Checkpoints and Feedback Circuits Time Insect Maturation

    DEFF Research Database (Denmark)

    Rewitz, Kim F.; Yamanaka, Naoki; O'Connor, Michael B.

    2013-01-01

    Abstract The transition from juvenile to adult is a fundamental process that allows animals to allocate resource toward reproduction after completing a certain amount of growth. In insects, growth to a species-specific target size induces pulses of the steroid hormone ecdysone that triggers...

  6. Putting checkpoints to work in thread level speculative execution

    OpenAIRE

    Khan, Salman

    2010-01-01

    With the advent of Chip Multi Processors (CMPs), improving performance relies on the programmers/compilers to expose thread level parallelism to the underlying hardware. Unfortunately, this is a difficult and error-prone process for the programmers, while state of the art compiler techniques are unable to provide significant benefits for many classes of applications. An interesting alternative is offered by systems that support Thread Level Speculation (TLS), which relieve the ...

  7. OOcyte Spindle Assembly Checkpoint in Space and Time

    Czech Academy of Sciences Publication Activity Database

    Anger, Martin; Awadová, S.; Kovačovicová, K.

    Prague : Czechoslovak Microscopy Society, 2014. s. 903-903. [International Microscopy Congress /18./. 07.09.2014-12.09.2014, Prague] R&D Projects: GA ČR GAP502/12/2201 Institutional support: RVO:67985904 Keywords : aneuploidy Subject RIV: EB - Genetics ; Molecular Biology

  8. Checkpoint NG VPN 1firewall 1 advanced configuration and troubleshooting

    CERN Document Server

    Syngress

    2003-01-01

    Check Point Software Technologies is the worldwide leader in securing the Internet. The company''s Secure Virtual Network (SVN) architecture provides the infrastructure that enables secure and reliable Internet communications. Check Point recently announced a ground-breaking user interface that meets the industry''s next generation Internet security requirements, including simplified security management for increasingly complex environments. Built upon Check Point''s Secure Virtual Network (SVN) architecture, the Next Generation User Interface revolutionizes the way security administrators define and manage enterprise security by further integrating management functions into a security dashboard and creating a visual picture of security operations. The Next Generation User Interface delivers unparalleled ease-of-use, improved security and true end-to-end security management. Check Point''s revenues have more than doubled in each of the last two years, while capturing over 50% of the VPN market and over 40% of...

  9. A cell cycle and nutritional checkpoint controlling bacterial surface adhesion.

    Directory of Open Access Journals (Sweden)

    Aretha Fiebig

    2014-01-01

    Full Text Available In natural environments, bacteria often adhere to surfaces where they form complex multicellular communities. Surface adherence is determined by the biochemical composition of the cell envelope. We describe a novel regulatory mechanism by which the bacterium, Caulobacter crescentus, integrates cell cycle and nutritional signals to control development of an adhesive envelope structure known as the holdfast. Specifically, we have discovered a 68-residue protein inhibitor of holdfast development (HfiA that directly targets a conserved glycolipid glycosyltransferase required for holdfast production (HfsJ. Multiple cell cycle regulators associate with the hfiA and hfsJ promoters and control their expression, temporally constraining holdfast development to the late stages of G1. HfiA further functions as part of a 'nutritional override' system that decouples holdfast development from the cell cycle in response to nutritional cues. This control mechanism can limit surface adhesion in nutritionally sub-optimal environments without affecting cell cycle progression. We conclude that post-translational regulation of cell envelope enzymes by small proteins like HfiA may provide a general means to modulate the surface properties of bacterial cells.

  10. More to Life than NF-κB in TNFR1 Signaling.

    Science.gov (United States)

    Ting, Adrian T; Bertrand, Mathieu J M

    2016-08-01

    TNF is a master proinflammatory cytokine whose pathogenic role in inflammatory disorders has long been attributed to induction of proinflammatory mediators. TNF also activates cell survival and death pathways, and recent studies demonstrated that TNF also causes inflammation by inducing cell death. The default response of most cells to TNF is survival and NF-κB-mediated upregulation of prosurvival molecules is a well-documented protective mechanism downstream of TNFR1. Recent studies revealed the existence of an NF-κB-independent cell death checkpoint that restricts cell demise by inactivating RIPK1. Disruption of this checkpoint leads to RIPK1 kinase-dependent death and causes inflammation in vivo. These revelations bring complexity to the control of TNF-induced cell death, and suggest clinical benefit of RIPK1 inhibitors in TNF-driven human inflammatory disorders. PMID:27424290

  11. Transforming Growth Factor β1 Inhibits Cystic Fibrosis Transmembrane Conductance Regulator-dependent cAMP-stimulated Alveolar Epithelial Fluid Transport via a Phosphatidylinositol 3-Kinase-dependent Mechanism*

    OpenAIRE

    Roux, Jérémie; Carles, Michel; Koh, Hidefumi; Goolaerts, Arnaud; Ganter, Michael T.; Chesebro, Brian B; Howard, Marybeth; Houseman, Benjamin T.; Finkbeiner, Walter; Shokat, Kevan M.; Paquet, Agnès C.; Matthay, Michael A; Pittet, Jean-François

    2009-01-01

    Exogenous or endogenous β2-adrenergic receptor agonists enhance alveolar epithelial fluid transport via a cAMP-dependent mechanism that protects the lungs from alveolar flooding in acute lung injury. However, impaired alveolar fluid clearance is present in most of the patients with acute lung injury and is associated with increased mortality, although the mechanisms responsible for this inhibition of the alveolar epithelial fluid transport are not completely understood. Here, we found that tr...

  12. Regulation of Lymphoid Enhancer Factor 1/T-Cell Factor by Mitogen-Activated Protein Kinase-Related Nemo-Like Kinase-Dependent Phosphorylation in Wnt/β-Catenin Signaling

    OpenAIRE

    Ishitani, Tohru; Ninomiya-Tsuji, Jun; Matsumoto, Kunihiro

    2003-01-01

    The Wnt/β-catenin signaling pathway regulates many developmental processes by modulating gene expression. Wnt signaling induces the stabilization of cytosolic β-catenin, which then associates with lymphoid enhancer factor and T-cell factor (LEF-1/TCF) to form a transcription complex that activates Wnt target genes. Previously, we have shown that a specific mitogen-activated protein (MAP) kinase pathway involving the MAP kinase kinase kinase TAK1 and MAP kinase-related Nemo-like kinase (NLK) s...

  13. Oncogenic Herpesvirus Utilizes Stress-Induced Cell Cycle Checkpoints for Efficient Lytic Replication.

    Science.gov (United States)

    Balistreri, Giuseppe; Viiliäinen, Johanna; Turunen, Mikko; Diaz, Raquel; Lyly, Lauri; Pekkonen, Pirita; Rantala, Juha; Ojala, Krista; Sarek, Grzegorz; Teesalu, Mari; Denisova, Oxana; Peltonen, Karita; Julkunen, Ilkka; Varjosalo, Markku; Kainov, Denis; Kallioniemi, Olli; Laiho, Marikki; Taipale, Jussi; Hautaniemi, Sampsa; Ojala, Päivi M

    2016-02-01

    Kaposi's sarcoma herpesvirus (KSHV) causes Kaposi's sarcoma and certain lymphoproliferative malignancies. Latent infection is established in the majority of tumor cells, whereas lytic replication is reactivated in a small fraction of cells, which is important for both virus spread and disease progression. A siRNA screen for novel regulators of KSHV reactivation identified the E3 ubiquitin ligase MDM2 as a negative regulator of viral reactivation. Depletion of MDM2, a repressor of p53, favored efficient activation of the viral lytic transcription program and viral reactivation. During lytic replication cells activated a p53 response, accumulated DNA damage and arrested at G2-phase. Depletion of p21, a p53 target gene, restored cell cycle progression and thereby impaired the virus reactivation cascade delaying the onset of virus replication induced cytopathic effect. Herpesviruses are known to reactivate in response to different kinds of stress, and our study now highlights the molecular events in the stressed host cell that KSHV has evolved to utilize to ensure efficient viral lytic replication. PMID:26891221

  14. A DNA damage-induced, SOS-independent checkpoint regulates cell division in Caulobacter crescentus.

    Directory of Open Access Journals (Sweden)

    Joshua W Modell

    2014-10-01

    Full Text Available Cells must coordinate DNA replication with cell division, especially during episodes of DNA damage. The paradigm for cell division control following DNA damage in bacteria involves the SOS response where cleavage of the transcriptional repressor LexA induces a division inhibitor. However, in Caulobacter crescentus, cells lacking the primary SOS-regulated inhibitor, sidA, can often still delay division post-damage. Here we identify didA, a second cell division inhibitor that is induced by DNA damage, but in an SOS-independent manner. Together, DidA and SidA inhibit division, such that cells lacking both inhibitors divide prematurely following DNA damage, with lethal consequences. We show that DidA does not disrupt assembly of the division machinery and instead binds the essential division protein FtsN to block cytokinesis. Intriguingly, mutations in FtsW and FtsI, which drive the synthesis of septal cell wall material, can suppress the activity of both SidA and DidA, likely by causing the FtsW/I/N complex to hyperactively initiate cell division. Finally, we identify a transcription factor, DriD, that drives the SOS-independent transcription of didA following DNA damage.

  15. A DNA damage-induced, SOS-independent checkpoint regulates cell division in Caulobacter crescentus.

    Science.gov (United States)

    Modell, Joshua W; Kambara, Tracy K; Perchuk, Barrett S; Laub, Michael T

    2014-10-01

    Cells must coordinate DNA replication with cell division, especially during episodes of DNA damage. The paradigm for cell division control following DNA damage in bacteria involves the SOS response where cleavage of the transcriptional repressor LexA induces a division inhibitor. However, in Caulobacter crescentus, cells lacking the primary SOS-regulated inhibitor, sidA, can often still delay division post-damage. Here we identify didA, a second cell division inhibitor that is induced by DNA damage, but in an SOS-independent manner. Together, DidA and SidA inhibit division, such that cells lacking both inhibitors divide prematurely following DNA damage, with lethal consequences. We show that DidA does not disrupt assembly of the division machinery and instead binds the essential division protein FtsN to block cytokinesis. Intriguingly, mutations in FtsW and FtsI, which drive the synthesis of septal cell wall material, can suppress the activity of both SidA and DidA, likely by causing the FtsW/I/N complex to hyperactively initiate cell division. Finally, we identify a transcription factor, DriD, that drives the SOS-independent transcription of didA following DNA damage. PMID:25350732

  16. The DNA damage checkpoint precedes activation of ARF in response to escalating oncogenic stress during tumorigenesis

    Czech Academy of Sciences Publication Activity Database

    Evangelou, K.; Bartkova, J.; Kotsinas, A.; Pateras, I.S.; Liontos, M.; Velimezi, G.; Košař, Martin; Liloglou, T.; Trougakos, I.P.; Dyrskjot, L.; Andersen, C.J.; Papaioannou, G.; Drosos, A.; Papafotiou, M.; Hodný, Zdeněk; Sosa-Pineda, B.; Wu, X.R.; Klinakis, A.; Orntoft, T.; Lukas, J.; Bartek, Jiří; Gorgoulis, V.G.

    2013-01-01

    Roč. 20, č. 11 (2013), s. 1485-1497. ISSN 1350-9047 Institutional support: RVO:68378050 Keywords : ARF * carcinogenesis * DDR * E2F1 * p16(INK4A) Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 8.385, year: 2013

  17. PSGL-1 Is an Immune Checkpoint Regulator that Promotes T Cell Exhaustion.

    Science.gov (United States)

    Tinoco, Roberto; Carrette, Florent; Barraza, Monique L; Otero, Dennis C; Magaña, Jonathan; Bosenberg, Marcus W; Swain, Susan L; Bradley, Linda M

    2016-05-17

    Chronic viruses and cancers thwart immune responses in humans by inducing T cell dysfunction. Using a murine chronic virus that models human infections, we investigated the function of the adhesion molecule, P-selectin glycoprotein ligand-1 (PSGL-1), that is upregulated on responding T cells. PSGL-1-deficient mice cleared the virus due to increased intrinsic survival of multifunctional effector T cells that had downregulated PD-1 as well as other inhibitory receptors. Notably, this response resulted in CD4(+)-T-cell-dependent immunopathology. Mechanistically, PSGL-1 ligation on exhausted CD8(+) T cells inhibited T cell receptor (TCR) and interleukin-2 (IL-2) signaling and upregulated PD-1, leading to diminished survival with TCR stimulation. In models of melanoma cancer in which T cell dysfunction occurs, PSGL-1 deficiency led to PD-1 downregulation, improved T cell responses, and tumor control. Thus, PSGL-1 plays a fundamental role in balancing viral control and immunopathology and also functions to regulate T cell responses in the tumor microenvironment. PMID:27192578

  18. Time-Memory Trade-Offs: False Alarm Detection Using Checkpoints, Extended Version

    OpenAIRE

    Avoine, Gildas; Junod, Pascal; Oechslin, Philippe

    2005-01-01

    Since the original publication of Martin Hellman's cryptanalytic time-memory trade-off, a few improvements on the method have been suggested. In all these variants, the cryptanalysis time decreases with the square of the available memory. However, a large amount of work is wasted during the cryptanalysis process due to so-called "false alarms". In this paper we present a method of detection of false alarms which can significantly reduce the cryptanalysis time while using a minute amount of me...

  19. Stable MCC binding to the APC/C is required for a functional spindle assembly checkpoint

    DEFF Research Database (Denmark)

    Hein, Jamin B; Nilsson, Jakob

    2014-01-01

    stably to the APC/C. Whether MCC formation per se is sufficient for a functional SAC or MCC association with the APC/C is required remains unclear. Here, we analyze the role of two conserved motifs in Cdc20, IR and C-Box, in binding of the MCC to the APC/C. Mutants in both motifs assemble the MCC...

  20. A checkpoint control orchestrates the replication of the two chromosomes of Vibrio cholerae

    DEFF Research Database (Denmark)

    Val, Marie-Eve; Marbouty, Martial; Martins, Francisco de Lemos;

    2016-01-01

    important differences between plasmids and chromosomes is that the latter replicate during a defined period of the cell cycle, ensuring a single round of replication per cell. Vibrio cholerae carries two circular chromosomes, Chr1 and Chr2, which are replicated in a well-orchestrated manner with the cell...

  1. The DNA damage checkpoint precedes activation of ARF in response to escalating oncogenic stress during tumorigenesis

    DEFF Research Database (Denmark)

    Evangelou, K.; Bartkova, J.; Kotsinas, A.;

    2013-01-01

    Oncogenic stimuli trigger the DNA damage response (DDR) and induction of the alternative reading frame (ARF) tumor suppressor, both of which can activate the p53 pathway and provide intrinsic barriers to tumor progression. However, the respective timeframes and signal thresholds for ARF induction...

  2. Checkpoint kinase inhibitors: SAR and radioprotective properties of a series of 2-arylbenzimidazoles.

    Science.gov (United States)

    Arienti, Kristen L; Brunmark, Anders; Axe, Frank U; McClure, Kelly; Lee, Alice; Blevitt, Jon; Neff, Danielle K; Huang, Liming; Crawford, Shelby; Pandit, Chennagiri R; Karlsson, Lars; Breitenbucher, J Guy

    2005-03-24

    The discovery of a series of novel, potent, and highly selective inhibitors of the DNA damage control kinase chk2 is disclosed. Here we report the first SAR study around inhibitors of this kinase. High-throughput screening of purified human chk2 led to the identification of a novel series of 2-arylbenzimidazole inhibitors of the kinase. Optimization was facilitated using homology models of chk2 and docking of inhibitors, leading to the highly potent 2-arylbenzimidazole 2h (IC(50) 15 nM). Compound 2h is an ATP-competitive inhibitor of chk2 that dose dependently protects human CD4(+) and CD8(+) T-cells from apoptosis due to ionizing radiation. This work suggests that a selective small molecule inhibitor of chk2 could be a useful adjuvant to radiotherapy, increasing the therapeutic window of such treatment. PMID:15771432

  3. Activation of a DNA damage checkpoint response in a TAF1-defective cell line.

    Science.gov (United States)

    Buchmann, Ann M; Skaar, Jeffrey R; DeCaprio, James A

    2004-06-01

    Although the link between transcription and DNA repair is well established, defects in the core transcriptional complex itself have not been shown to elicit a DNA damage response. Here we show that a cell line with a temperature-sensitive defect in TBP-associated factor 1 (TAF1), a component of the TFIID general transcription complex, exhibits hallmarks of an ATR-mediated DNA damage response. Upon inactivation of TAF1, ATR rapidly localized to subnuclear foci and contributed to the phosphorylation of several downstream targets, including p53 and Chk1, resulting in cell cycle arrest. The increase in p53 expression and the G(1) phase arrest could be blocked by caffeine, an inhibitor of ATR. In addition, dominant negative forms of ATR but not ATM were able to override the arrest in G(1). These results suggest that a defect in TAF1 can elicit a DNA damage response. PMID:15169897

  4. Activation of a DNA Damage Checkpoint Response in a TAF1-Defective Cell Line

    OpenAIRE

    Buchmann, Ann M.; Skaar, Jeffrey R.; DeCaprio, James A.

    2004-01-01

    Although the link between transcription and DNA repair is well established, defects in the core transcriptional complex itself have not been shown to elicit a DNA damage response. Here we show that a cell line with a temperature-sensitive defect in TBP-associated factor 1 (TAF1), a component of the TFIID general transcription complex, exhibits hallmarks of an ATR-mediated DNA damage response. Upon inactivation of TAF1, ATR rapidly localized to subnuclear foci and contributed to the phosphoryl...

  5. Regulation of APC/C activity in oocytes by a Bub1-dependent spindle assembly checkpoint

    Czech Academy of Sciences Publication Activity Database

    McGuinness, B. E.; Anger, Martin; Kouznetsova, A.; Gil-Barnabé, A. M.; Helmhart, W.; Kudo, Nobuaki R.; Wuensche, A.; Taylor, S.; Hoog, Ch.; Novak, B.; Nasmyth, K.

    2009-01-01

    Roč. 19, č. 5 (2009), s. 369-380. ISSN 0960-9822 Institutional research plan: CEZ:AV0Z50450515 Keywords : Oocyte * Aneuploidy Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 10.992, year: 2009

  6. Checkpoints in Ohm's Law%欧姆定律考点聚集

    Institute of Scientific and Technical Information of China (English)

    刘跃; 杨元俊

    2004-01-01

    @@ 欧姆定律这一章是在介绍了电流、电压、电阻的基础上,通过实验、归纳、总结这三个重要物理量之间的关系.欧姆定律是初中物理的重点又是难点,它贯穿于整个电学的计算,是中考的热点和必考内容,在中考试卷中的题量很大,题型也多种多样,包括填空、选择、实验、问答、计算题等.且常常与电功、电功率和焦耳定律等知识结合起来,作为压轴题的频率较高.综观近几年各省、市的中考题,命题主要包含以下几个知识点.

  7. Eukaryotic checkpoints are absent in the cell division cycle of Entamoeba histolytica

    Indian Academy of Sciences (India)

    Sulagna Banerjee; Suchismita Das; Anuradha Lohia

    2002-11-01

    Fidelity in transmission of genetic characters is ensured by the faithful duplication of the genome, followed by equal segregation of the genetic material in the progeny. Thus, alternation of DNA duplication (S-phase) and chromosome segregation during the M-phase are hallmarks of most well studied eukaryotes. Several rounds of genome reduplication before chromosome segregation upsets this cycle and leads to polyploidy. Polyploidy is often witnessed in cells prior to differentiation, in embryonic cells or in diseases such as cancer. Studies on the protozoan parasite, Entamoeba histolytica suggest that in its proliferative phase, this organism may accumulate polyploid cells. It has also been shown that although this organism contains sequence homologs of genes which are known to control the cell cycle of most eukaryotes, these genes may be structurally altered and their equivalent function yet to be demonstrated in amoeba. The available information suggests that surveillance mechanisms or ‘checkpoints’ which are known to regulate the eukaryotic cell cycle may be absent or altered in E. histolytica.

  8. Diffusion MRI and novel texture analysis in osteosarcoma xenotransplants predicts response to anti-checkpoint therapy.

    Science.gov (United States)

    Foroutan, Parastou; Kreahling, Jenny M; Morse, David L; Grove, Olya; Lloyd, Mark C; Reed, Damon; Raghavan, Meera; Altiok, Soner; Martinez, Gary V; Gillies, Robert J

    2013-01-01

    Combinations of targeted drugs have been employed to treat sarcomas, however, response rates have not improved notably, therefore emphasizing the need for novel treatments. In addition, imaging approaches to assess therapeutic response is lacking, as currently measurable indices, such as volume and/or diameter, do not accurately correlate with changes in tumor biology. In this study, quantitative and profound analyses of magnetic resonance imaging (MRI) were developed to evaluate these as imaging biomarkers for MK1775 and Gem in an osteosarcoma xenotransplant model at early time-points following treatment. Notably, we showed that Gem and Gem+MK1775 groups had significantly inhibited tumor growth by day 4, which was presaged by elevations in mean ADC by 24 hours post treatment. Significant differences were also observed at later time points for the Gem+MK1775 combination and MK1775 therapy. ADC distribution and entropy (randomness of ADC values) were also elevated by 24 hours following therapy. Immunohistochemistry demonstrated that these treatment-related increases in ADC correlated with apoptosis and observed cell condensations (dense- and exploded bodies). These findings underline the role of ADC as a quantitative imaging biomarker for therapy-induced response and show promising clinical relevance in the sarcoma patient population. PMID:24358232

  9. Polycomb proteins control proliferation and transformation independently of cell cycle checkpoints by regulating DNA replication

    DEFF Research Database (Denmark)

    Piunti, Andrea; Rossi, Alessandra; Cerutti, Aurora;

    2014-01-01

    PRCs regulate cellular proliferation and transformation independently of the Ink4a/Arf-pRb-p53 pathway. We provide evidence that PRCs localize at replication forks, and that loss of their function directly affects the progression and symmetry of DNA replication forks. Thus, we have identified a novel...

  10. CtIP-dependent DNA resection is required for DNA damage checkpoint maintenance but not initiation

    DEFF Research Database (Denmark)

    Kousholt, Arne Nedergaard; Fugger, Kasper; Hoffmann, Saskia;

    2012-01-01

    To prevent accumulation of mutations, cells respond to DNA lesions by blocking cell cycle progression and initiating DNA repair. Homology-directed repair of DNA breaks requires CtIP-dependent resection of the DNA ends, which is thought to play a key role in activation of ATR (ataxia telangiectasia...

  11. Oncogene-induced senescence is part of the tumorigenesis barrier imposed by DNA damage checkpoints

    DEFF Research Database (Denmark)

    Bartkova, Jirina; Rezaei, Nousin; Liontos, Michalis;

    2006-01-01

    DNA double-strand breaks. Inhibiting the DNA double-strand break response kinase ataxia telangiectasia mutated (ATM) suppressed the induction of senescence and in a mouse model led to increased tumour size and invasiveness. Analysis of human precancerous lesions further indicated that DNA damage and...

  12. Immune checkpoint inhibitors: the new frontier in non–small cell lung cancer treatment

    OpenAIRE

    El-Osta, Hazem; Shahid,Kamran; Mills, Glenn; Peddi, Prakash

    2016-01-01

    Hazem El-Osta, Kamran Shahid, Glenn M Mills, Prakash Peddi Department of Medicine, Division of Hematology-Oncology, Louisiana State University Health Sciences Center, Shreveport, LA, USA Abstract: Lung cancer is the major cause for cancer-related death in the US. Although advances in chemotherapy and targeted therapy have improved the outcome of metastatic non-small-cell lung cancer, its prognosis remains dismal. A deeper understanding of the complex interaction between the immune system an...

  13. Arginase 1 is an innate lymphoid-cell-intrinsic metabolic checkpoint controlling type 2 inflammation.

    Science.gov (United States)

    Monticelli, Laurel A; Buck, Michael D; Flamar, Anne-Laure; Saenz, Steven A; Tait Wojno, Elia D; Yudanin, Naomi A; Osborne, Lisa C; Hepworth, Matthew R; Tran, Sara V; Rodewald, Hans-Reimer; Shah, Hardik; Cross, Justin R; Diamond, Joshua M; Cantu, Edward; Christie, Jason D; Pearce, Erika L; Artis, David

    2016-06-01

    Group 2 innate lymphoid cells (ILC2s) regulate tissue inflammation and repair after activation by cell-extrinsic factors such as host-derived cytokines. However, the cell-intrinsic metabolic pathways that control ILC2 function are undefined. Here we demonstrate that expression of the enzyme arginase-1 (Arg1) during acute or chronic lung inflammation is a conserved trait of mouse and human ILC2s. Deletion of mouse ILC-intrinsic Arg1 abrogated type 2 lung inflammation by restraining ILC2 proliferation and dampening cytokine production. Mechanistically, inhibition of Arg1 enzymatic activity disrupted multiple components of ILC2 metabolic programming by altering arginine catabolism, impairing polyamine biosynthesis and reducing aerobic glycolysis. These data identify Arg1 as a key regulator of ILC2 bioenergetics that controls proliferative capacity and proinflammatory functions promoting type 2 inflammation. PMID:27043409

  14. Oncogenic Herpesvirus Utilizes Stress-Induced Cell Cycle Checkpoints for Efficient Lytic Replication.

    Directory of Open Access Journals (Sweden)

    Giuseppe Balistreri

    2016-02-01

    Full Text Available Kaposi's sarcoma herpesvirus (KSHV causes Kaposi's sarcoma and certain lymphoproliferative malignancies. Latent infection is established in the majority of tumor cells, whereas lytic replication is reactivated in a small fraction of cells, which is important for both virus spread and disease progression. A siRNA screen for novel regulators of KSHV reactivation identified the E3 ubiquitin ligase MDM2 as a negative regulator of viral reactivation. Depletion of MDM2, a repressor of p53, favored efficient activation of the viral lytic transcription program and viral reactivation. During lytic replication cells activated a p53 response, accumulated DNA damage and arrested at G2-phase. Depletion of p21, a p53 target gene, restored cell cycle progression and thereby impaired the virus reactivation cascade delaying the onset of virus replication induced cytopathic effect. Herpesviruses are known to reactivate in response to different kinds of stress, and our study now highlights the molecular events in the stressed host cell that KSHV has evolved to utilize to ensure efficient viral lytic replication.

  15. Oncogenic Herpesvirus Utilizes Stress-Induced Cell Cycle Checkpoints for Efficient Lytic Replication.

    OpenAIRE

    Giuseppe Balistreri; Johanna Viiliäinen; Mikko Turunen; Raquel Diaz; Lauri Lyly; Pirita Pekkonen; Juha Rantala; Krista Ojala; Grzegorz Sarek; Mari Teesalu; Oxana Denisova; Karita Peltonen; Ilkka Julkunen; Markku Varjosalo; Denis Kainov

    2016-01-01

    Kaposi’s sarcoma herpesvirus (KSHV) causes Kaposi’s sarcoma and certain lymphoproliferative malignancies. Latent infection is established in the majority of tumor cells, whereas lytic replication is reactivated in a small fraction of cells, which is important for both virus spread and disease progression. A siRNA screen for novel regulators of KSHV reactivation identified the E3 ubiquitin ligase MDM2 as a negative regulator of viral reactivation. Depletion of MDM2, a repressor of p53, favored...

  16. Social and ethical checkpoints for bottom-up synthetic biology, or protocells

    OpenAIRE

    Bedau M.A.; Parke E.C.; Tangen U.; Hantsche-Tangen B.

    2009-01-01

    An alternative to creating novel organisms through the traditional “top-down” approach to synthetic biology involves creating them from the “bottom up” by assembling them from non-living components; the products of this approach are called “protocells.” In this paper we describe how bottom-up and top-down synthetic biology differ, review the current state of protocell research and development, and examine the unique ethical, social, and regulatory issues raised by bottom-up synthetic biology....

  17. ATR Mediates a Checkpoint at the Nuclear Envelope in Response to Mechanical Stress

    Czech Academy of Sciences Publication Activity Database

    Kumar, A.; Mazzanti, M.; Mistrik, M.; Košař, Martin; Beznoussenko, G.V.; Mironov, A. A.; Garrè, M.; Parazolli, D.; Shivashankar, G. V.; Scita, G.; Bartek, Jiří; Foiani, M.

    2014-01-01

    Roč. 158, č. 3 (2014), s. 633-646. ISSN 0092-8674 Grant ostatní: Marie Curie Intra-European(IT) 274093 Institutional support: RVO:68378050 Keywords : ATR * Mechanical Stress * cell cycle Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 32.242, year: 2014

  18. GATA-3 dose-dependent checkpoints in early T cell commitment1

    OpenAIRE

    Scripture-Adams, Deirdre D.; Damle, Sagar S.; Li, Long; Elihu, Koorosh J.; Qin, Shuyang; Arias, Alexandra M.; Butler, Robert R.; Champhekar, Ameya; Zhang, Jingli A.; Rothenberg, Ellen V.

    2014-01-01

    GATA-3 expression is crucial for T cell development and peaks during commitment to the T-cell lineage, midway through the CD4−CD8− (DN) 1-3 stages. We used RNA interference and conditional deletion to reduce GATA-3 protein acutely at specific points during T-cell differentiation in vitro. Even moderate GATA-3 reduction killed DN1 cells, delayed progression to DN2 stage, skewed DN2 gene regulation, and blocked appearance of DN3 phenotype. Although a Bcl-2 transgene rescued DN1 survival and imp...

  19. Sequence Classification: 894788 [

    Lifescience Database Archive (English)

    Full Text Available volved in transcription-coupled repair nucleotide excision repair of UV-induced DNA lesions; homolog of human CSB protein; Rad26p || http://www.ncbi.nlm.nih.gov/protein/6322495 ...

  20. Gain-of-function mutations of PPM1D/Wip1 impair the p53-dependent G1 checkpoint

    Czech Academy of Sciences Publication Activity Database

    Kleiblová, P.; Shaltiel, I.A.; Benada, Jan; Ševčík, J.; Pecháčková, Soňa; Pohlreich, P.; Voest, E.E.; Dundr, P.; Bartek, Jiří; Kleibl, Z.; Medema, R.H.; Macůrek, Libor

    2013-01-01

    Roč. 201, č. 4 (2013), s. 511-521. ISSN 0021-9525 R&D Projects: GA ČR GAP301/10/1525; GA ČR GAP305/12/2485; GA ČR GA13-18392S Grant ostatní: GA MZd(CZ) NT13343; EK(XE) 223575; EK(XE) ED0030/01/01 Institutional support: RVO:68378050 Keywords : DNA damage * cell cycle * cancer Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 9.688, year: 2013

  1. Chronic Exposure to Zinc Chromate Induces Centrosome Amplification and Spindle Assembly Checkpoint Bypass in Human Lung Fibroblasts

    OpenAIRE

    Holmes, Amie L.; Wise, Sandra S.; Pelsue, Stephen C.; Aboueissa, AbouEl-Makarim; Lingle, Wilma; Salisbury, Jeffery; Gallagher, Jamie; Wise, John Pierce

    2010-01-01

    Hexavalent chromium (Cr(VI)) compounds are known human lung carcinogens. Solubility plays an important role in its carcinogenicity with the particulate or insoluble form being the most potent. Of the particulate Cr(VI) compounds, zinc chromate appears to be the most potent carcinogen, however, very few studies have investigated its carcinogenic mechanism. In this study, we investigated the ability of chronic exposure to zinc chromate to induce numerical chromosome instability. We found no inc...

  2. ERK1/2 Signaling Plays an Important Role in Topoisomerase II Poison-Induced G2/M Checkpoint Activation

    OpenAIRE

    Kolb, Ryan H.; Greer, Patrick M.; Cao, Phu T.; Cowan, Kenneth H.; Ying Yan

    2012-01-01

    Topo II poisons, which target topoisomerase II (topo II) to generate enzyme mediated DNA damage, have been commonly used for anti-cancer treatment. While clinical evidence demonstrate a capability of topo II poisons in inducing apoptosis in cancer cells, accumulating evidence also show that topo II poison treatment frequently results in cell cycle arrest in cancer cells, which was associated with subsequent resistance to these treatments. Results in this report indicate that treatment of MCF-...

  3. Cop9/signalosome subunits and Pcu4 regulate ribonucleotide reductase by both checkpoint-dependent and -independent mechanisms

    OpenAIRE

    Liu, Cong; Powell, Kelly A.; Mundt, Kirsten; Wu, LeJung; Carr, Antony M.; Caspari, Thomas

    2003-01-01

    The signalosome is implicated in regulating cullin-dependent ubiquitin ligases. We find that two signalosome subunits, Csn1 and Csn2, are required to regulate ribonucleotide reductase (RNR) through the degradation of a small protein, Spd1, that acts to anchor the small RNR subunit in the nucleus. Spd1 destruction correlates with the nuclear export of the small RNR subunit, which, in turn, correlates with a requirement for RNR in replication and repair. Spd1 degradation is promoted by two sepa...

  4. The immune checkpoint regulator PD-L1 is a specific target for naturally occurring CD4+ T cells

    OpenAIRE

    Munir, Shamaila; Andersen, Gitte Holmen; Svane, Inge Marie; Andersen, Mads Hald

    2013-01-01

    Programmed cell death 1 ligand 1 (PD-L1) is an important regulator of T-cell responses and may consequently limit anticancer immunity. We have recently identified PD-L1-specific, cytotoxic CD8+ T cells. In the present study, we develop these findings and report that CD4+ helper T cells spontaneously recognize PD-L1. We examined the locality of a previously identified HLA-A*0201-restricted PD-L1-epitope for the presence of possible CD4+ T-cell epitopes. Thus, we identified naturally occurring ...

  5. The immune checkpoint regulator PD-L1 is a specific target for naturally occurring CD4(+) T cells

    DEFF Research Database (Denmark)

    Munir, Shamaila; Andersen, Gitte Holmen; Svane, Inge Marie; Andersen, Mads Hald

    2013-01-01

    Programmed cell death 1 ligand 1 (PD-L1) is an important regulator of T-cell responses and may consequently limit anticancer immunity. We have recently identified PD-L1-specific, cytotoxic CD8(+) T cells. In the present study, we develop these findings and report that CD4(+) helper T cells...... spontaneously recognize PD-L1. We examined the locality of a previously identified HLA-A*0201-restricted PD-L1-epitope for the presence of possible CD4(+) T-cell epitopes. Thus, we identified naturally occurring PD-L1-specific CD4(+) T cells among the peripheral blood lymphocytes of cancer patients and - to...... lesser extents - healthy donors, by means of ELISPOT assays. PD-L1-specific CD4(+) T cells appeared to be TH17 cells exhibiting an effector T-cell cytokine profile. Hence, PD-L1-specific CD4(+) T cells released interferon γ (IFNγ), tumor necrosis factor α (TNFα) and interleukin-17 (IL-17) in response to...

  6. Understanding the function and dysfunction of the immune system in lung cancer: the role of immune checkpoints

    International Nuclear Information System (INIS)

    Survival rates for metastatic lung cancer, including non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC), are poor with 5-year survivals of less than 5%. The immune system has an intricate and complex relationship with tumorigenesis; a groundswell of research on the immune system is leading to greater understanding of how cancer progresses and presenting new ways to halt disease progress. Due to the extraordinary power of the immune system—with its capacity for memory, exquisite specificity and central and universal role in human biology—immunotherapy has the potential to achieve complete, long-lasting remissions and cures, with few side effects for any cancer patient, regardless of cancer type. As a result, a range of cancer therapies are under development that work by turning our own immune cells against tumors. However deeper understanding of the complexity of immunomodulation by tumors is key to the development of effective immunotherapies, especially in lung cancer

  7. MITRAC7 Acts as a COX1-Specific Chaperone and Reveals a Checkpoint during Cytochrome c Oxidase Assembly.

    Science.gov (United States)

    Dennerlein, Sven; Oeljeklaus, Silke; Jans, Daniel; Hellwig, Christin; Bareth, Bettina; Jakobs, Stefan; Deckers, Markus; Warscheid, Bettina; Rehling, Peter

    2015-09-01

    Cytochrome c oxidase, the terminal enzyme of the respiratory chain, is assembled from mitochondria- and nuclear-encoded subunits. The MITRAC complex represents the central assembly intermediate during this process as it receives imported subunits and regulates mitochondrial translation of COX1 mRNA. The molecular processes that promote and regulate the progression of assembly downstream of MITRAC are still unknown. Here, we identify MITRAC7 as a constituent of a late form of MITRAC and as a COX1-specific chaperone. MITRAC7 is required for cytochrome c oxidase biogenesis. Surprisingly, loss of MITRAC7 or an increase in its amount causes selective cytochrome c oxidase deficiency in human cells. We demonstrate that increased MITRAC7 levels stabilize and trap COX1 in MITRAC, blocking progression in the assembly process. In contrast, MITRAC7 deficiency leads to turnover of newly synthesized COX1. Accordingly, MITRAC7 affects the biogenesis pathway by stabilizing newly synthesized COX1 in assembly intermediates, concomitantly preventing turnover. PMID:26321642

  8. Disruption of Murine mp29/Syf2/Ntc31 Gene Results in Embryonic Lethality with Aberrant Checkpoint Response

    OpenAIRE

    Chia-Hsin Chen; Po-Chen Chu; Liekyeow Lee; Huang-Wei Lien; Tse-Ling Lin; Chi-Chen Fan; Peter Chi; Chang-Jen Huang; Mau-Sun Chang

    2012-01-01

    Human p29 is a putative component of spliceosomes, but its role in pre-mRNA is elusive. By siRNA knockdown and stable overexpression, we demonstrated that human p29 is involved in DNA damage response and Fanconi anemia pathway in cultured cells. In this study, we generated p29 knockout mice (mp29(GT/GT)) using the mp29 gene trap embryonic stem cells to study the role of mp29 in DNA damage response in vivo. Interruption of mp29 at both alleles resulted in embryonic lethality. Embryonic abnorma...

  9. The Level of Europium-154 Contaminating Samarium-153-EDTMP Activates the Radiation Alarm System at the US Homeland Security Checkpoints

    OpenAIRE

    Najeeb Al Hallak, Mohammed; McCurdy, Matt; Zouain, Nicolas; Hayes, Justin

    2009-01-01

    153Sm-EDTMP is a radiopharmaceutical composed of EDTMP (ethylenediamine-tetramethylenephosphonate) and Samarium-153 [1]. 153Sm-EDTMP has an affinity for skeletal tissue and concentrates in areas with increased bone turnover; thus, it is successfully used in relieving pain related to diffuse bone metastases [1]. The manufacturing process of 153Sm-EDTMP leads to contamination with 154Eu (Europium-154) [2]. A previous study only alluded to the retention of 154Eu in the bones after receiving trea...

  10. Sox7 is an independent checkpoint for β-catenin function in prostate and colon epithelial cells

    OpenAIRE

    Guo, Lizheng; Diansheng ZHONG; Lau, Stephen; Liu, Xiuju; Dong, Xue-Yuan; Sun, Xiaodong; Yang, Vincent W.; Vertino, Paula M.; Moreno, Carlos; Varma, Vijay; Jin-Tang, Dong; Wei, Zhou

    2008-01-01

    The presence of somatic β-catenin mutations in some prostate cancers implies that aberrant WNT signaling is involved in the cancer’s development. Although β-catenin stability is regulated by a multi-component destruction complex, mutational alterations of β-catenin or other components of the destruction complexes are rare in prostate tumors. Therefore, β-catenin may be regulated by another protein in the prostate. In fact, recent linkage and somatic deletion analyses in prostate cancers revea...

  11. Cisplatin-Induced DNA Damage Activates Replication Checkpoint Signaling Components that Differentially Affect Tumor Cell SurvivalS⃞

    OpenAIRE

    Wagner, Jill M.; Karnitz, Larry M.

    2009-01-01

    Cisplatin and other platinating agents are some of the most widely used chemotherapy agents. These drugs exert their antiproliferative effects by creating intrastrand and interstrand DNA cross-links, which block DNA replication. The cross-links mobilize signaling and repair pathways, including the Rad9-Hus1-Rad1-ATR-Chk1 pathway, a pathway that helps tumor cells survive the DNA damage inflicted by many chemotherapy agents. Here we show that Rad9 and ATR play critical r...

  12. Serum-resistant CpG-STAT3 decoy for targeting survival and immune checkpoint signaling in acute myeloid leukemia.

    Science.gov (United States)

    Zhang, Qifang; Hossain, Dewan Md Sakib; Duttagupta, Priyanka; Moreira, Dayson; Zhao, Xingli; Won, Haejung; Buettner, Ralf; Nechaev, Sergey; Majka, Marcin; Zhang, Bin; Cai, Qi; Swiderski, Piotr; Kuo, Ya-Huei; Forman, Stephen; Marcucci, Guido; Kortylewski, Marcin

    2016-03-31

    Targeting oncogenic transcription factor signal transducer and activator of transcription 3 (STAT3) in acute myeloid leukemia (AML) can reduce blast survival and tumor immune evasion. Decoy oligodeoxynucleotides (dODNs), which comprise STAT3-specific DNA sequences are competitive inhibition of STAT3 transcriptional activity. To deliver STAT3dODN specifically to myeloid cells, we linked STAT3dODN to the Toll-like receptor 9 (TLR9) ligand, cytosine guanine dinucleotide (CpG). The CpG-STAT3dODN conjugates are quickly internalized by human and mouse TLR9(+)immune cells (dendritic cells, B cells) and the majority of patients' derived AML blasts, including leukemia stem/progenitor cells. Following uptake, CpG-STAT3dODNs are released from endosomes, and bind and sequester cytoplasmic STAT3, thereby inhibiting downstream gene expression in target cells. STAT3 inhibition in patients' AML cells limits their immunosuppressive potential by reduced arginase expression, thereby partly restoring T-cell proliferation. Partly chemically modified CpG-STAT3dODNs have >60 hours serum half-life which allows for IV administration to leukemia-bearing mice (50% effective dose ∼ 2.5 mg/kg). Repeated administration of CpG-STAT3dODN resulted in regression of human MV4-11 AML in mice. The antitumor efficacy of this strategy is further enhanced in immunocompetent mice by combining direct leukemia-specific cytotoxicity with immunogenic effects of STAT3 blocking/TLR9 triggering. CpG-STAT3dODN effectively reducedCbfb/MYH11/MplAML burden in various organs and eliminated leukemia stem/progenitor cells, mainly through CD8/CD4 T-cell-mediated immune responses. In contrast, small-molecule Janus kinase 2/STAT3 inhibitor failed to reproduce therapeutic effects of cell-selective CpG-STAT3dODN strategy. These results demonstrate therapeutic potential of CpG-STAT3dODN inhibitors with broad implications for treatement of AML and potentially other hematologic malignancies. PMID:26796361

  13. Understanding the function and dysfunction of the immune system in lung cancer:the role of immune checkpoints

    Institute of Scientific and Technical Information of China (English)

    Niki Karachaliou; Maria Gonzalez Cao; Cristina Teixid; Santiago Viteri; Daniela Morales-Espinosa; Mariacarmela Santarpia; Rafael Rosell

    2015-01-01

    AbstrAct Survival rates for metastatic lung cancer, including non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC), are poor with 5-year survivals of less than 5%. The immune system has an intricate and complex relationship with tumorigenesis;a groundswell of research on the immune system is leading to greater understanding of how cancer progresses and presenting new ways to halt disease progress. Due to the extraordinary power of the immune system—with its capacity for memory, exquisite speciifcity and central and universal role in human biology—immunotherapy has the potential to achieve complete, long-lasting remissions and cures, with few side effects for any cancer patient, regardless of cancer type. As a result, a range of cancer therapies are under development that work by turning our own immune cells against tumors. However deeper understanding of the complexity of immunomodulation by tumors is key to the development of effective immunotherapies, especially in lung cancer.

  14. Temporal Gradient in the Clock Gene and Cell-Cycle Checkpoint Kinase Wee1 Expression along the Gut

    Czech Academy of Sciences Publication Activity Database

    Polidarová, Lenka; Soták, Matúš; Sládek, Martin; Pácha, Jiří; Sumová, Alena

    2009-01-01

    Roč. 26, č. 4 (2009), s. 607-620. ISSN 0742-0528 R&D Projects: GA AV ČR(CZ) IAA500110605; GA ČR(CZ) GA305/09/0321; GA MŠk(CZ) LC554 EU Projects: European Commission(XE) 18741 - EUCLOCK Institutional research plan: CEZ:AV0Z50110509 Keywords : intestine epithelium * circadian clock gene * cell cycle Subject RIV: ED - Physiology Impact factor: 3.987, year: 2009

  15. The internal Cdc20 binding site in BubR1 facilitates both spindle assembly checkpoint signalling and silencing

    DEFF Research Database (Denmark)

    Lischetti, Tiziana; Zhang, Gang; Sedgwick, Garry G; Bolanos-Garcia, Victor M; Nilsson, Jakob

    2014-01-01

    in the SAC, we delineate the molecular details of its interaction with kinetochores. We find that BubR1 recruits the bulk of Cdc20 to kinetochores through its internal Cdc20 binding domain (IC20BD). We show that preventing Cdc20 kinetochore localization by removal of the IC20BD has a limited effect...

  16. RHINO forms a stoichiometric complex with the 9-1-1 checkpoint clamp and mediates ATR-Chk1 signaling

    OpenAIRE

    Lindsey-Boltz, Laura A.; Kemp, Michael G.; Capp, Christopher; Sancar, Aziz

    2015-01-01

    The ATR-Chk1 signaling pathway mediates cellular responses to DNA damage and replication stress and is composed of a number of core factors that are conserved throughout eukaryotic organisms. However, humans and other higher eukaryotic species possess additional factors that are implicated in the regulation of this signaling network but that have not been extensively studied. Here we show that RHINO (for Rad9, Rad1, Hus1 interacting nuclear orphan) forms complexes with both the 9-1-1 checkpoi...

  17. DNA-damage response network at the crossroads of cell-cycle checkpoints, cellular senescence and apoptosis*

    OpenAIRE

    Schmitt, Estelle; Paquet, Claudie; Beauchemin, Myriam; Bertrand, Richard

    2007-01-01

    Tissue homeostasis requires a carefully-orchestrated balance between cell proliferation, cellular senescence and cell death. Cells proliferate through a cell cycle that is tightly regulated by cyclin-dependent kinase activities. Cellular senescence is a safeguard program limiting the proliferative competence of cells in living organisms. Apoptosis eliminates unwanted cells by the coordinated activity of gene products that regulate and effect cell death. The intimate link between the cell cycl...

  18. [Therapy monitoring and management of adverse events in PD-1/PD-L1 immune checkpoint inhibition].

    Science.gov (United States)

    Oppel-Heuchel, H; Grimm, M-O

    2016-05-01

    Nivolumab was recently approved as the first inhibitor of the programmed death 1 (PD-1) receptor and its ligand (PD-L1) for the treatment of urological cancer, namely metastasized renal cell carcinoma after prior therapy. The use of this new immunotherapy requires special therapy monitoring and management of side effects. An increase of immune cells around the tumor can initially mimic progression (so-called pseudoprogression). Treatment-associated side effects of higher grade according to the common terminology criteria for adverse events (CTCAE grades 3 or 4) are relatively rare; however, new immune-mediated side effects can occur and affect the skin, liver (hepatitis), kidneys (nephritis), gastrointestinal tract (diarrhea and colitis), lungs (pneumonitis) and endocrine organs (hyperthyroidism, hypothyroidism and hypophysitis). Treatment has to be delayed or discontinued depending on the kind and degree of side effects; furthermore, corticosteroids can be administered as immunosuppressants. When recognized in time and with correct management, immune-mediated side effects are basically reversible. PMID:27146870

  19. Akt Switches TopBP1 Function from Checkpoint Activation to Transcriptional Regulation through Phosphoserine Binding-Mediated Oligomerization

    OpenAIRE

    Liu, Kang; Graves, Joshua D.; Scott, Jessica D.; Li, Rongbao; Lin, Weei-Chin

    2013-01-01

    Our previous study showed that Akt phosphorylates TopBP1 at the Ser-1159 residue and induces its oligomerization. Oligomerization is required for TopBP1 to bind and repress E2F1 activity. However, the mechanism through which phosphorylation of TopBP1 by Akt leads to its oligomerization remains to be determined. Here, we demonstrate that binding between the phosphorylated Ser-1159 (pS1159) residue and the 7th and 8th BRCT domains of TopBP1 mediates TopBP1 oligomerization. Mutations within the ...

  20. Therapeutic antitumor immunity by checkpoint blockade is enhanced by ibrutinib, an inhibitor of both BTK and ITK

    OpenAIRE

    Sagiv-Barfi, Idit; Kohrt, Holbrook E. K.; Czerwinski, Debra K.; Ng, Patrick P.; Chang, Betty Y.; Levy, Ronald

    2015-01-01

    Antibodies that block the negative signals between PD1-Ligand on tumor cells and PD-1 on T cells are effective therapies against several types of cancer. Ibrutinib, a covalent inhibitor of BTK is an approved therapy for B-cell leukemia and lymphoma. But ibrutinib also inactivates ITK, an enzyme required for certain subsets of T lymphocytes (Th2 T cells). We found that the combination of anti–PD-L1 antibodies and ibrutinib led to impressive therapeutic effects not only in animal models of lymp...

  1. Regulation of a senescence checkpoint response by the E2F1 transcription factor and p14ARF tumor suppressor

    Energy Technology Data Exchange (ETDEWEB)

    Dimri, Goberdhan P.; Itahana, Koji; Acosta, Meileen; Campisi, Judith

    1999-11-05

    Normal cells do not divide indefinitely due to a process known as replicative senescence. Human cells arrest growth with a senescent phenotype when they acquire one or more critically short telomere as a consequence of cell division. Recent evidence suggests that certain types of DNA damage, chromatin remodeling, or oncogenic forms of Rasor Raf can also elicit a senescence response. We show here that E2F1, a multifunctional transcription factor that binds the retinoblastoma (pRb) tumor suppressor and can either promote or suppress tumorigenesis, induces a senescent phenotype when overexpressed in normal human fibroblasts. Normal human cells stably arrested proliferation and expressed several markers of replicative senescence in response to E2F1. This activity of E2F1 was independent of its pRb binding activity, but dependent on its ability to stimulate gene expression. The E2F1 target gene critical for the senescence response appeared to be the p14ARF tumor suppressor. Replicatively senescent human fibroblasts overexpressed p14ARF, and ectopic expression of p14ARF in presenescent cells induced a phenotype similar to that induced by E2F1. Consistent with a critical role for p14ARF, cells with compromised p53 function were immune to senescence induction by E2F1, as were cells deficient in p14ARF. Our findings support the idea that the senescence response is a critical tumor suppressive mechanism, provide an explanation for the apparently paradoxical roles of E2F1 in oncogenesis, and identify p14ARF as a potentially important mediator of the senescent phenotype.

  2. Integrated genomic analysis identifies the mitotic checkpoint kinase WEE1 as a novel therapeutic target in medulloblastoma

    OpenAIRE

    Harris, Peter S; Venkataraman, Sujatha; Alimova, Irina; Birks, Diane K; Balakrishnan, Ilango; Cristiano, Brian; Donson, Andrew M; Dubuc, Adrian M.; Taylor, Michael D.; Foreman, Nicholas K.; Reigan, Philip; Vibhakar, Rajeev

    2014-01-01

    Background Medulloblastoma is the most common type of malignant brain tumor that afflicts children. Although recent advances in chemotherapy and radiation have improved outcomes, high-risk patients do poorly with significant morbidity. Methods To identify new molecular targets, we performed an integrated genomic analysis using structural and functional methods. Gene expression profiling in 16 medulloblastoma patient samples and subsequent gene set enrichment analysis indicated that cell cycle...

  3. MITRAC7 Acts as a COX1-Specific Chaperone and Reveals a Checkpoint during Cytochrome c Oxidase Assembly

    Directory of Open Access Journals (Sweden)

    Sven Dennerlein

    2015-09-01

    Full Text Available Cytochrome c oxidase, the terminal enzyme of the respiratory chain, is assembled from mitochondria- and nuclear-encoded subunits. The MITRAC complex represents the central assembly intermediate during this process as it receives imported subunits and regulates mitochondrial translation of COX1 mRNA. The molecular processes that promote and regulate the progression of assembly downstream of MITRAC are still unknown. Here, we identify MITRAC7 as a constituent of a late form of MITRAC and as a COX1-specific chaperone. MITRAC7 is required for cytochrome c oxidase biogenesis. Surprisingly, loss of MITRAC7 or an increase in its amount causes selective cytochrome c oxidase deficiency in human cells. We demonstrate that increased MITRAC7 levels stabilize and trap COX1 in MITRAC, blocking progression in the assembly process. In contrast, MITRAC7 deficiency leads to turnover of newly synthesized COX1. Accordingly, MITRAC7 affects the biogenesis pathway by stabilizing newly synthesized COX1 in assembly intermediates, concomitantly preventing turnover.

  4. MGT 245 uop Courses/ uophelp

    OpenAIRE

    Uophelp

    2015-01-01

    For more course tutorials visit www.uophelp.com   MGT 245 Assignment: Organizational Culture MGT 245 Capstone Discussion Question MGT 245 Assignment: Effective Teams MGT 245 CheckPoint: Employee Profile MGT 245 CheckPoint 1: Organizational Structure MGT 245 CheckPoint: Conflict Analysis MGT 245 CheckPoint 2: Peer Review MGT 245 CheckPoint: Employee Programs MGT 245 CheckPoint: Performance Evaluation MGT 245 CheckPoint: Change Management ...

  5. MGT 245 UOP Course Tutorial / Tutorialoutlet

    OpenAIRE

    karthi

    2015-01-01

    For more course tutorials visit www.tutorialoutlet.com MGT 245 Assignment: Organizational Culture MGT 245 Capstone Discussion Question MGT 245 Assignment: Effective Teams MGT 245 CheckPoint: Employee Profile MGT 245 CheckPoint 1: Organizational Structure MGT 245 CheckPoint: Conflict Analysis MGT 245 CheckPoint 2: Peer Review MGT 245 CheckPoint: Employee Programs MGT 245 CheckPoint: Performance Evaluation MGT 245 CheckPoint: Change Management MGT...

  6. MGT 245 Courses/Indigohelp

    OpenAIRE

    charles

    2015-01-01

    For more classes visits www.indigohelp.com   MGT 245 Assignment: Organizational Culture MGT 245 Capstone Discussion Question MGT 245 Assignment: Effective Teams MGT 245 CheckPoint: Employee Profile MGT 245 CheckPoint 1: Organizational Structure MGT 245 CheckPoint: Conflict Analysis MGT 245 CheckPoint 2: Peer Review MGT 245 CheckPoint: Employee Programs MGT 245 CheckPoint: Performance Evaluation MGT 245 CheckPoint: Change Management ...

  7. GenBank blastx search result: AK104850 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK104850 001-043-A07 AL161454.10 Human DNA sequence from clone RP11-72B4 on chromosome 9 Contain ... ir and recombination helicase RAD26L (LOC56959), a parkinson ... disease 7 (PARK7) pseudogene, the 5' end of a nove ...

  8. GenBank blastx search result: AK064456 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK064456 002-110-F11 AL161454.10 Human DNA sequence from clone RP11-72B4 on chromosome 9 Contain ... ir and recombination helicase RAD26L (LOC56959), a parkinson ... disease 7 (PARK7) pseudogene, the 5' end of a nove ...

  9. GenBank blastn search result: AK242341 [KOME

    Lifescience Database Archive (English)

    Full Text Available chromosome 9 Contains the gene for a novel protein similar to putative repair and recombination helicase RAD26L (LOC56959), a parkin...son disease 7 (PARK7) pseudogene, the 5' end of a novel gene and a CpG island, complete sequence. PRI 2e-15 1 -1 ...

  10. GenBank blastx search result: AK242828 [KOME

    Lifescience Database Archive (English)

    Full Text Available chromosome 9 Contains the gene for a novel protein similar to putative repair and recombination helicase RAD26L (LOC56959), a parkin...son disease 7 (PARK7) pseudogene, the 5' end of a novel gene and a CpG island, complete sequence. PRI 1e-58 1 ...

  11. MGT 245 Course Tutorial/snaptutorial

    OpenAIRE

    roman8034

    2015-01-01

    MGT 245 Entire Course For more classes visit www.snaptutorial.com   MGT 245 Assignment: Organizational Culture MGT 245 Capstone Discussion Question MGT 245 Assignment: Effective Teams MGT 245 CheckPoint: Employee Profile MGT 245 CheckPoint 1: Organizational Structure MGT 245 CheckPoint: Conflict Analysis MGT 245 CheckPoint 2: Peer Review MGT 245 CheckPoint: Employee Programs MGT 245 CheckPoint: Performance Evaluation MGT 245 CheckPo...

  12. CIS 105 UOP Course Tutorial/ Tutorialrank

    OpenAIRE

    SINCENK

    2015-01-01

    For more course tutorials visit www.tutorialrank.com Tutorial Purchased: 5 Times, Rating: A   CIS 105 Capstone Discussion Question CIS 105 Assignment: Technology of the Future CIS 105 Assignment: Security Systems CIS 105 CheckPoint: Internet Scavenger Hunt CIS 105 CheckPoint: Computer Comparison CIS 105 CheckPoint: Data Versus Information CIS 105 CheckPoint: Microsoft® Word CIS 105 CheckPoint: Types of Networks CIS 105 CheckPoint: Cyber...

  13. Plumbagin shows anticancer activity in human osteosarcoma (MG-63) cells via the inhibition of S-Phase checkpoints and down-regulation of c-myc

    OpenAIRE

    Yan, Chao-Hua; Li, Feng; Ma, Yuan-Chen

    2015-01-01

    Objective: Plumbagin, a naphthoquinone constituent of Plumbago zeylanica L. (Plumbaginaceae), has been extensively studied for its pharmacological activities and reported to show a good anti-cancer activity in different human cancer cell lines. It is known to exhibit proapoptotic, antiangiogenic and antimetastatic effects in cancer cells. Plumbagin is also known to inhibit NF-κB, JNK (Hsu), PKCε, and STAT-3. However, the anti-proliferatory activity and their core molecular mechanisms have bee...

  14. Targeting the DNA replication checkpoint by pharmacologic inhibition of Chk1 kinase: a strategy to sensitize APC mutant colon cancer cells to 5-fluorouracil chemotherapy

    OpenAIRE

    Martino-Echarri, Estefania; Henderson, Beric R.; Brocardo, Mariana G.

    2014-01-01

    5-fluorouracil (5-FU) is the first line component used in colorectal cancer (CRC) therapy however even in combination with other chemotherapeutic drugs recurrence is common. Mutations of the adenomatous polyposis coli (APC) gene are considered as the initiating step of transformation in familial and sporadic CRCs. We have previously shown that APC regulates the cellular response to DNA replication stress and recently hypothesized that APC mutations might therefore influence 5-FU resistance. T...

  15. Checkpoints Studies Using the Budding Yeast Saccharomyces cerevisiae: Analysis of changes in protein level and subcellular localization during cell cycle progression

    OpenAIRE

    Wu, Xiaorong; Liu, Lili; HUANG, Mingxia

    2011-01-01

    Methods are described here to monitor changes in protein level and subcellular localization during the cell cycle progression in the budding yeast S. cerevisiae. Cell synchronization is achieved by an α-factor mediated block-and-release protocol. Cells are collected at different time points for the first two cell cycles upon release. Cellular DNA contents are analyzed by flow cytometry. Trichloroacetic acid protein precipitates are prepared for monitoring levels of cell cycle regulated protei...

  16. Immune checkpoint regulator PD-L1 expression on tumor cells by contacting CD11b positive bone marrow derived stromal cells

    OpenAIRE

    Noh, Hyangsoon; Hu, Jiemiao; Wang, Xiaohong; Xia, Xueqing; Satelli, Arun; Li, Shulin

    2015-01-01

    Background Expression of programmed cell death ligand 1 (PD-L1) is an important process by which tumor cells suppress antitumor immunity in the tumor microenvironment. Bone marrow (BM)–derived immune cells are an important component of the tumor microenvironment. However, the link between PD-L1 induction on tumor cells and communication with BM cells is unknown. Results This study demonstrates that BM cells have a direct effect in inducing PD-L1 expression on tumor cells, which contributes to...

  17. TBL1 and TBLR1 Phosphorylation on Regulated Gene Promoters Overcomes Dual CtBP and NCoR/SMRT Transcriptional Repression Checkpoints

    OpenAIRE

    Perissi, Valentina; Scafoglio, Claudio; Zhang, Jie; Ohgi, Kenneth A.; Rose, David W.; Glass, Christopher K.; Rosenfeld, Michael G.

    2008-01-01

    A key strategy to achieve regulated gene expression in higher eukaryotes is to prevent illegitimate signal-independent activation by imposing robust control on the dismissal of corepressors. Here, we report that many signaling pathways, including Notch, NFkB, and nuclear receptor ligands, are subjected to a dual repression “check point” based on distinct corepressor complexes. Gene activation requires the release of both CtBP1/2- and NCoR/SMRT-dependent repression, through the coordinate acti...

  18. Analysis of Genomic Integrity and p53-Dependent G1 Checkpoint in Telomerase-Induced Extended-Life-Span Human Fibroblasts

    OpenAIRE

    Vaziri, Homayoun; Jeremy A Squire; Pandita, Tej K.; Bradley, Grace; Kuba, Robert M.; Zhang, Haihua; Gulyas, Sandor; Hill, Richard P.; Nolan, Garry P.; Benchimol, Samuel

    1999-01-01

    Life span determination in normal human cells may be regulated by nucleoprotein structures called telomeres, the physical ends of eukaryotic chromosomes. Telomeres have been shown to be essential for chromosome stability and function and to shorten with each cell division in normal human cells in culture and with age in vivo. Reversal of telomere shortening by the forced expression of telomerase in normal cells has been shown to elongate telomeres and extend the replicative life span (H. Vazi...

  19. CHK1 ensures the preservation of DNA integrity during meiosis I but it is not a component of spindle assembly checkpoint

    Czech Academy of Sciences Publication Activity Database

    Mayer, Alexandra; Baran, V.; Kitajima, T.; Brzáková, Adéla; Ellenberg, J.; Motlík, Jan; Šolc, Petr

    Heidelberg : EMBL, 2012. 107-107. [EMBO/EMBL Symposium Germline Immortality through Totipotency. 13.10.2012-16.10.2012, Heidelberg] Institutional support: RVO:67985904 Keywords : CHK1 Subject RIV: EB - Genetics ; Molecular Biology

  20. Anti-tumor effects in head and neck cancer in response to toll-like receptor activation, checkpoint inhibition, and chemotherapy

    OpenAIRE

    Zhang, Shannon Shueyin

    2016-01-01

    Head and neck cancer (HNC) affects approximately 600,000 individuals annually and occurs when squamous cells lining the oral cavity, nasal cavity, and throat become cancerous. Certain problems are associated with current therapies. Surgery can lead to a lower quality of life due to functional and cosmetic disturbances while chemotherapy and radiation have high toxicity levels. In addition, chemotherapy has low response rates and high recurrence rates. Thus, it is necessary to utilize immune-d...

  1. 14-3-3 checkpoint regulatory proteins interact specifically with DNA repair protein human exonuclease 1 (hEXO1) via a semi-conserved motif

    DEFF Research Database (Denmark)

    Andersen, Sofie Dabros; Keijzers, Guido; Rampakakis, Emmanouil;

    2012-01-01

    Human exonuclease 1 (hEXO1) acts directly in diverse DNA processing events, including replication, mismatch repair (MMR), and double strand break repair (DSBR), and it was also recently described to function as damage sensor and apoptosis inducer following DNA damage. In contrast, 14-3-3 proteins...

  2. Suppression of the p53- or pRB-mediated G1 checkpoint is required for E2F-induced S-phase entry

    DEFF Research Database (Denmark)

    Lomazzi, Marina; Moroni, M Cristina; Jensen, Michael R;

    2002-01-01

    Deregulation of the retinoblastoma protein (pRB) pathway is a hallmark of cancer. In the absence of other genetic alterations, this deregulation results in lack of differentiation, hyperproliferation and apoptosis. The pRB protein acts as a transcriptional repressor by targeting the E2F transcrip......Deregulation of the retinoblastoma protein (pRB) pathway is a hallmark of cancer. In the absence of other genetic alterations, this deregulation results in lack of differentiation, hyperproliferation and apoptosis. The pRB protein acts as a transcriptional repressor by targeting the E2F...

  3. A Topoisomerase II-Dependent Checkpoint in G2-Phase Plat Cells Can Be Bypassed by Ectopic Expression of Mitotic Cyclin B2

    Czech Academy of Sciences Publication Activity Database

    Giménez-Abián, J. F.; Weingartner, M.; Binarová, Pavla; Clarke, D. J.; Anthony, R. G.; Calderini, O.; Heberle-Bors, E.; Díaz de la Espina, S. M.; Bogre, L.; De la Torre, C.

    2002-01-01

    Roč. 1, č. 3 (2002), s. 187-192. ISSN 1538-4101 Grant ostatní: GA BBSRC(ES) 111/P13340 Institutional research plan: CEZ:AV0Z5020903 Keywords : plant cyclin b2 * topoisomeraseii * icrf-193 Subject RIV: EE - Microbiology, Virology

  4. Activation of the nimA protein kinase plays a unique role during mitosis that cannot be bypassed by absence of the bimE checkpoint.

    OpenAIRE

    Osmani, A H; O'Donnell, K; Pu, R T; Osmani, S A

    1991-01-01

    Mutation of nimA reversibly arrests cells in late G2 and nimA overexpression promotes premature mitosis. Here we demonstrate that the product of nimA (designated NIMA) has protein kinase activity that can phosphorylate beta-casein but not histone proteins. NIMA kinase activity is cell cycle regulated being 20-fold higher at mitosis when compared to S-phase arrested cells. NIMA activation is normally required in G2 to initiate chromosome condensation, to nucleate spindle pole body microtubules...

  5. HLA-Restricted CTL That Are Specific for the Immune Checkpoint Ligand PD-L1 Occur with High Frequency in Cancer Patients

    DEFF Research Database (Denmark)

    Munir, Shamaila; Andersen, Gitte Holmen; Met, Özcan;

    2013-01-01

    PD-L1 (CD274) contributes to functional exhaustion of T cells and limits immune responses in patients with cancer. In this study, we report the identification of an human leukocyte antigen (HLA)-A2-restricted epitope from PD-L1, and we describe natural, cytolytic T-cell reactivity against PD-L1 in......, by incubating nonprofessional antigen-presenting cells with long peptides from PD-L1, we found that PD-L1 was rapidly internalized, processed, and cross-presented by HLA-A2 on the cell surface. Apparently, this cross-presentation was TAP-independent, as it was conducted not only by B cells but in...

  6. Human Rif1, ortholog of a yeast telomeric protein, is regulated by ATM and 53BP1 and functions in the S-phase checkpoint

    OpenAIRE

    Silverman, Joshua; Takai, Hiroyuki; Buonomo, Sara B C; Eisenhaber, Frank; de Lange, Titia

    2004-01-01

    We report on the function of the human ortholog of Saccharomyces cerevisiae Rif1 (Rap1-interacting factor 1). Yeast Rif1 associates with telomeres and regulates their length. In contrast, human Rif1 did not accumulate at functional telomeres, but localized to dysfunctional telomeres and to telomeric DNA clusters in ALT cells, a pattern of telomere association typical of DNA-damage-response factors. After induction of double-strand breaks (DSBs), Rif1 formed foci that colocalized with other DN...

  7. HTT 250 uop course tutorial/uop helP

    OpenAIRE

    baily

    2015-01-01

    HTT 250 Week 1 Assignment: Organization Chart Hotel Front Office HTT 250 Week 1 CheckPoint: Total Quality Management HTT 250 Week 1 CheckPoint: Lodging Properties HTT 250 Week 2 CheckPoint: Hotel Price Comparison HTT 250 Week 2 DQs HTT 250 Week 3 CheckPoint: Average Daily Rate HTT 250 Week 4 CheckPoint: Front Office FAQs HTT 250 Week 4 DQs HTT 250 Week 5 Assignment: Night Audit Application HTT 250 Week 5 CheckPoint: Night Audit HTT 250 Week 6 CheckPoint...

  8. Cell cycle checkpoints in Caenorhabditis elegans: the 14-3-3 gene par-5 is required for germline development and DNA damage response / Checkpoints del ciclo celular en Caenorhabditis elegans: el gen 14-3-3, par-5, es necesario para el desarrollo y respuesta al daño genómico de la línea germinal

    OpenAIRE

    Aristizábal Corrales, David

    2012-01-01

    [spa] Las proteínas 14-3-3 han sido ampliamente estudiadas desde levadura hasta mamíferos y han sido asociadas con múltiples roles en procesos como ciclo celular, apoptosis y la respuesta al estrés. Así mismo estas proteínas se han visto involucradas en enfermedades neurodegenerativas y cáncer. De hecho, las proteínas 14-3-3 han sido propuestas como posibles agentes terapéuticos en el tratamiento contra el cáncer. En mamíferos existen 7 genes que codifican para proteínas 14-3-3, mientras en C...

  9. ADJ 225 Courses Tutorial / indigohelp

    OpenAIRE

    nesa

    2015-01-01

    ADJ 225 Assignment Final Project ADJ 225 Week 8 Assignment Prevention Program Resource Guides- Appendix F ADJ 225 CheckPoint 2 Mission Statement ADJ 225 CheckPoint Levels of Prevention ADJ 225 Assignment Juvenile Court Comparison ADJ 225 Assignment Corrections Comparison Summary ADJ 225 CheckPoint Case Study Sequence ADJ 225 Assignment Risk Factor Handout ADJ 225 CheckPoint Community Agency Goals ADJ 225 CheckPoint 1 Juvenile Court Characteristics ADJ 2...

  10. CIS 105 UOP COURSES TUTORIAL/UOPHELP

    OpenAIRE

    hubli

    2015-01-01

    For more course tutorials visit www.uophelp.com   CIS 105 Capstone Discussion Question   CIS 105 Assignment: Technology of the Future   CIS 105 Assignment: Security Systems   CIS 105 CheckPoint: Internet Scavenger Hunt   CIS 105 CheckPoint: Computer Comparison   CIS 105 CheckPoint: Data Versus Information   CIS 105 CheckPoint: Microsoft® Word   CIS 105 CheckPoint: Types of Networks ...

  11. HIS 115 Course Tutorial/snaptutorial

    OpenAIRE

    roman8034

    2015-01-01

    HIS 115 Entire Course For more classes visit www.snaptutorial.com   HIS 115 Capstone CheckPoint HIS 115 CheckPoint: The Confederation Government Table HIS 115 CheckPoint: Compare and Contrast Matrix HIS 115 CheckPoint: Civil War Matrix HIS 115 Assignment: North American Civilization Paper HIS 115 Final Project: Historical Timeline and Essay HIS 115 Assignment: Seven Years’ War Paper HIS 115 CheckPoint: European Societal Changes HIS 11...

  12. HIS 115 UOP course tutorial/tutorialoutlet

    OpenAIRE

    NARESH 1

    2015-01-01

    For more course tutorials visit www.tutorialoutlet.com     HIS 115 Capstone CheckPoint HIS 115 CheckPoint: The Confederation Government Table HIS 115 CheckPoint: Compare and Contrast Matrix HIS 115 CheckPoint: Civil War Matrix HIS 115 Assignment: North American Civilization Paper HIS 115 Final Project: Historical Timeline and Essay HIS 115 Assignment: Seven Years’ War Paper HIS 115 CheckPoint: European Societal Changes HIS 115 WE...

  13. HIS 115 uop course/uophelp

    OpenAIRE

    uop help

    2015-01-01

    For more course tutorials visit www.uophelp.com         HIS 115 Capstone CheckPoint HIS 115 CheckPoint: The Confederation Government Table HIS 115 CheckPoint: Compare and Contrast Matrix HIS 115 CheckPoint: Civil War Matrix HIS 115 Assignment: North American Civilization Paper HIS 115 Final Project: Historical Timeline and Essay HIS 115 Assignment: Seven Years’ War Paper HIS 115 CheckPoint: European Societal Changes...

  14. ADJ 225 UOP Course Tutorial/TutotorialRank

    OpenAIRE

    hidden

    2015-01-01

    For more course tutorials visit www.tutorialrank.com Tutorial Purchased: 3 Times, Rating: A+ ADJ 225 Assignment Corrections Comparison Summary  ADJ 225 Assignment Final Project ADJ 225 Assignment Juvenile Court Comparison ADJ 225 Assignment Risk Factor Handout ADJ 225 CheckPoint 1 Juvenile Court Characteristics ADJ 225 CheckPoint 2 Mission Statement ADJ 225 CheckPoint Case Study Sequence ADJ 225 CheckPoint Community Agency Goals ADJ 225 CheckPoint Juveni...

  15. PSY 285 Uop Material - psy285dotcom

    OpenAIRE

    honey

    2015-01-01

    PSY 285 Entire Course For more course tutorials visit www.psy285.com   PSY 285 Week 1 Assignment: Experimental Research PSY 285 Week 1 CheckPoint: Causation and Correlation PSY 285 Week 2 CheckPoint: Locus of Control PSY 285 Week 3 CheckPoint: Confirmation Bias PSY 285 Week 3 Assignment: Attitudes and Behaviors PSY 285 Week 4 CheckPoint: Obedience PSY 285 Week 5 Assignment: Persuasion, Indoctrination, and Inoculation PSY 285 Week 5 CheckPoint: ...

  16. ADJ 235 UOP COURSE Tutorial/UOPHELP

    OpenAIRE

    vasu

    2015-01-01

    ADJ 235 Week 1 Checkpoint Legalizing Morality ADJ 235 Week 1 DQ 1 and DQ 2 ADJ 235 Week 2 Checkpoint Interpretation of Ethical Systems ADJ 235 Week 2 Assignment Deontological Versus Teleological Ethical Systems ADJ 235 Week 3 Checkpoint Undercover Operations ADJ 235 Week 3 DQ 1 and DQ 2 ADJ 235 Week 4 Checkpoint Law Enforcement Code of Ethics ADJ 235 Week 4 Assignment Law Enforcement Deviance ADJ 235 Week 5 Checkpoint Attorneys Ethical Responsibilities ADJ ...

  17. ADJ 235 Courses Tutorial / indigohelp

    OpenAIRE

    nesa

    2015-01-01

    ADJ 235 Week 1 Checkpoint Legalizing Morality ADJ 235 Week 1 DQ 1 and DQ 2 ADJ 235 Week 2 Checkpoint Interpretation of Ethical Systems ADJ 235 Week 2 Assignment Deontological Versus Teleological Ethical Systems ADJ 235 Week 3 Checkpoint Undercover Operations ADJ 235 Week 3 DQ 1 and DQ 2 ADJ 235 Week 4 Checkpoint Law Enforcement Code of Ethics ADJ 235 Week 4 Assignment Law Enforcement Deviance ADJ 235 Week 5 Checkpoint Attorneys Ethical Responsibilities ADJ ...

  18. SCI 230 ENTIRE COURSE *INTRODUCTION TO LIFE SCIENCE* UPDATE COURSE

    OpenAIRE

    SCI 230 ENTIRE COURSE

    2016-01-01

    http://www.helperstudy.com/products/sci-230 SCI 230 ALL DQS SCI 230 WEEK 1 ASSIGNMENT THE SCIENTIFIC METHOD SCI 230 WEEK 1 CHECKPOINT LEVELS OF LIFE SCI 230 WEEK 1 CHECKPOINT THEORIES OF BIOLOGY SCI 230 WEEK 2 CHECKPOINT PLANTS VS. ANIMALS SCI 230 WEEK 3 ASSIGNMENT PHOTOSYNTHESIS AND CELLULAR RESPIRATION SCI 230 WEEK 3 CHECKPOINT MITOSIS AND MEIOSIS SCI 230 WEEK 3 EXERCISE UNESCO RESEARCH SCI 230 WEEK 4 CHECKPOINT MENDEL ON PATTERNS OF INHERITANCE SCI 2...

  19. HTT200 phoenix Material-htt200dotcom

    OpenAIRE

    sandy

    2015-01-01

    HTT 200 Entire Course (UOP) For more course tutorials visit www.htt200.com   HTT 200 Final Project: Foreign Destination Research Project HTT 200 Week 1 CheckPoint: Job Opportunities HTT 200 Week 2 Assignment: Demographics HTT 200 Week 2 CheckPoint: Diversity HTT 200 Week 3 CheckPoint: Casual Restaurants HTT 200 Week 4 Assignment: Franchising HTT 200 Week 4 CheckPoint: Chain and Independent Restaurants HTT 200 Week 5 CheckPoint: Customer Types ...

  20. HTT 200 UOP Course Tutorial / Tutoriloutlet

    OpenAIRE

    JANU

    2015-01-01

    For more course tutorials visit www.tutorialoutlet.com     HTT 200 Final Project: Foreign Destination Research Project HTT 200 Week 1 CheckPoint: Job Opportunities HTT 200 Week 2 Assignment: Demographics HTT 200 Week 2 CheckPoint: Diversity HTT 200 Week 3 CheckPoint: Casual Restaurants HTT 200 Week 4 Assignment: Franchising HTT 200 Week 4 CheckPoint: Chain and Independent Restaurants HTT 200 Week 5 CheckPoint: Customer Types HTT 200 We...

  1. HTT 200 UOP Coureses/UOP Help

    OpenAIRE

    royal12

    2015-01-01

     HTT 200 Entire Course For more course tutorials visit www.uophelp.com HTT 200 Final Project: Foreign Destination Research Project HTT 200 Week 1 CheckPoint: Job Opportunities HTT 200 Week 2 Assignment: Demographics HTT 200 Week 2 CheckPoint: Diversity HTT 200 Week 3 CheckPoint: Casual Restaurants HTT 200 Week 4 Assignment: Franchising HTT 200 Week 4 CheckPoint: Chain and Independent Restaurants HTT 200 Week 5 CheckPoint: Customer Types HTT 200...

  2. HSM 270 UOP course tutorial/uop helP

    OpenAIRE

    HUTN

    2015-01-01

    For more course tutorials visit www.uophelp.com HTT 200 Final Project: Foreign Destination Research Project HTT 200 Week 1 CheckPoint: Job Opportunities HTT 200 Week 2 Assignment: Demographics HTT 200 Week 2 CheckPoint: Diversity HTT 200 Week 3 CheckPoint: Casual Restaurants HTT 200 Week 4 Assignment: Franchising HTT 200 Week 4 CheckPoint: Chain and Independent Restaurants HTT 200 Week 5 CheckPoint: Customer Types HTT 200 Week 6 Assignment: Hotel Securi...

  3. AcEST: DK954482 [AcEST

    Lifescience Database Archive (English)

    Full Text Available sp|Q9Y620|RA54B_HUMAN DNA repair and recombination protein RAD54... 81 3e-15 sp|Q6PDQ2|CHD4_MOUSE Chromodomain-helicase-DNA-bindin..... 81 3e-15 sp|Q6PFE3|RA54B_MOUSE DNA repair and recombination protein RAD54... 81 4e-15 sp|Q22516|CHD3_CAEEL Chromodomain...QPTALQLELYRKLLSS 572 >sp|P40352|RAD26_YEAST DNA repair and recombination protein RAD26 OS=Saccharomyces cere...32863|RAD54_YEAST DNA repair and recombination protein RAD54 OS=Saccharomyces cerevisiae GN=RAD54 PE=1 SV=1 ...NJT5_COPC7 Putative uncharacterized protein OS=Copri... 129 9e-29 tr|B2WKB2|B2WKB2_PYRTR DNA repair and recombination protein

  4. Low Overhead Minimum Process Global Snapshot Collection Algorithm for Mobile Distributed System

    Directory of Open Access Journals (Sweden)

    Surender Kumar

    2010-05-01

    Full Text Available Coordinated checkpointing is an effective fault tolerant technique in distributed system as it avoids the domino effect and require minimum storage requirement. Most of the earlier coordinated checkpoint algorithms block their computation during checkpointing and forces minimum-process or non-blocking but forces all nodes to takes checkpoint even though many of them may not be necessary or non-blocking minimum-process but takes useless checkpoints or reduced useless checkpoint but has higher synchronization message overhead or has high checkpoint request propagation time. Hence in mobile distributed systems there is a great need of minimizing the number of communication message and checkpointing overhead as it raise new issues such as mobility, low bandwidth of wireless channels, frequently disconnections, limited battery power and lack of reliable stable storage on mobile nodes. In this paper, we propose a minimum-process coordinated checkpointing algorithm for mobile distributed system where no useless checkpoints are taken, no blocking of processes takes place and enforces a minimum-number of processes to take checkpoints. Our algorithm imposes low memory and computation overheads on MH’s and low communication overheads on wireless channels. It avoids awakening of an MH if it is not required to take its checkpoint and has reduced latency time as each process involved in a global checkpoint can forward its own decision directly to the checkpoint initiator.

  5. AED 200 UOP course/uophelp

    OpenAIRE

    uophelp

    2015-01-01

    For more course tutorials visit www.uophelp.com   AED 200 Week 1 DQ1 and DQ2 AED 200 Week 1 CheckPoint Teacher Certification Licensure AED 200 Week 2 CheckPoint Students Social Challenges AED 200 Week 2 CheckPoint Social Justice AED 200 Week 2 Assignment Multicultural Education Scenario AED 200 Week 3 DQ 1 and DQ 2 AED 200 Week 3 CheckPoint School Funding Issues AED 200 Week 4 CheckPoint Religion in Schools AED 200 Week 4 CheckPoint Debate Proj...

  6. AED 200 Courses Tutorial / indigohelp

    OpenAIRE

    nesa

    2015-01-01

    AED 200 Week 1 DQ1 and DQ2 AED 200 Week 1 CheckPoint Teacher Certification Licensure AED 200 Week 2 CheckPoint Students Social Challenges AED 200 Week 2 CheckPoint Social Justice AED 200 Week 2 Assignment Multicultural Education Scenario AED 200 Week 3 DQ 1 and DQ 2 AED 200 Week 3 CheckPoint School Funding Issues AED 200 Week 4 CheckPoint Religion in Schools AED 200 Week 4 CheckPoint Debate Project Outline AED 200 Week 4 Assignment Students Rights and Teach...

  7. SPM 220 uop course tutorial/tutorial rank

    OpenAIRE

    tiplod

    2015-01-01

    SPM 220 Week 1 CheckPoint: Moral and Nonmoral Values SPM 220 Week 2 Assignment: Principles and Exceptions SPM 220 Week 2 CheckPoint: Moral Principles SPM 220 Week 3 CheckPoint: Intimidation in Sport SPM 220 Week 4 Assignment: Kids Speak Out SPM 220 Week 4 CheckPoint: Violence in Sport SPM 220 Week 5 CheckPoint: Eligibility and Elimination in Sport SPM 220 Week 6 Assignment: Exploitation in Sport SPM 220 Week 6 CheckPoint: Commercialized Sport SPM 220 Week 7...

  8. PSY 285 UOP Courses / uoptutorial

    OpenAIRE

    THANU

    2015-01-01

    PSY 285 Week 1 Assignment: Experimental Research PSY 285 Week 1 CheckPoint: Causation and Correlation PSY 285 Week 2 CheckPoint: Locus of Control PSY 285 Week 3 CheckPoint: Confirmation Bias PSY 285 Week 3 Assignment: Attitudes and Behaviors PSY 285 Week 4 CheckPoint: Obedience PSY 285 Week 5 Assignment: Persuasion, Indoctrination, and Inoculation PSY 285 Week 5 CheckPoint: Presence of Others PSY 285 Week 6 CheckPoint: Symptoms and Remedies of Groupthink PS...

  9. AED 200 uop course tutorial/uop help

    OpenAIRE

    baily

    2015-01-01

     For more course tutorials visit www.uophelp.com   AED 200 Week 1 DQ1 and DQ2 AED 200 Week 1 CheckPoint Teacher Certification Licensure AED 200 Week 2 CheckPoint Students Social Challenges AED 200 Week 2 CheckPoint Social Justice AED 200 Week 2 Assignment Multicultural Education Scenario AED 200 Week 3 DQ 1 and DQ 2 AED 200 Week 3 CheckPoint School Funding Issues AED 200 Week 4 CheckPoint Religion in Schools AED 200 Week 4 CheckPoint Debat...

  10. ADJ 275 Courses Tutorial / indigohelp

    OpenAIRE

    NESA

    2015-01-01

    ADJ 275 Week 2 Checkpoint 1 Criminal Procedures ADJ 275 Week 2 CheckPoint 2 Criminal Procedure Variation ADJ 275 Week 2 Assignment Jurisdiction Case Review ADJ 275 Week 3 Checkpoint Arrest ADJ 275 Week 4 CheckPoint 1 Exclusionary Rule Arguments ADJ 275 Week 4 CheckPoint 2 Stop and Frisk and Arrest ADJ 275 Week 4 Assignment Exclusionary Rule Presentation ADJ 275 Week 5 CheckPoint Search and Seizure ADJ 275 Week 6 Assignment Plain View Open Fields Case Study ...

  11. PSY 240 Course Tutorial / Snaptutorial

    OpenAIRE

    Aster

    2015-01-01

    PSY 240 Week 1 CheckPoint The Nature-Nurture Issue PSY 240 Week 1 DQ 1 and DQ 2 PSY 240 Week 2 CheckPoint The Nervous System PSY 240 Week 2 Assignment The Brain PSY 240 Week 3 CheckPoint Brain Studies PSY 240 Week 3 DQ 1 and DQ 2 PSY 240 Week 4 CheckPoint Eating What, When, and How Much PSY 240 Week 4 CheckPoint Set and Settling Points PSY 240 Week 4 Assignment To Eat or Not to Eat PSY 240 Week 5 CheckPoint Sexual Orientation and Identity PSY 240 Week 5...

  12. PSY 285 Course Tutorial / Snaptutorial

    OpenAIRE

    Aster

    2015-01-01

    PSY 285 Week 1 Assignment: Experimental Research PSY 285 Week 1 CheckPoint: Causation and Correlation PSY 285 Week 2 CheckPoint: Locus of Control PSY 285 Week 3 CheckPoint: Confirmation Bias PSY 285 Week 3 Assignment: Attitudes and Behaviors PSY 285 Week 4 CheckPoint: Obedience PSY 285 Week 5 Assignment: Persuasion, Indoctrination, and Inoculation PSY 285 Week 5 CheckPoint: Presence of Others PSY 285 Week 6 CheckPoint: Symptoms and Remedies of Groupthink PS...

  13. Yeast Interacting Proteins Database: YER118C, YER118C [Yeast Interacting Proteins Database

    Lifescience Database Archive (English)

    Full Text Available YER118C SHO1 Transmembrane osmosensor, participates in activation of both the Cdc42p- and MAP ... ki ... participates in activation of both the Cdc42p- and MAP ... kinase-dependent filamentous growth pathway and th ... participates in activation of both the Cdc42p- and MAP ... kinase-dependent filamentous growth pathway and th ...

  14. Arabidopsis CDS blastp result: AK065579 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK065579 J013031C20 At3g05480.1 cell cycle checkpoint control protein family contains Pfam profi ... ein N-myristoyltransferase signature 2; similar to radio -resistance/chemo-resistance/cell cycle checkpoint ...

  15. FIN 200 (UOP) course tutorial/tutorialoutlet

    OpenAIRE

    naresh 1

    2015-01-01

    For more course tutorials visit www.tutorialoutlet.com     FIN 200 Week 1 CheckPoint Financial Management Goals (UOP) FIN 200 Week 1 Assignment Cash Flow Preparation (UOP) FIN 200 Week 2 Checkpoint Financial Ratios (UOP) FIN 200 Week 2 DQ 1 & DQ 2 (UOP) FIN 200 Week 3 CheckPoint Financial Forecasting (UOP) FIN 200 Week 3 Assignment Pro Forma Statements (UOP) FIN 200 Week 4 Checkpoint Break Even Analysis (UOP) FIN 200 Week 4 DQ 1 & ...

  16. GEN 105 Course tutorial/ indigohelp

    OpenAIRE

    SCD

    2015-01-01

    For more classes visit www.indigohelp.com   GEN 105 Assignment: Reading and Retention GEN 105 Assignment: Elevator Speech GEN 105 CheckPoint: Technological Tools GEN 105 CheckPoint: Distance Learning I GEN 105 CheckPoint: Distance Learning II GEN 105 CheckPoint: Communicating in Forums GEN 105 Week 2 Discussion Questions GEN 105 Week 4 Discussion Questions GEN 105 Week 6 Discussion Questions GEN 105 Week 8 Discussion Questions GEN 105 A...

  17. ADJ 225 uop course tutorial/uop help

    OpenAIRE

    sando

    2015-01-01

    For more course tutorials visit www.uophelp.com       ADJ 225 Assignment Final Project ADJ 225 Week 8 Assignment Prevention Program Resource Guides- Appendix F ADJ 225 CheckPoint 2 Mission Statement ADJ 225 CheckPoint Levels of Prevention ADJ 225 Assignment Juvenile Court Comparison ADJ 225 Assignment Corrections Comparison Summary ADJ 225 CheckPoint Case Study Sequence ADJ 225 Assignment Risk Factor Handout ADJ 225 CheckPoint...

  18. PHI 105 Course tutorial/ indigohelp

    OpenAIRE

    SAS

    2015-01-01

    For more classes visit www.indigohelp.com   PHI 105 Final Project: Personal Philosophy PowerPoint® Presentation PHI 105 CheckPoint: Pragmatism and Analytic Philosophy PHI 105 Assignment: Ethical Decisions Scenario Analysis PHI 105 CheckPoint: Final Project Preview PHI 105 CheckPoint: Argument and Logic PHI 105 CheckPoint: Final Project Topic PHI 105 WEEK 2 DISCUSSION QUESTION 1 PHI 105 WEEK 2 DISCUSSION QUESTION 2 PHI 105 Assignment: Letter...

  19. POS 110 Course Tutorial / Snaptutorial

    OpenAIRE

    Selye

    2015-01-01

    For more classes visit www.snaptutorial.com POS 110 Exercise: Amending the Constitution POS 110 Assignment: Bureaucracy and You POS 110 Capstone CheckPoint POS 110 CheckPoint: Federal Court Structure POS 110 Assignment: Get to Know Your Member of Congress POS 110 Assignment: Civil Liberties and Civil Rights POS 110 Final Project: Democracy in America POS 110 CheckPoint: Defining Democracy POS 110 CheckPoint: American Civic Values POS 110 Week 2 Disc...

  20. HTT 230 UOP Coureses/UOP Help

    OpenAIRE

    royal8

    2015-01-01

    HTT 230 Entire Course For more course tutorials visit www.uophelp.com       HTT 230 Week 1 CheckPoint: Primary Financial Statements HTT 230 Week 2 Assignment: Rosabella and Canyon Hotel Concept Checks HTT 230 Week 2 CheckPoint: The CP3 Accounting System HTT 230 Week 3 CheckPoint: Credit Card Research HTT 230 Week 3 CheckPoint: Crescendo Concept Check HTT 230 Week 4 Assignment: Watree Lodge and Greenville Restaurant Concept Checks ...

  1. CJS 200 UOP Course Tutorial/ Tutorialrank

    OpenAIRE

    SICENK

    2015-01-01

    For more course tutorials visit www.tutorialrank.com Tutorial Purchased: 3 Times, Rating: A+   CJS 200 Assignment: The Prison System CJS 200 Assignment: The Effects of Punishment and Sentencing CJS 200 CheckPoint: Prison Populations CJS 200 Assignment: Inside Criminal Law CJS 200 CheckPoint: Causation of Crime CJS 200 CheckPoint: Law Enforcement Agencies CJS 200 Assignment: The Rules of Law Enforcement CJS 200 CheckPoint: Terrorism and Cyber Cr...

  2. XACC 280 courses/snaptutorial

    OpenAIRE

    Ronald James Marsh

    2015-01-01

    For more classes visit www.snaptutorial.com   XACC 280 Week 1 CheckPoint Accounting Assumptions, Principles, and Constraints XACC 280 Week 1 CheckPoint The Accounting Equation XACC 280 Week 1 Discussion Question 1 & 2 XACC 280 Week 2 CheckPoint Debits and Credits XACC 280 Week 2 Assignment Journalizing, Posting, and Preparing a Trial Balance XACC 280 Week 3 CheckPoint Adjusting Entries, Posting, and Preparing an Adjusted Trial Balance XACC 280 Week...

  3. POS 110 UOP Course Tutorial/ Tutorialrank

    OpenAIRE

    john

    2015-01-01

    For more course tutorials visit www.tutorialrank.com   POS 110 Exercise: Amending the Constitution POS 110 Assignment: Bureaucracy and You POS 110 Capstone CheckPoint POS 110 CheckPoint: Federal Court Structure POS 110 Assignment: Get to Know Your Member of Congress POS 110 Assignment: Civil Liberties and Civil Rights POS 110 Final Project: Democracy in America POS 110 CheckPoint: Defining Democracy POS 110 CheckPoint: American Civic Values ...

  4. XACC 280 Course tutorial/uophelp

    OpenAIRE

    SZXDSCX

    2015-01-01

    XACC 280 Entire Course For more course tutorials visit www.uophelp.com     XACC 280 Week 1 CheckPoint Accounting Assumptions, Principles, and Constraints XACC 280 Week 1 CheckPoint The Accounting Equation XACC 280 Week 1 Discussion Question 1 & 2 XACC 280 Week 2 CheckPoint Debits and Credits XACC 280 Week 2 Assignment Journalizing, Posting, and Preparing a Trial Balance XACC 280 Week 3 CheckPoint Adjusting Entries, Posting, and Preparing a...

  5. GEN 105 UOP course tutorial/tutorialoutlet

    OpenAIRE

    NARESH 1

    2015-01-01

    For more course tutorials visit www.tutorialoutlet.com     GEN 105 Assignment: Reading and Retention GEN 105 Assignment: Elevator Speech GEN 105 CheckPoint: Technological Tools GEN 105 CheckPoint: Distance Learning I GEN 105 CheckPoint: Distance Learning II GEN 105 CheckPoint: Communicating in Forums GEN 105 Week 2 Discussion Questions GEN 105 Week 4 Discussion Questions GEN 105 Week 6 Discussion Questions GEN 105 Week 8 Discussion...

  6. POS 110 COURSE TUTORIAL/ indigohelp

    OpenAIRE

    jayadev

    2015-01-01

    For more classes visit www.indigohelp.com POS 110 Exercise: Amending the Constitution POS 110 Assignment: Bureaucracy and You POS 110 Capstone CheckPoint POS 110 CheckPoint: Federal Court Structure POS 110 Assignment: Get to Know Your Member of Congress POS 110 Assignment: Civil Liberties and Civil Rights POS 110 Final Project: Democracy in America POS 110 CheckPoint: Defining Democracy POS 110 CheckPoint: American Civic Values POS 110 Week 2 Discus...

  7. XCOM 285 Courses Tutorial / indigohelp

    OpenAIRE

    sana

    2015-01-01

    XCOM 285 Week 1 CheckPoint Technology Trendy Communication XCOM 285 Week 1 Assignment Business Communication Trends XCOM 285 Week 2 Discussion Question 1 & 2 XCOM 285 Week 2 CheckPoint Audience Focused Communication Matrix XCOM 285 Week 3 CheckPoint Technological Advancements in Communication XCOM 285 Week 3 Assignment Cross Cultural Communication Matrix XCOM 285 Week 4 Discussion Question 1 & 2XCOM 285 Week 4 CheckPoint Graphic Organizer XCOM 285 Week 5 Ch...

  8. ADJ 225 UOP COURSE Tutorial/UOPHELP

    OpenAIRE

    mangush

    2015-01-01

    For more course tutorials visit www.uophelp.com       ADJ 225 Assignment Final Project ADJ 225 Week 8 Assignment Prevention Program Resource Guides- Appendix F ADJ 225 CheckPoint 2 Mission Statement ADJ 225 CheckPoint Levels of Prevention ADJ 225 Assignment Juvenile Court Comparison ADJ 225 Assignment Corrections Comparison Summary ADJ 225 CheckPoint Case Study Sequence ADJ 225 Assignment Risk Factor Handout ADJ 225 CheckPoint...

  9. GEN 105 Courses/sanptutorial

    OpenAIRE

    ncbdf

    2015-01-01

    GEN 105 Assignment: Reading and Retention GEN 105 Assignment: Elevator Speech GEN 105 CheckPoint: Technological Tools GEN 105 CheckPoint: Distance Learning I GEN 105 CheckPoint: Distance Learning II GEN 105 CheckPoint: Communicating in Forums GEN 105 Week 2 Discussion Questions GEN 105 Week 4 Discussion Questions GEN 105 Week 6 Discussion Questions GEN 105 Week 8 Discussion Questions GEN 105 Assignment: University Library Article Search GEN 105 Chec...

  10. ADJ 215 UOP Course Tutorial/TutorialRank

    OpenAIRE

    apj

    2015-01-01

    For more course tutorials visit www.tutorialrank.com Tutorial Purchased: 1 Times, Rating:     ADJ 215 Week 1 Checkpoint Survey of Local Crime Prevention Methods ADJ 215 Week 1 DQ 1 and DQ 2 ADJ 215 Week 2 Assignment Just Desserts ADJ 215 Week 3 DQ 1 and DQ 2 ADJ 215 Week 4 Checkpoint Social Structure Crime ADJ 215 Week 4 Checkpoint Diversion Program ADJ 215 Week 4 Assignment Socialization and Criminal Behavior ADJ 215 Week 5 Checkpoint...

  11. ADJ 235 uop course tutorial/uop help

    OpenAIRE

    andor

    2015-01-01

     For more course tutorials visit www.uophelp.com     ADJ 235 Week 1 Checkpoint Legalizing Morality ADJ 235 Week 1 DQ 1 and DQ 2 ADJ 235 Week 2 Checkpoint Interpretation of Ethical Systems ADJ 235 Week 2 Assignment Deontological Versus Teleological Ethical Systems ADJ 235 Week 3 Checkpoint Undercover Operations ADJ 235 Week 3 DQ 1 and DQ 2 ADJ 235 Week 4 Checkpoint Law Enforcement Code of Ethics ADJ 235 Week 4 Assignment Law Enforcemen...

  12. ADJ 235 Tutorials / adj235dotcom

    OpenAIRE

    munna9

    2015-01-01

    ADJ 235 Entire Course (UOP)   For more course tutorials visit www.adj235.com   ADJ 235 Week 1 Checkpoint Legalizing Morality ADJ 235 Week 1 DQ 1 and DQ 2 ADJ 235 Week 2 Checkpoint Interpretation of Ethical Systems ADJ 235 Week 2 Assignment Deontological Versus Teleological Ethical Systems ADJ 235 Week 3 Checkpoint Undercover Operations ADJ 235 Week 3 DQ 1 and DQ 2 ADJ 235 Week 4 Checkpoint Law Enforcement Code of Ethics ADJ 235 Week 4 ...

  13. ADJ 235 UOP Course Tutorial / adj235dotcom

    OpenAIRE

    geenid

    2015-01-01

    ADJ 235 Entire Course (UOP) For more course tutorials visit www.adj235.com ADJ 235 Week 1 Checkpoint Legalizing Morality ADJ 235 Week 1 DQ 1 and DQ 2 ADJ 235 Week 2 Checkpoint Interpretation of Ethical Systems ADJ 235 Week 2 Assignment Deontological Versus Teleological Ethical Systems ADJ 235 Week 3 Checkpoint Undercover Operations ADJ 235 Week 3 DQ 1 and DQ 2 ADJ 235 Week 4 Checkpoint Law Enforcement Code of Ethics ADJ 235 Week 4 Assignment Law Enforce...

  14. ADJ 235 Courses/snaptutorial

    OpenAIRE

    charles

    2015-01-01

    ­For more classes visits www.indigohelp.com                 ADJ 235 Week 1 Checkpoint Legalizing Morality ADJ 235 Week 1 DQ 1 and DQ 2 ADJ 235 Week 2 Checkpoint Interpretation of Ethical Systems ADJ 235 Week 2 Assignment Deontological Versus Teleological Ethical Systems ADJ 235 Week 3 Checkpoint Undercover Operations ADJ 235 Week 3 DQ 1 and DQ 2 ADJ 235 Week 4 Checkpoint Law Enforcement...

  15. SCI 241 WEEKS 1-9 ENTIRE COURSE

    OpenAIRE

    SCI 241 WEEKS 1-9 ENTIRE COURSE

    2016-01-01

    http://www.helperstudy.com/products/sci-241 SCI 241 WEEK 9 CAPSTONE CHECKPOINT REFLECTIONS ON NUTRITION SCI 241 WEEK 9 ASSIGNMENT FINAL PROJECT PERSONALIZED NUTRITION AND EXERCISE PLAN SCI 241 WEEK 7 CHECKPOINT LIFESPAN NUTRITIONAL NEEDS POWERPOINT PRESENTATION SCI 241 WEEK 7 ASSIGNMENT HEALTHY EATING PLAN COMPARISON SCI 241 WEEK 6 CHECKPOINT PERSONAL EXERCISE PLAN SCI 241 WEEK 5 CHECKPOINT MULTIVITIAMIN REVIEW SCI 241 WEEK 5 ASSIGNMENT DEHYDRATION SCI 241 WEEK...

  16. XMGT 216 courses/snaptutorial

    OpenAIRE

    Paul Marlowe

    2015-01-01

    For more classes visit www.snaptutorial.com   XMGT 216 Week 1 CheckPoint Ethical Theories Chart XMGT 216 Week 1 Discussion Question 1 & 2 XMGT 216 Week 2 CheckPoint Clarifying Personal Values XMGT 216 Week 2 Assignment Business Ethics Reflection XMGT 216 Week 3 CheckPoint Ethics Awareness Inventory XMGT 216 Week 3 Discussion Question 1 & 2 XMGT 216 Week 4 Assignment Ethical Issues and Management Paper XMGT 216 Week 4 CheckPoint Short Case M...

  17. The design and implementation of Berkeley Lab's linuxcheckpoint/restart

    Energy Technology Data Exchange (ETDEWEB)

    Duell, Jason

    2005-04-30

    This paper describes Berkeley Linux Checkpoint/Restart (BLCR), a linux kernel module that allows system-level checkpoints on a variety of Linux systems. BLCR can be used either as a stand alone system for checkpointing applications on a single machine, or as a component by a scheduling system or parallel communication library for checkpointing and restoring parallel jobs running on multiple machines. Integration with Message Passing Interface (MPI) and other parallel systems is described.

  18. HTT 250 UOP Tutorial/ Uoptutorial

    OpenAIRE

    Moon

    2015-01-01

    For more course tutorials visit www.uoptutorial.com   HTT 250 Week 1 Assignment Organization Chart Hotel Front Office   HTT 250 Week 1 CheckPoint Total Quality Management   HTT 250 Week 1 CheckPoint Lodging Properties   HTT 250 Week 2 CheckPoint Hotel Price Comparison   HTT 250 Week 2 DQs   HTT 250 Week 3 CheckPoint Average Daily Rate   HTT 250 Week 3 Assignment Revenue Management HTT 250 Week ...

  19. HTT 250 UOP Course Tutorial / Tutorialoutlet

    OpenAIRE

    keerthi

    2015-01-01

    For more course tutorials visit www.tutorialoutlet.com     HTT 250 Week 1 Assignment Organization Chart Hotel Front Office   HTT 250 Week 1 CheckPoint Total Quality Management   HTT 250 Week 1 CheckPoint Lodging Properties   HTT 250 Week 2 CheckPoint Hotel Price Comparison   HTT 250 Week 2 DQs   HTT 250 Week 3 CheckPoint Average Daily Rate   HTT 250 Week 3 Assignment Revenue Management ...

  20. Función de Dot1 en el checkpoint de recombinación meiótica y en la tolerancia al daño en el DNA

    OpenAIRE

    Ontoso Picón, David

    2013-01-01

    [ES] Las modificaciones post-traduccionales de las histonas repercuten en numerosas funciones biológicas, regulando procesos esenciales como la transcripción, la replicación, la reparación del DNA, la recombinación o el control del ciclo celular. En la presente tesis doctoral se ha estudiado la influencia de la metilación de la lisina 79 de la histona H3 (H3K79me), mediada en exclusiva por la singular enzima Dot1, en mecanismos celulares implicados en el mantenimiento de la integridad genómic...