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Cells are constantly challenged by genotoxic stresses that can lead to genome instability. The integrity of the nuclear genome is preserved by the DNA damage response (DDR) and repair. Additionally, these stresses can induce mitochondria to transiently hyperfuse; however, it remains unclear whether canonical DDR is linked to these mitochondrial morphological changes. Here, we report that the abolition of mitochondrial fusion causes a substantial defect in the ATM-mediated DDR signaling. This deficiency is overcome by the restoration of mitochondria fusion. In cells with fragmented mitochondria, genotoxic stress-induced activation of JNK and its translocation to DNA lesion are lost. Importantly, the mitochondrial fusion machinery of MFN1/MFN2 associates with Sab (SH3BP5) and JNK, and these interactions are indispensable for the Sab-mediated activation of JNK and the ATM-mediated DDR signaling. Accordingly, the formation of BRCA1 and 53BP1 foci, as well as homology and end-joining repair are impaired in cells with fragmented mitochondria. Together, these data show that mitochondrial fusion-dependent JNK signaling is essential for the DDR, providing vital insight into the integration of nuclear and cytoplasmic stress signals.

The NLRP3 inflammasome is a global immune-sensor that is activated by a repertoire of endogenous and exogenous stimuli. NLRP3 translocates to mitochondria but whether mitochondria involvement in the inflammasome assembly is unclear. Here, we show that the mitochondrial E3 ligase MARCH5 is a key regulator of NLRP3 inflammasome assembly. Myeloid cell-specificMarch5 conditional knockout (March5cKO) mice exhibited an attenuated mortality rate upon LPS orPseudomonas aeruginosachallenge. Macrophages derived fromMarch5cKO mice failed to secrete IL-1β and IL-18 after microbial infection. Mechanistically, MARCH5 interacts with the NACHT domain of NLRP3 and promotes K27-linked polyubiquitination of K324 and K430 residues of NLRP3. Ubiquitination-defective NLRP3 mutants neither bind to NEK7, nor form NLRP3 oligomers, but remain binding to MAVS. Accordingly, NLRP3 mutants led to abortive ASC speck formation and diminished IL-1β production. We propose that MARCH5-dependent NLRP3 ubiquitination creates a docking site for NEK7 binding, playing as a fundamental step-wise regulator on the mitochondria.

Infection of hepatitis B virus (HBV) increase the incidence of chronic liver disease and hepatocellular carcinoma (HCC). The hepatitis B viral x (HBx) protein encoded by the HBV genome contributes to the pathogenesis of HCC and thus, negative regulation of HBx is beneficial for the alleviation of the disease pathogenesis. MARCH5 is a mitochondrial E3 ubiquitin ligase and here, we show that high MARCH5 expression levels are correlated with improved survival in HCC patients. MARCH5 interacts with HBx protein mainly accumulated in mitochondria and targets it for degradation. The N-terminal RING domain of MARCH5 was required for the interaction with HBx, and MARCH5H43W lacking E3 ligase activity failed to reduce HBx protein levels. High expression of HBx results in the formation of protein aggregates in semi-denaturing detergent agarose gels and MARCH5 mediates the elimination of protein aggregates through the proteasome pathway. HBx-induced ROS production, mitophagy, and cyclooxygenase-2 gene expression were suppressed in the presence of high MARCH5 expression. These results suggest MARCH5 as a target for alleviating HBV-mediated liver disease.

The chromatin remodeler RSF1 enriched at mitotic centromeres is essential for proper chromosome alignment and segregation and underlying mechanisms remain to be disclosed. We here show that PLK1 recruitment by RSF1 at centromeres creates an activating phosphorylation on Thr236 in the activation loop of Aurora B and this is indispensable for the Aurora B activation. In structural modeling the phosphorylated Thr236 enhances the base catalysis by Asp200 nearby, facilitating the Thr232 autophosphorylation. Accordingly, RSF1-PLK1 is central for Aurora B-mediated microtubule destabilization in error correction. However, under full microtubule-kinetochore attachment RSF1-PLK1 positions at kinetochores, halts activating Aurora B and phosphorylates BubR1, regardless of tension. Spatial movement of RSF1-PLK1 to kinetochores is triggered by Aurora B-mediated phosphorylation of centromeric histone H3 on Ser28. We propose a regulatory RSF1-PLK1 axis that spatiotemporally controls on/off switch on Aurora B. This feedback circuit among RSF1-PLK1-Aurora B may coordinate dynamic microtubule-kinetochore attachment in early mitosis when full tension yet to be generated., During cell division, chromosome alignment is engendered by connection of microtubules to kinetochores, coordinated by Aurora B and PLK1. Here, the authors show that the RSF1-PLK1 axis creates an activating phosphorylation on T236 in the GT motif of Aurora B and this is indispensable for Aurora B activation.

연구의 목적은 스페인의 테러리즘 대응 체계와 입법을 검토·분석하여 한국에서의 테러방지법 관련 논의에 시사점을 제공하는 것이다. 스페인의 테러리즘 대응 관련 입법의 주요 특징을 보면, 첫째, 테러리즘 관련 특별법이 없다. 9·11 테러 이후 서구의 주요 국가들이 테러방지법을 마련한 것과는 달리, 스페인은 형법이나 형사소송법 등의 법체계를 활용하여 테러리즘에...

본 연구의 목적은 미국의 테러리즘 대응 입법 과정과 테러리즘 대응법제의 특징을 분석함으로써 한국 테러리즘 대응법제에 시사점을 제공하는 것이다. 미국 테러리즘 대응법제의 특징은 다음과 같다. 첫째, 테러리즘대응 체제를 정비하였다. 둘째, 테러리즘의 개념을 확장하였다. 셋째, 테러리즘 수사권과 국가안보를 강화하였다. 넷째, 테러리즘 범죄에 대한 처벌을 강화하였다. 다섯째, 테러자금 에 대한 감시를 강화하였다. 여섯째, 범죄수사 정보의 공유체계를 수립하였다. 일곱째, 시민의 사생 활 보호와 정보수집의 투명성을 강화하였다. 이처럼 미국 테러리즘 대응법제에서는 테러리즘 대응 체계를 확립하고 테러리즘에 대한 강력한 대응으로 국가안보를 강화하면서도 시민의 사생활과 자유 를 보호하고 정보수집의 투명성을 확보하고자 노력하고 있다. 이는 ‘어떻게 국가안보와 시민의 자유 보호를 절충할 것인가’와 관련하여 최근 테러방지법 입법화에 성공한 우리사회에 시사하는 바가 크다.

Chromatin remodelers regulate the nucleosome barrier during transcription, DNA replication, and DNA repair. The chromatin remodeler RSF1 is enriched at mitotic centromeres, but the functional consequences of this enrichment are not completely understood. Shugoshin (Sgo1) protects centromeric cohesion during mitosis and requires BuB1-dependent histone H2A phosphorylation (H2A-pT120) for localization. Loss of Sgo1 at centromeres causes chromosome missegregation. Here, we show that RSF1 regulates Sgo1 localization to centromeres through coordinating a crosstalk between histone acetylation and phosphorylation. RSF1 interacts with and recruits HDAC1 to centromeres, where it counteracts TIP60-mediated acetylation of H2A at K118. This deacetylation is required for the accumulation of H2A-pT120 and Sgo1 deposition, as H2A-K118 acetylation suppresses H2A-T120 phosphorylation by Bub1. Centromeric tethering of HDAC1 prevents premature chromatid separation in RSF1 knockout cells. Our results indicate that RSF1 regulates the dynamics of H2A histone modifications at mitotic centromeres and contributes to the maintenance of chromosome stability.

Wip1 belongs to the protein phosphatase C (PP2C) family, of which expression is up-regulated by a number of external stresses, and serves as a stress modulator in normal physiological conditions. When overexpressed, premature dephosphorylation of stress-mediators by Wip1 results in abrogation of tumor surveillance, thus Wip1 acts as an oncogene. Previously, the functional regulation of Wip1 in cell-cycle progression by counteracting cellular G1 and G2/M checkpoint activity in response to DNA damage was reported. However, other than in stress conditions, the function and regulatory mechanism of Wip1 has not been fully determined. Herein, we demonstrated that protein regulation of Wip1 occurs in a cell cycle-dependent manner, which is directly governed by APC/C(Cdh1) at the end of mitosis. In particular, we also showed evidence that Wip1 phosphatase activity is closely associated with its own protein stability, suggesting that reduced phosphatase activity of Wip1 during mitosis could trigger its degradation. Furthermore, to verify the physiological role of its phosphatase activity during mitosis, we established doxycycline-inducible cell models, including a Wip1 wild type (WT) and phosphatase dead mutant (Wip1 DA). When ectopically expressing Wip1 WT, we observed a delay in the transition from metaphase to anaphase. In conclusion, these studies show that mitotic degradation of Wip1 by APC/C(Cdh1) is important for normal mitotic progression.

As a member of imitation switch (ISWI) family in ATP-dependent chromatin remodeling factors, RSF complex consists of SNF2h ATPase and Rsf-1. Although it has been reported that SNF2h ATPase is recruited to DNA damage sites (DSBs) in a poly(ADP-ribosyl) polymerase 1 (PARP1)-dependent manner in DNA damage response (DDR), the function of Rsf-1 is still elusive. Here we show that Rsf-1 is recruited to DSBs confirmed by various cellular analyses. Moreover, the initial recruitment of Rsf-1 and SNF2h to DSBs shows faster kinetics than that of γH2AX after micro-irradiation. Signals of Rsf-1 and SNF2h are retained over 30 min after micro-irradiation, whereas γH2AX signals are gradually reduced at 10 min. In addition, Rsf-1 is accumulated at DSBs in ATM-dependent manner, and the putative pSQ motifs of Rsf-1 by ATM are required for its accumulation at DSBs. Furtheremore, depletion of Rsf-1 attenuates the activation of DNA damage checkpoint signals and cell survival upon DNA damage. Finally, we demonstrate that Rsf-1 promotes homologous recombination repair (HRR) by recruiting resection factors RPA32 and Rad51. Thus, these findings reveal a new function of chromatin remodeler Rsf-1 as a guard in DNA damage checkpoints and homologous recombination repair.

Hepatitis B virus (HBV) X protein (HBx), encoded by the HBV genome, is involved in the development of HBV-mediated liver cancer, whose frequency is highly correlated with chromosomal instability (CIN). We reported previously that HBx induces mitotic checkpoint dysfunction by targeting the human serine/threonine kinase BubR1 (hBubR1). However, the underlying mechanism remained unresolved. Here, we show that HBx protein-associated protein α (HBxAPα)/Rsf-1 associates with hBubR1 and HBx in the chromatin fraction during mitosis. Depletion of HBxAPα/Rsf-1 abolished the interaction between HBx and hBubR1, indicating that HBxAPα/Rsf-1 mediates these interactions. Knockdown of HBxAPα/Rsf-1 with small interfering RNA did not affect the recruitment of hBubR1 to kinetochores; however, it did significantly impair HBx targeting to kinetochores. A deletion mutant analysis revealed that two Kunitz domains of HBx, the Cdc20-binding domain of hBubR1 and full-length of HBxAPα/Rsf-1 were essential for these interactions. Thus, binding of HBx to hBubR1, stabilized by HBxAPα/Rsf-1, significantly attenuated hBubR1 binding to Cdc20 and consequently increased the rate of mitotic aberrations. Collectively, our data show that the HBx impairs hBubR1 function and induces CIN through HBxAPα/Rsf-1, providing a novel mechanism for induction of genomic instability by a viral pathogen in hepatocarcinogenesis.

The trabecular microstructure of normal lunates and lunates with Kienböck’s disease was investigated using micro-computed tomography (micro-CT). Five lunates with advanced Kienböck’s disease were obtained during lunate excision and scaphocapitate fusion, and five control lunates were from embalmed cadavers. Microstructural morphometric parameters were measured using micro-CT images. Trabeculations of lunates with Kienböck’s disease were 2.67 times denser and 1.84 times thicker than those of normal lunates. Furthermore, bone surface areas were 1.43 times greater and bone volume 2.67 times greater, and structural model indices were significantly lower in lunates with Kienböck’s disease. The study estimated that high mechanical stress would be applied to lunates with Kienböck’s disease, and suggests that new bone formation and collapse may play important roles in the microstructural changes in the lunate with advanced Kienböck’s disease.

This study estimated the kinetics of the mono- and co-combustion of sewage sludge pellets and combustible wastes such as municipal solid waste (MSW) and refuse-derived fuel (RDF). Sewage sludge was manufactured into pellets with a diameter of 8, 12, or 16 mm and a length of 30 mm. The RDF was composed of paper and plastics and was formed into pellets with a diameter of 8 mm and a length of 30 mm. MSW samples were synthesized using combustible wastes such as garbage, paper, plastics, and wood. The MSW was adjusted to have a moisture content of around 40% after shredding to under 10 mm. A laboratory-scale batch type stoker incinerator was used for the combustion and the gas composition of the flue gas was measured. The activation energy was calculated using the experimental results, and then the relation of the decomposition rate and reaction time was evaluated using the shrinking core model. The decomposition rate of the sludge pellets decreased as their diameter and moisture content increased, and the co-combustion of sludge pellets and combustible waste was affected by the amount of combustible waste. The individual combustion rates of the cylindrical sludge pellets or RDF were mainly controlled by the chemical reaction, but in the case of shredded MSW it was mostly influenced by gas diffusion. The rate for the co-combustion of sludge pellets and combustible wastes was mainly determined by the combustion rate of the combustible waste. The activation energy of the 8-mm-diameter sludge pellets was between 6.70 and 10.0 kcal/mol, according to the moisture content, but it was lower for MSW and RDF. In the case of MSW co-combustion, the reaction rate accelerated as the moisture content of the sludge pellets decreased, but it was markedly increased by the addition of RDF, regardless of the sludge moisture content.

Mitochondria serve as platforms for innate immunity. The mitochondrial antiviral signalling (MAVS) protein forms aggregates that elicit robust type-I interferon induction on viral infection, but persistent MAVS signalling leads to host immunopathology; it remains unknown how these signalling aggregates are resolved. Here we identify the mitochondria-resident E3 ligase, MARCH5, as a negative regulator of MAVS aggregates. March5+/− mice and MARCH5-deficient immune cells exhibit low viral replication and elevated type-I interferon responses to RNA viruses. MARCH5 binds MAVS only during viral stimulation when MAVS forms aggregates, and these interactions require the RING domain of MARCH5 and the CARD domain of MAVS. MARCH5, but not its RING mutant (MARCH5H43W), reduces the level of MAVS aggregates. MARCH5 transfers ubiquitin to Lys7 and Lys500 of MAVS and promotes its proteasome-mediated degradation. Our results indicate that MARCH5 modulates MAVS-mediated antiviral signalling, preventing excessive immune reactions., RNA viral infections trigger an immune response mediated by the formation of aggregates of the MAVS protein. Here the authors show that the mitochondrial protein MARCH5 modulates this response by transferring ubiquitin to MAVS aggregates, thus promoting their proteasomal degradation.

Genetic instability is intimately associated with tumour development. In particular, liver cancers associated with hepatitis B virus (HBV) exhibit high genetic instability; however, our understanding of the underlying molecular mechanisms remains limited. In this study, we found that γ-H2AX, a marker of DNA double-strand breaks (DSBs), and the levels of phospho-Chk2 (p-Chk2, the activated form) were significantly elevated in HBV-associated hepatocellular carcinomas and neighbouring regenerating nodules. Likewise, introduction of the pHBV or pMyc-HBx plasmids into cells induced accumulation of γ-H2AX foci and increased the p-Chk2 level. In these cells, inhibitory phosphorylation of Cdc25C phosphatase (Ser(216)) and CDK1 (Tyr(15)) was elevated; consequently, cell-cycle progression was delayed at G2/M phase, suggesting that activation of the ATM-Chk2 pathway by the HBV X protein (HBx) induces cell-cycle delay. Accordingly, inhibition of ataxia telangiectasia mutated (ATM) by caffeine or siRNA abolished the increase in the p-Chk2 level and restored the delayed CDK1 kinase activity in ChangX cells. We also found that cytoplasmic HBx, but not nuclear HBx, induced reactive oxygen species (ROS) production and led to the accumulation of γ-H2AX foci and the increased p-Chk2 level. Together, these data indicate that HBx-induced ROS accumulation induces DNA damage that activates the ATM-Chk2 pathway. Our findings provide insight into the mechanisms of HBV pathogenesis.

Accumulation of PLK1 at kinetochores is essential for chromosome alignment and segregation; however, the mechanism underlying PLK1 recruitment to kinetochores remains unresolved. The chromatin remodeller RSF1 tightly associates with centromere proteins, but its mitotic function is unknown. Here we show that RSF1 localizes at mitotic kinetochores and directly binds PLK1. RSF1 depletion disrupts localization of PLK1 at kinetochores; the C-terminal fragment of RSF1, which can bind PLK1, is sufficient to restore PLK1 localization. Moreover, CDK1 phosphorylates RSF1 at Ser1375, and this phosphorylation is necessary for PLK1 recruitment. Subsequently, PLK1 phosphorylates RSF1 at Ser1359, stabilizing PLK1 deposition. Importantly, RSF1 depletion mimicks the chromosome misalignment phenotype resulting from PLK1 knockdown; these defects are rescued by RSF1 S1375D or RSF1 S1359D but not RSF1 S1375A, showing a functional link between phosphorylation of RSF1 and chromosome alignment. Together, these data show that RSF1 is an essential centromeric component that recruits PLK1 to kinetochores and plays a crucial role in faithful cell division., Polo-like kinase 1 (PLK1) is recruited to kinetochores during mitosis, where it is required for proper chromosome alignment. Lee et al. show that the chromatin-remodelling factor RSF1 is required for PLK1 recruitment, and that this function depends on phosphorylation of RSF1 by the mitotic kinase CDK1.

One of the key elements of e-commerce systems is the online product catalog. It provides sellers with a content management system that assembles, aggregates, normalizes, and distributes product information. It also provides potential buyers with an interactive interface that offers a multimedia representation of the product information as well as retrieval, classification and ordering services. In this paper, we discuss the interface aspect of online product catalogs, focusing on its ability to help shoppers navigate and analyze product information. Specifically, we present a new interactive interface for online product catalogs that effectively enables the shopper to navigate through the product information space and analytically select suitable products.

Planners in the steel industry design a set of steel slabs to satisfy the order book subject to constraints on (1) achieving a total designed weight for each order using multiples of an order-specific production size range, (2) minimum and maximum sizes for each slab, and (3) feasible assignments of multiple orders to the same slab. We developed a heuristic solution based on matchings and bin packing that a large steel plant uses daily in mill operations.

The increasingly dynamic nature of business-to-business electronic commerce has produced a recent shift away from fixed pricing and toward flexible pricing. Flexible pricing, as defined here, includes both differential pricing, in which different buyers may receive different prices based on expected valuations, and dynamic-pricing mechanisms, such as auctions, where prices and conditions are based on bids by market participants. In this paper we survey ongoing work in flexible pricing in the context of the supply chain, including revenue management, procurement, and supply-chain coordination. We review negotiation mechanisms for procurement, including optimization approaches to the evaluation of complex, multidimensional bids. We also discuss several applications of flexible pricing on the sell side, including pricing strategies for response to requests for quotes, dynamic pricing in a reverse logistics application, and pricing in the emerging area of hosted applications services. We conclude with a discussion of future research directions in this rapidly growing area.

Wip1 belongs to the protein phosphatase C (PP2C) family, of which expression is up-regulated by a number of external stresses, and serves as a stress modulator in normal physiological conditions. When overexpressed, premature dephosphorylation of stress-mediators by Wip1 results in abrogation of tumor surveillance, thus Wip1 acts as an oncogene. Previously, the functional regulation of Wip1 in cell-cycle progression by counteracting cellular G1 and G2/M checkpoint activity in response to DNA damage was reported. However, other than in stress conditions, the function and regulatory mechanism of Wip1 has not been fully determined. Herein, we demonstrated that protein regulation of Wip1 occurs in a cell cycle-dependent manner, which is directly governed by APC/C(Cdh1) at the end of mitosis. In particular, we also showed evidence that Wip1 phosphatase activity is closely associated with its own protein stability, suggesting that reduced phosphatase activity of Wip1 during mitosis could trigger its degradation. Furthermore, to verify the physiological role of its phosphatase activity during mitosis, we established doxycycline-inducible cell models, including a Wip1 wild type (WT) and phosphatase dead mutant (Wip1 DA). When ectopically expressing Wip1 WT, we observed a delay in the transition from metaphase to anaphase. In conclusion, these studies show that mitotic degradation of Wip1 by APC/C(Cdh1) is important for normal mitotic progression.

Granulocytic sarcoma is rare extramedullary tumor composed of myeloblasts and other granulocytic precursors. The majority of cases have been reported in association with acute myeloid leukemia (AML) or myeloproliferative disorders. Granulocytic sarcoma may occur in patients with myelodysplastic syndromes. Reports are very rare, especially in the brain. We report an unusual case of granulocytic sarcoma of the parenchyma of the brain, occurring in a patient with myelodysplastic syndrome, diagnosed by cerebro-spinal fluid cytology and magnetic resonance imaging brain scan.

Up until now, electronic negotiations have primarily focused on the trading of simple goods and services, where products can be described either by price alone or as a set of attributes. There is little support for trading complex, configurable products, such as computer systems, automobiles, insurances and services in general. Companies need to communicate offers including complex rules and business policies, and they need decision support to evaluate these offers. This paper describes RECO, a decision support system for the representation and evaluation of configurable offers. Configurable offers allow multiple values for each attribute and they include rules on how to combine the various attribute values and how to price a desired configuration. From a configurable offer, a user can extract offers for individual configurations. RECO provides a compact representation for configurable offers using prepositional logic, and helps a user in finding the top L configurations based on her preferences. In a multi-sourcing setting, it provides support for identifying the optimal sourcing strategy subject to considerations such as minimum/maximium number of winners and homogeneity of attributes across bids. We draw on mathematical programming, propositional logic and decision analysis.

Efficient selection of bids and product offerings is a key requirement in today's procurement processes. Purchasing managers have to make comprehensible selections among hundreds of alternatives considering several dozens of criteria. Selection problems like this are challenging, because they require the balancing of multiple, often conflicting objectives. Traditional approaches to multi-attribute decision making (MADM) are often insufficient in situations with a large number of attributes and alternatives. ABSolute is an application framework for electronic sourcing. The core component of ABSolute is an intelligent decision analysis engine for electronic procurement, which is designed for large amounts of alternatives and attributes. ABSolute enhances traditional approaches to decision making with advanced visualization capabilities and WORA, a new technique for weight assessment based on ordinal rankings of alternatives.

We present a new technique for automatic data reduction and pattern recognition of time-domain signals such as electrocardiogram (ECG) waveforms. Data reduction is important because only a few significant features of each heart beat are of interest in pattern analysis, while the patient data collection system acquires an enormous number of data samples. We present a significant point extraction algorithm, based on the analysis of curvature, that identifies data samples that represent clinically significant information in the ECG waveform. Data reduction rates of up to 1:10 are possible without significantly distorting the appearance of the waveform. This method is unique in that common procedures help in both data reduction as well as pattern recognition. Part II of this work deals specifically with pattern analysis of normal and abnormal heart beats.