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Sample records for deacetylases controls g1

  1. The periodic down regulation of Cyclin E gene expression from exit of mitosis to end of G(1) is controlled by a deacetylase- and E2F-associated bipartite repressor element.

    PubMed

    Polanowska, J; Fabbrizio, E; Le Cam, L; Trouche, D; Emiliani, S; Herrera, R; Sardet, C

    2001-07-12

    The expression of cyclin E and that of a few other bona fide cell cycle regulatory genes periodically oscillates every cycle in proliferating cells. Although numerous experiments have documented the role of E2F sites and E2F activities in the control of these genes as cells exit from G(0) to move through the initial G(1)/S phase transition, almost nothing is known on the role of E2Fs during the subsequent cell cycles. Here we show that a variant E2F-site that is part of the Cyclin E Repressor Module (CERM) (Le Cam et al., 1999b) accounts for the periodic down regulation of the cyclin E promoter observed between the exit from mitosis until the mid/late G(1) phase in exponentially cycling cells. This cell cycle-dependent repression correlates with the periodic binding of an atypical G(1)-specific high molecular weight p107-E2F complex (Cyclin E Repressor Complex: CERC2) that differs in both size and DNA binding behaviors from known p107-E2F complexes. Notably, affinity purified CERC2 displays a TSA-sensitive histone deacetylase activity and, consistent with this, derepression of the cyclin E promoter by trichostatin A depends on the CERM element. Altogether, this shows that the cell cycle-dependent control of cyclin E promoter in cycling cells is embroiled in acetylation pathways via the CERM-like E2F element.

  2. Invasive Cell Fate Requires G1 Cell-Cycle Arrest and Histone Deacetylase-Mediated Changes in Gene Expression.

    PubMed

    Matus, David Q; Lohmer, Lauren L; Kelley, Laura C; Schindler, Adam J; Kohrman, Abraham Q; Barkoulas, Michalis; Zhang, Wan; Chi, Qiuyi; Sherwood, David R

    2015-10-26

    Despite critical roles in development and cancer, the mechanisms that specify invasive cellular behavior are poorly understood. Through a screen of transcription factors in Caenorhabditis elegans, we identified G1 cell-cycle arrest as a precisely regulated requirement of the anchor cell (AC) invasion program. We show that the nuclear receptor nhr-67/tlx directs the AC into G1 arrest in part through regulation of the cyclin-dependent kinase inhibitor cki-1. Loss of nhr-67 resulted in non-invasive, mitotic ACs that failed to express matrix metalloproteinases or actin regulators and lack invadopodia, F-actin-rich membrane protrusions that facilitate invasion. We further show that G1 arrest is necessary for the histone deacetylase HDA-1, a key regulator of differentiation, to promote pro-invasive gene expression and invadopodia formation. Together, these results suggest that invasive cell fate requires G1 arrest and that strategies targeting both G1-arrested and actively cycling cells may be needed to halt metastatic cancer.

  3. Mechanisms of G1 cell cycle arrest and apoptosis in myeloma cells induced by hybrid-compound histone deacetylase inhibitor

    SciTech Connect

    Fujii, Seiko; Okinaga, Toshinori; Ariyoshi, Wataru; Takahashi, Osamu; Iwanaga, Kenjiro; Nishino, Norikazu; Tominaga, Kazuhiro; Nishihara, Tatsuji

    2013-05-10

    Highlights: •Novel histone deacetylase inhibitor Ky-2, remarkably inhibits myeloma cell growth. •Ky-2 demonstrates no cytotoxicity against normal lymphocytic cells. •Ky-2 induces cell cycle arrest through the cell cycle-associated proteins. •Ky-2 induces Bcl-2-inhibitable apoptosis through a caspase-dependent cascade. -- Abstract: Objectives: Histone deacetylase (HDAC) inhibitors are new therapeutic agents, used to treat various types of malignant cancers. In the present study, we investigated the effects of Ky-2, a hybrid-compound HDAC inhibitor, on the growth of mouse myeloma cells. Materials and methods: Myeloma cells, HS-72, P3U1, and mouse normal cells were used in this study. Effect of HDAC inhibitors on cell viability was determined by WST-assay and trypan blue assay. Cell cycle was analyzed using flow cytometer. The expression of cell cycle regulatory and the apoptosis associated proteins were examined by Western blot analysis. Hoechst’s staining was used to detect apoptotic cells. Results: Our findings showed that Ky-2 decreased the levels of HDACs, while it enhanced acetylation of histone H3. Myeloma cell proliferation was inhibited by Ky-2 treatment. Interestingly, Ky-2 had no cytotoxic effects on mouse normal cells. Ky-2 treatment induced G1-phase cell cycle arrest and accumulation of a sub-G1 phase population, while Western blotting analysis revealed that expressions of the cell cycle-associated proteins were up-regulated. Also, Ky-2 enhanced the cleavage of caspase-9 and -3 in myeloma cells, followed by DNA fragmentation. In addition, Ky-2 was not found to induce apoptosis in bcl-2 overexpressing myeloma cells. Conclusion: These findings suggest that Ky-2 induces apoptosis via a caspase-dependent cascade and Bcl-2-inhibitable mechanism in myeloma cells.

  4. G1/S control of anchorage-independent growth in the fibroblast cell cycle

    PubMed Central

    1991-01-01

    We have developed methodology to identify the block to anchorage- independent growth and position it within the fibroblast cell cycle. Results with NRK fibroblasts show that mitogen stimulation of the G0/G1 transition and G1-associated increases in cell size are minimally affected by loss of cell anchorage. In contrast, the induction of G1/S cell cycle genes and DNA synthesis is markedly inhibited when anchorage is blocked. Moreover, we demonstrate that the anchorage-dependent transition maps to late G1 and shortly before activation of the G1/S p34cdc2-like kinase. The G1/S block was also detectable in NIH-3T3 cells. Our results: (a) distinguish control of cell cycle progression by growth factors and anchorage; (b) indicate that anchorage mediates G1/S control in fibroblasts; and (c) identify a physiologic circumstance in which the phenotype of mammalian cell cycle arrest would closely resemble Saccharomyces cerevisiae START. The close correlation between anchorage independence in vitro and tumorigenicity in vivo emphasizes the key regulatory role for G1/S control in mammalian cells. PMID:1955482

  5. Defective in Mitotic Arrest 1 (Dma1) Ubiquitin Ligase Controls G1 Cyclin Degradation*

    PubMed Central

    Hernández-Ortega, Sara; Bru, Samuel; Ricco, Natalia; Ramírez, Sara; Casals, Núria; Jiménez, Javier; Isasa, Marta; Crosas, Bernat; Clotet, Josep

    2013-01-01

    Progression through the G1 phase of the cell cycle is controlled by diverse cyclin-dependent kinases (CDKs) that might be associated to numerous cyclin isoforms. Given such complexity, regulation of cyclin degradation should be crucial for coordinating progression through the cell cycle. In Saccharomyces cerevisiae, SCF is the only E3 ligase known to date to be involved in G1 cyclin degradation. Here, we report the design of a genetic screening that uncovered Dma1 as another E3 ligase that targets G1 cyclins in yeast. We show that the cyclin Pcl1 is ubiquitinated in vitro and in vivo by Dma1, and accordingly, is stabilized in dma1 mutants. We demonstrate that Pcl1 must be phosphorylated by its own CDK to efficiently interact with Dma1 and undergo degradation. A nonphosphorylatable version of Pcl1 accumulates throughout the cell cycle, demonstrating the physiological relevance of the proposed mechanism. Finally, we present evidence that the levels of Pcl1 and Cln2 are independently controlled in response to nutrient availability. This new previously unknown mechanism for G1 cyclin degradation that we report here could help elucidate the specific roles of the redundant CDK-cyclin complexes in G1. PMID:23264631

  6. Production of stable bispecific IgG1 by controlled Fab-arm exchange

    PubMed Central

    Gramer, Michael J; van den Bremer, Ewald TJ; van Kampen, Muriel D; Kundu, Amitava; Kopfmann, Peter; Etter, Eric; Stinehelfer, David; Long, Justin; Lannom, Tom; Noordergraaf, Esther H; Gerritsen, Jolanda; Labrijn, Aran F; Schuurman, Janine; van Berkel, Patrick HC; Parren, Paul WHI

    2013-01-01

    The manufacturing of bispecific antibodies can be challenging for a variety of reasons. For example, protein expression problems, stability issues, or the use of non-standard approaches for manufacturing can result in poor yield or poor facility fit. In this paper, we demonstrate the use of standard antibody platforms for large-scale manufacturing of bispecific IgG1 by controlled Fab-arm exchange. Two parental antibodies that each contain a single matched point mutation in the CH3 region were separately expressed in Chinese hamster ovary cells and manufactured at 1000 L scale using a platform fed-batch and purification process that was designed for standard antibody production. The bispecific antibody was generated by mixing the two parental molecules under controlled reducing conditions, resulting in efficient Fab-arm exchange of >95% at kg scale. The reductant was removed via diafiltration, resulting in spontaneous reoxidation of interchain disulfide bonds. Aside from the bispecific nature of the molecule, extensive characterization demonstrated that the IgG1 structural integrity was maintained, including function and stability. These results demonstrate the suitability of this bispecific IgG1 format for commercial-scale manufacturing using standard antibody manufacturing techniques. PMID:23995617

  7. A Whi7-anchored loop controls the G1 Cdk-cyclin complex at start.

    PubMed

    Yahya, Galal; Parisi, Eva; Flores, Alba; Gallego, Carme; Aldea, Martí

    2014-01-09

    Cells commit to a new cell cycle at Start by activation of the G1 Cdk-cyclin complex which, in turn, triggers a genome-wide transcriptional wave that executes the G1/S transition. In budding yeast, the Cdc28-Cln3 complex is regulated by an ER-retention mechanism that is important for proper cell size control. We have isolated small-cell-size CDC28 mutants showing impaired retention at the ER and premature accumulation of the Cln3 cyclin in the nucleus. The differential interactome of a quintuple Cdc28(wee) mutant pinpointed Whi7, a Whi5 paralog targeted by Cdc28 that associates to the ER in a phosphorylation-dependent manner. Our results demonstrate that the Cln3 cyclin and Whi7 act in a positive feedback loop to release the G1 Cdk-cyclin complex and trigger Start once a critical size has been reached, thus uncovering a key nonlinear mechanism at the earliest known events of cell-cycle entry.

  8. Synthesis of labile, serum-dependent protein in early G1 controls animal cell growth

    PubMed Central

    Rossow, Peter W.; Riddle, Veronica G. H.; Pardee, Arthur B.

    1979-01-01

    We present a model to account for several major observations on growth control of animal cells in culture. This model is tested by means of kinetic experiments which show that exponentially growing animal cells whose ability to synthesize total protein has been inhibited with cycloheximide (by up to 70%) grow at rates approximately proportional to their rates of protein synthesis. However, virtually the entire elongation of the cell cycle occurs in the part of the G1 phase that depends on a high concentration of serum in the medium. This part of the cycle has earlier been suggested to lie prior to the restriction point—i.e., the point beyond the main regulatory processes of G1. The remainder of the cycle, from restriction point to mitosis, is markedly insensitive to these concentrations of cycloheximide as well as to growth regulation. We quantitatively account for the specific lengthening of that part of the cycle involved in growth regulation by assuming that cells must accumulate a specific protein in a critical amount before they can proceed beyond the restriction point. The lability of this protein (half-life about 2 hr) makes its accumulation unusually sensitive to inhibition of total protein synthesis by cycloheximide. Its production appears to depend on growth factors provided by serum. The model can also account for greater variations of G1 durations as the growth of cell populations is made slower. It also predicts two sorts of quiescence: one of cells slowly traversing G1, in slightly suboptimal conditions; the other of cells that enter G0 under inadequate conditions. Transformation of different sorts could create cells with altered variables for initiation, synthesis, or inactivation of the regulatory protein or could altogether eliminate the need for the protein. PMID:291975

  9. Recruitment of Cln3 Cyclin to Promoters Controls Cell Cycle Entry via Histone Deacetylase and Other Targets

    PubMed Central

    Cai, Ying; Wijnen, Herman; Futcher, Bruce

    2009-01-01

    In yeast, the G1 cyclin Cln3 promotes cell cycle entry by activating the transcription factor SBF. In mammals, there is a parallel system for cell cycle entry in which cyclin dependent kinase (CDK) activates transcription factor E2F/Dp. Here we show that Cln3 regulates SBF by at least two different pathways, one involving the repressive protein Whi5, and the second involving Stb1. The Rpd3 histone deacetylase complex is also involved. Cln3 binds to SBF at the CLN2 promoter, and removes previously bound Whi5 and histone deacetylase. Adding extra copies of the SBF binding site to the cell delays Start, possibly by titrating Cln3. Since Rpd3 is the yeast ortholog of mammalian HDAC1, there is now a virtually complete analogy between the proteins regulating cell cycle entry in yeast (SBF, Cln3, Whi5 and Stb1, Rpd3) and mammals (E2F, Cyclin D, Rb, HDAC1). The cell may titrate Cln3 molecules against the number of SBF binding sites, and this could be the underlying basis of the size-control mechanism for Start. PMID:19823669

  10. p53 controls CDC7 levels to reinforce G1 cell cycle arrest upon genotoxic stress

    PubMed Central

    Tudzarova, Slavica; Dey, Ayona; Stoeber, Kai; Okorokov, Andrei L.; Williams, Gareth H.

    2016-01-01

    ABSTRACT DNA replication initiation is a key event in the cell cycle, which is dependent on 2 kinases - CDK2 and CDC7. Here we report a novel mechanism in which p53 induces G1 checkpoint and cell cycle arrest by downregulating CDC7 kinase in response to genotoxic stress. We demonstrate that p53 controls CDC7 stability post-transcriptionally via miR-192/215 and post-translationally via Fbxw7β E3 ubiquitin ligase. The p53-dependent pathway of CDC7 downregulation is interlinked with the p53-p21-CDK2 pathway, as p21-mediated inhibition of CDK2-dependent phosphorylation of CDC7 on Thr376 is required for GSK3ß-phosphorylation and Fbxw7ß-dependent degradation of CDC7. Notably, sustained oncogenic high levels of active CDC7 exert a negative feedback onto p53, leading to unrestrained S-phase progression and accumulation of DNA damage. Thus, p53-dependent control of CDC7 levels is essential for blocking G1/S cell-cycle transition upon genotoxic stress, thereby safeguarding the genome from instability and thus representing a novel general stress response. PMID:27611229

  11. Plant D-type cyclins and the control of G1 progression.

    PubMed Central

    Oakenfull, E Ann; Riou-Khamlichi, Catherine; Murray, James A H

    2002-01-01

    The basic pattern of controls that operate during the G1 phase of the plant cell cycle shows much closer similarity to animals than to the yeasts and other fungi. The activity of D-type cyclin (CycD) kinases is induced in response to stimulatory signals, and these phosphorylate the plant homologue of the retinoblastoma tumour susceptibility (Rb) protein. It is likely that Rb phosphorylation results in the activation of genes under the control of E2F transcription factors, including those required for S phase entry. As the initial triggers of the cascade, attention has focused on the CycDs, and a family of 10 genes is present in Arabidopsis, divided into three major and three minor groups. Analysis to date suggests that these groups are functionally distinct. PMID:12079670

  12. CDK-Dependent Hsp70 Phosphorylation Controls G1 Cyclin Abundance and Cell-Cycle Progression

    PubMed Central

    Truman, Andrew W.; Kristjansdottir, Kolbrun; Wolfgeher, Donald; Hasin, Naushaba; Polier, Sigrun; Zhang, Hong; Perrett, Sarah; Prodromou, Chrisostomos; Jones, Gary W.; Kron, Stephen J.

    2012-01-01

    Summary In budding yeast, the essential functions of Hsp70 chaperones Ssa1–4 are regulated through expression level, isoform specificity, and cochaperone activity. Suggesting a novel regulatory paradigm, we find that phosphorylation of Ssa1 T36 within a cyclin-dependent kinase (CDK) consensus site conserved among Hsp70 proteins alters cochaperone and client interactions. T36 phosphorylation triggers displacement of Ydj1, allowing Ssa1 to bind the G1 cyclin Cln3 and promote its degradation. The stress CDK Pho85 phosphorylates T36 upon nitrogen starvation or pheromone stimulation, destabilizing Cln3 to delay onset of S phase. In turn, the mitotic CDK Cdk1 phosphorylates T36 to block Cln3 accumulation in G2/M. Suggesting broad conservation from yeast to human, CDK-dependent phosphorylation of Hsc70 T38 similarly regulates Cyclin D1 binding and stability. These results establish an active role for Hsp70 chaperones as signal transducers mediating growth control of G1 cyclin abundance and activity. PMID:23217712

  13. Genes involved in cell cycle G1 checkpoint control are frequently mutated in human melanoma metastases.

    PubMed Central

    Platz, A.; Sevigny, P.; Norberg, T.; Ring, P.; Lagerlöf, B.; Ringborg, U.

    1996-01-01

    A common characteristic of cancer cells is unrestrained cell division. This may be caused by mutational changes in genes coding for components of cell cycle-controlling networks. Alterations in genes involved in G1 checkpoint control have been registered in many human tumours, and investigations from several laboratories show that such alterations, taken together, are the most frequent changes detected in cancer cells. The present paper describes mutational analysis by polymerase chain reaction-single-strand conformation polymorphism (PCR/SSCP) and nucleotide sequence analysis of the genes coding for the p15, p53 and N-ras proteins in 26 metastases from 25 melanoma patients. The registered mutation frequencies add together with previously registered mutations in p16 in the same patient samples to a substantial total frequency of 44% of patients with mutation in at least one of the investigated genes. These results show the occurrence of heterogeneous defects among components of the cell cycle controlling machinery in a human melanoma tumour sample collection and demonstrate that the total frequency of detected alterations increases with the number of cell cycle controlling genes included in the screening panel. Images Figure 1 PMID:8826861

  14. First passage times in M2[X ]|G |1 |R queue with hysteretic overload control policy

    NASA Astrophysics Data System (ADS)

    Pechinkin, Alexander V.; Razumchik, Rostislav R.; Zaryadov, Ivan S.

    2016-06-01

    One of the reported approaches towards the solution of overload problem in networks of SIP servers is the implementation of multi-level hysteretic control of arrivals in SIP servers. Each level, being the parameter of the policy, specifies operation mode of SIP server i.e. it implicitly indicates what SIP server must do with the arriving packets. The choice of parameters' values is not guided by standards and is usually left for the network owner. In general, all operation modes of the considered policy can be grouped into two groups: normal mode (when all arriving packets are accepted) and congested mode (when part or all arriving packets are being dropped). Such grouping may serve as the criteria for choosing parameters' values of the policy: pick those values which minimize SIP server sojourn time in congested mode. In this short note we propose some analytical results which facilitate the solution of stated minimization problem. The considered mathematical model of SIP server is the queueing system M2[X ]|G |1 |R with batch arrivals and bi-level hysteretic control policy, which specifies three operation modes: normal (customers both flows are accepted), overload (only customers from one flow are accepted), discard (customers from both flows are blocked/lost)). The switching between modes can occur only on service completions. Analytical method allowing computation of stationary sojourn times in different operation modes (as well as first passage times between modes) is presented in brief. Numerical example is given.

  15. Singularity-conquering ZG controllers of z2g1 type for tracking control of the IPC system

    NASA Astrophysics Data System (ADS)

    Zhang, Yunong; Yu, Xiaotian; Yin, Yonghua; Peng, Chen; Fan, Zhengping

    2014-09-01

    With wider investigations and applications of autonomous robotics and intelligent vehicles, the inverted pendulum on a cart (IPC) system has become more attractive for numerous researchers due to its concise and representative structure. In this article, the tracking-control problem of the IPC system is considered and investigated. Based on Zhang dynamics (ZD) and gradient dynamics (GD), a novel kind of ZG controllers are developed and investigated for achieving the tracking-control purpose, which contains controllers of z2g0 and z2g1 types according to the number of times of using the ZD and GD methods. Besides, theoretical analyses are presented to guarantee the global and exponential convergence performance of both z2g0 and z2g1 controllers. Computer simulations are further performed to substantiate the feasibility and effectiveness of ZG controllers. More importantly, comparative simulation results demonstrate that controllers of z2g1 type can conquer the singularity problem (i.e. the division-by-zero problem).

  16. Whi5 phosphorylation embedded in the G1/S network dynamically controls critical cell size and cell fate

    PubMed Central

    Palumbo, Pasquale; Vanoni, Marco; Cusimano, Valerio; Busti, Stefano; Marano, Francesca; Manes, Costanzo; Alberghina, Lilia

    2016-01-01

    In budding yeast, overcoming of a critical size to enter S phase and the mitosis/mating switch—two central cell fate events—take place in the G1 phase of the cell cycle. Here we present a mathematical model of the basic molecular mechanism controlling the G1/S transition, whose major regulatory feature is multisite phosphorylation of nuclear Whi5. Cln3–Cdk1, whose nuclear amount is proportional to cell size, and then Cln1,2–Cdk1, randomly phosphorylate both decoy and functional Whi5 sites. Full phosphorylation of functional sites releases Whi5 inhibitory activity, activating G1/S transcription. Simulation analysis shows that this mechanism ensures coherent release of Whi5 inhibitory action and accounts for many experimentally observed properties of mitotically growing or conjugating G1 cells. Cell cycle progression and transcriptional analyses of a Whi5 phosphomimetic mutant verify the model prediction that coherent transcription of the G1/S regulon and ensuing G1/S transition requires full phosphorylation of Whi5 functional sites. PMID:27094800

  17. Whi5 phosphorylation embedded in the G1/S network dynamically controls critical cell size and cell fate.

    PubMed

    Palumbo, Pasquale; Vanoni, Marco; Cusimano, Valerio; Busti, Stefano; Marano, Francesca; Manes, Costanzo; Alberghina, Lilia

    2016-04-20

    In budding yeast, overcoming of a critical size to enter S phase and the mitosis/mating switch--two central cell fate events--take place in the G1 phase of the cell cycle. Here we present a mathematical model of the basic molecular mechanism controlling the G1/S transition, whose major regulatory feature is multisite phosphorylation of nuclear Whi5. Cln3-Cdk1, whose nuclear amount is proportional to cell size, and then Cln1,2-Cdk1, randomly phosphorylate both decoy and functional Whi5 sites. Full phosphorylation of functional sites releases Whi5 inhibitory activity, activating G1/S transcription. Simulation analysis shows that this mechanism ensures coherent release of Whi5 inhibitory action and accounts for many experimentally observed properties of mitotically growing or conjugating G1 cells. Cell cycle progression and transcriptional analyses of a Whi5 phosphomimetic mutant verify the model prediction that coherent transcription of the G1/S regulon and ensuing G1/S transition requires full phosphorylation of Whi5 functional sites.

  18. Division of labour between Myc and G1 cyclins in cell cycle commitment and pace control.

    PubMed

    Dong, Peng; Maddali, Manoj V; Srimani, Jaydeep K; Thélot, François; Nevins, Joseph R; Mathey-Prevot, Bernard; You, Lingchong

    2014-09-01

    A body of evidence has shown that the control of E2F transcription factor activity is critical for determining cell cycle entry and cell proliferation. However, an understanding of the precise determinants of this control, including the role of other cell-cycle regulatory activities, has not been clearly defined. Here, recognizing that the contributions of individual regulatory components could be masked by heterogeneity in populations of cells, we model the potential roles of individual components together with the use of an integrated system to follow E2F dynamics at the single-cell level and in real time. These analyses reveal that crossing a threshold amplitude of E2F accumulation determines cell cycle commitment. Importantly, we find that Myc is critical in modulating the amplitude, whereas cyclin D/E activities have little effect on amplitude but do contribute to the modulation of duration of E2F activation, thereby affecting the pace of cell cycle progression.

  19. Physiological electric fields control the G1/S phase cell cycle checkpoint to inhibit endothelial cell proliferation.

    PubMed

    Wang, Entong; Yin, Yili; Zhao, Min; Forrester, John V; McCaig, Colin D

    2003-03-01

    Vascular endothelial cell (VEC) proliferation is a key event in angiogenesis and is tightly regulated. Electric potential differences exist around the vascular endothelium and give rise to endogenous electric fields (EFs), whether these EFs influence VEC proliferation is unclear. We exposed cultured VECs to applied EFs of physiological strengths for up to 72 h. EF at 50 or 100 mV/mm did not influence cell proliferation, but at 200 mV/mm, cell density, cell growth rate, and mitosis index decreased significantly. EF-induced reduction in VEC proliferation was not due to increased apoptosis, because caspase apoptosis inhibitor Z-VAD-FMK (20 microM), had no effect on this response. Rather, EF responses were mediated via decreased entry of cells into S phase from G1 phase, as shown by flow cytometry. Western blot showed that EFs decreased G1-specific cyclin E expression and increased cyclin/cyclin-dependent kinase complex inhibitor p27kipl expression. Thus EFs controlled VEC proliferation through induction of cell cycle arrest at G1 by down-regulation of cyclin E expression and up-regulation of p27kipl expression, rather than by promoting apoptosis. If control of the cell cycle by endogenous EFs extends beyond VECs, this would be of widespread biological significance in vivo.

  20. PHYTOCHROME B and HISTONE DEACETYLASE 6 Control Light-Induced Chromatin Compaction in Arabidopsis thaliana

    PubMed Central

    Pavlova, Penka; Clifton, Rachel; Pontvianne, Frédéric; Snoek, L. Basten; Millenaar, Frank F.; Schulkes, Roeland Kees; van Driel, Roel; Voesenek, Laurentius A. C. J.; Spillane, Charles; Pikaard, Craig S.; Fransz, Paul; Peeters, Anton J. M.

    2009-01-01

    Natural genetic variation in Arabidopsis thaliana exists for many traits and often reflects acclimation to local environments. Studying natural variation has proven valuable in the characterization of phenotypic traits and, in particular, in identifying genetic factors controlling these traits. It has been previously shown that chromatin compaction changes during development and biotic stress. To gain more insight into the genetic control of chromatin compaction, we investigated the nuclear phenotype of 21 selected Arabidopsis accessions from different geographic origins and habitats. We show natural variation in chromatin compaction and demonstrate a positive correlation with latitude of geographic origin. The level of compaction appeared to be dependent on light intensity. A novel approach, combining Quantitative Trait Locus (QTL) mapping and microscopic examination, pointed at PHYTOCHROME-B (PHYB) and HISTONE DEACETYLASE-6 (HDA6) as positive regulators of light-controlled chromatin compaction. Indeed, mutant analyses demonstrate that both factors affect global chromatin organization. HDA6, in addition, strongly promotes the light-mediated compaction of the Nucleolar Organizing Regions (NORs). The accession Cape Verde Islands-0 (Cvi-0), which shows sequence polymorphism in the PHYB gene and in the HDA6 promotor, resembles the hda6 mutant in having reduced chromatin compaction and decreased methylation levels of DNA and histone H3K9 at the NORs. We provide evidence that chromatin organization is controlled by light intensity. We propose that chromatin plasticity is associated with acclimation of Arabidopsis to its environment. The polymorphic alleles such as PHYB and HDA6 control this process. PMID:19730687

  1. Murine hematopoietic stem cell dormancy controlled by induction of a novel short form of PSF1 by histone deacetylase inhibitors

    SciTech Connect

    Han, Yinglu; Gong, Zhi-Yuan; Takakura, Nobuyuki

    2015-06-10

    Hematopoietic stem cells (HSCs) can survive long-term in a state of dormancy. Little is known about how histone deacetylase inhibitors (HDACi) affect HSC kinetics. Here, we use trichostatin A (TSA), a histone deacetylase inhibitor, to enforce histone acetylation and show that this suppresses cell cycle entry by dormant HSCs. Previously, we found that haploinsufficiency of PSF1, a DNA replication factor, led to attenuation of the bone marrow (BM) HSC pool size and lack of acute proliferation after 5-FU ablation. Because PSF1 protein is present in CD34{sup +} transiently amplifying HSCs but not in CD34{sup −} long-term reconstituting-HSCs which are resting in a dormant state, we analyzed the relationship between dormancy and PSF1 expression, and how a histone deacetylase inhibitor affects this. We found that CD34{sup +} HSCs produce long functional PSF1 (PSF1a) but CD34{sup −} HSCs produce a shorter possibly non-functional PSF1 (PSF1b, c, dominantly PSF1c). Using PSF1a-overexpressing NIH-3T3 cells in which the endogenous PSF1 promoter is suppressed, we found that TSA treatment promotes production of the shorter form of PSF1 possibly by inducing recruitment of E2F family factors upstream of the PSF1 transcription start site. Our data document one mechanism by which histone deacetylase inhibitors affect the dormancy of HSCs by regulating the DNA replication factor PSF1. - Highlights: • Hematopoetic stem cell dormancy is controlled by histone deacetylation inhibitors. • Dormancy of HSCs is associated with a shorter form of non-functional PSF1. • Histone deacetylase inhibitors suppress PSF1 promoter activity.

  2. Histone deacetylases control module-specific phenotypic plasticity in beetle weapons

    PubMed Central

    Ozawa, Takane; Mizuhara, Tomoko; Arata, Masataka; Shimada, Masakazu; Niimi, Teruyuki; Okada, Kensuke; Okada, Yasukazu; Ohta, Kunihiro

    2016-01-01

    Nutritional conditions during early development influence the plastic expression of adult phenotypes. Among several body modules of animals, the development of sexually selected exaggerated traits exhibits striking nutrition sensitivity, resulting in positive allometry and hypervariability distinct from other traits. Using de novo RNA sequencing and comprehensive RNA interference (RNAi) for epigenetic modifying factors, we found that histone deacetylases (HDACs) and polycomb group (PcG) proteins preferentially influence the size of mandibles (exaggerated male weapon) and demonstrate nutrition-dependent hypervariability in the broad-horned flour beetle, Gnatocerus cornutus. RNAi-mediated HDAC1 knockdown (KD) in G. cornutus larvae caused specific curtailment of mandibles in adults, whereas HDAC3 KD led to hypertrophy. Notably, these KDs conferred opposite effects on wing size, but little effect on the size of the core body and genital modules. PcG RNAi also reduced adult mandible size. These results suggest that the plastic development of exaggerated traits is controlled in a module-specific manner by HDACs. PMID:27956627

  3. Cytoplasmic-nuclear trafficking of G1/S cell cycle molecules and adult human β-cell replication: a revised model of human β-cell G1/S control.

    PubMed

    Fiaschi-Taesch, Nathalie M; Kleinberger, Jeffrey W; Salim, Fatimah G; Troxell, Ronnie; Wills, Rachel; Tanwir, Mansoor; Casinelli, Gabriella; Cox, Amy E; Takane, Karen K; Srinivas, Harish; Scott, Donald K; Stewart, Andrew F

    2013-07-01

    Harnessing control of human β-cell proliferation has proven frustratingly difficult. Most G1/S control molecules, generally presumed to be nuclear proteins in the human β-cell, are in fact constrained to the cytoplasm. Here, we asked whether G1/S molecules might traffic into and out of the cytoplasmic compartment in association with activation of cell cycle progression. Cdk6 and cyclin D3 were used to drive human β-cell proliferation and promptly translocated into the nucleus in association with proliferation. In contrast, the cell cycle inhibitors p15, p18, and p19 did not alter their location, remaining cytoplasmic. Conversely, p16, p21, and p27 increased their nuclear frequency. In contrast once again, p57 decreased its nuclear frequency. Whereas proliferating β-cells contained nuclear cyclin D3 and cdk6, proliferation generally did not occur in β-cells that contained nuclear cell cycle inhibitors, except p21. Dynamic cytoplasmic-nuclear trafficking of cdk6 was confirmed using green fluorescent protein-tagged cdk6 and live cell imaging. Thus, we provide novel working models describing the control of cell cycle progression in the human β-cell. In addition to known obstacles to β-cell proliferation, cytoplasmic-to-nuclear trafficking of G1/S molecules may represent an obstacle as well as a therapeutic opportunity for human β-cell expansion.

  4. Histone Deacetylases

    PubMed Central

    Parbin, Sabnam; Kar, Swayamsiddha; Shilpi, Arunima; Sengupta, Dipta; Deb, Moonmoon; Rath, Sandip Kumar

    2014-01-01

    In the current era of genomic medicine, diseases are identified as manifestations of anomalous patterns of gene expression. Cancer is the principal example among such maladies. Although remarkable progress has been achieved in the understanding of the molecular mechanisms involved in the genesis and progression of cancer, its epigenetic regulation, particularly histone deacetylation, demands further studies. Histone deacetylases (HDACs) are one of the key players in the gene expression regulation network in cancer because of their repressive role on tumor suppressor genes. Higher expression and function of deacetylases disrupt the finely tuned acetylation homeostasis in both histone and non-histone target proteins. This brings about alterations in the genes implicated in the regulation of cell proliferation, differentiation, apoptosis and other cellular processes. Moreover, the reversible nature of epigenetic modulation by HDACs makes them attractive targets for cancer remedy. This review summarizes the current knowledge of HDACs in tumorigenesis and tumor progression as well as their contribution to the hallmarks of cancer. The present report also describes briefly various assays to detect histone deacetylase activity and discusses the potential role of histone deacetylase inhibitors as emerging epigenetic drugs to cure cancer. PMID:24051359

  5. A General G1/S-Phase Cell-Cycle Control Module in the Flowering Plant Arabidopsis thaliana

    PubMed Central

    Zhao, Xin'Ai; Harashima, Hirofumi; Dissmeyer, Nico; Pusch, Stefan; Weimer, Annika K.; Bramsiepe, Jonathan; Bouyer, Daniel; Rademacher, Svenja; Nowack, Moritz K.; Novak, Bela; Sprunck, Stefanie; Schnittger, Arp

    2012-01-01

    The decision to replicate its DNA is of crucial importance for every cell and, in many organisms, is decisive for the progression through the entire cell cycle. A comparison of animals versus yeast has shown that, although most of the involved cell-cycle regulators are divergent in both clades, they fulfill a similar role and the overall network topology of G1/S regulation is highly conserved. Using germline development as a model system, we identified a regulatory cascade controlling entry into S phase in the flowering plant Arabidopsis thaliana, which, as a member of the Plantae supergroup, is phylogenetically only distantly related to Opisthokonts such as yeast and animals. This module comprises the Arabidopsis homologs of the animal transcription factor E2F, the plant homolog of the animal transcriptional repressor Retinoblastoma (Rb)-related 1 (RBR1), the plant-specific F-box protein F-BOX-LIKE 17 (FBL17), the plant specific cyclin-dependent kinase (CDK) inhibitors KRPs, as well as CDKA;1, the plant homolog of the yeast and animal Cdc2+/Cdk1 kinases. Our data show that the principle of a double negative wiring of Rb proteins is highly conserved, likely representing a universal mechanism in eukaryotic cell-cycle control. However, this negative feedback of Rb proteins is differently implemented in plants as it is brought about through a quadruple negative regulation centered around the F-box protein FBL17 that mediates the degradation of CDK inhibitors but is itself directly repressed by Rb. Biomathematical simulations and subsequent experimental confirmation of computational predictions revealed that this regulatory circuit can give rise to hysteresis highlighting the here identified dosage sensitivity of CDK inhibitors in this network. PMID:22879821

  6. G0/G1 Switch Gene 2 controls adipose triglyceride lipase activity and lipid metabolism in skeletal muscle.

    PubMed

    Laurens, Claire; Badin, Pierre-Marie; Louche, Katie; Mairal, Aline; Tavernier, Geneviève; Marette, André; Tremblay, Angelo; Weisnagel, S John; Joanisse, Denis R; Langin, Dominique; Bourlier, Virginie; Moro, Cedric

    2016-07-01

    Recent data suggest that adipose triglyceride lipase (ATGL) plays a key role in providing energy substrate from triglyceride pools and that alterations of its expression/activity relate to metabolic disturbances in skeletal muscle. Yet little is known about its regulation. We here investigated the role of the protein G0/G1 Switch Gene 2 (G0S2), recently described as an inhibitor of ATGL in white adipose tissue, in the regulation of lipolysis and oxidative metabolism in skeletal muscle. We first examined G0S2 protein expression in relation to metabolic status and muscle characteristics in humans. We next overexpressed and knocked down G0S2 in human primary myotubes to assess its impact on ATGL activity, lipid turnover and oxidative metabolism, and further knocked down G0S2 in vivo in mouse skeletal muscle. G0S2 protein is increased in skeletal muscle of endurance-trained individuals and correlates with markers of oxidative capacity and lipid content. Recombinant G0S2 protein inhibits ATGL activity by about 40% in lysates of mouse and human skeletal muscle. G0S2 overexpression augments (+49%, p < 0.05) while G0S2 knockdown strongly reduces (-68%, p < 0.001) triglyceride content in human primary myotubes and mouse skeletal muscle. We further show that G0S2 controls lipolysis and fatty acid oxidation in a strictly ATGL-dependent manner. These metabolic adaptations mediated by G0S2 are paralleled by concomitant changes in glucose metabolism through the modulation of Pyruvate Dehydrogenase Kinase 4 (PDK4) expression (5.4 fold, p < 0.001). Importantly, downregulation of G0S2 in vivo in mouse skeletal muscle recapitulates changes in lipid metabolism observed in vitro. Collectively, these data indicate that G0S2 plays a key role in the regulation of skeletal muscle ATGL activity, lipid content and oxidative metabolism.

  7. TEAD4-YAP interaction regulates tumoral growth by controlling cell-cycle arrest at the G1 phase.

    PubMed

    Takeuchi, Shin; Kasamatsu, Atsushi; Yamatoji, Masanobu; Nakashima, Dai; Endo-Sakamoto, Yosuke; Koide, Nao; Takahara, Toshikazu; Shimizu, Toshihiro; Iyoda, Manabu; Ogawara, Katsunori; Shiiba, Masashi; Tanzawa, Hideki; Uzawa, Katsuhiro

    2017-04-29

    TEA domain transcription factor 4 (TEAD4), which has critical functions in the process of embryonic development, is expressed in various cancers. However, the important role of TEAD4 in human oral squamous cell carcinomas (OSCCs) remain unclear. Here we investigated the TEAD4 expression level and the functional mechanism in OSCC using quantitative reverse transcriptase-polymerase chain reaction, Western blot analysis, and immunohistochemistry. Furthermore, TEAD4 knockdown model was used to evaluate cellular proliferation, cell-cycle analysis, and the interaction between TEAD4 and Yes-associated protein (YAP) which was reported to be a transcription coactivator of cellular proliferation. In the current study, we found that TEAD4 expression increased significantly in vitro and in vivo and correlated with tumoral size in OSCC patients. TEAD4 knockdown OSCC cells showed decreased cellular proliferation resulting from cell-cycle arrest in the G1 phase by down-regulation of cyclins, cyclin-dependent kinases (CDKs), and up-regulation of CDK inhibitors. We also found that the TEAD4-YAP complex in the nuclei may be related closely to transcriptions of G1 arrest-related genes. Taken together, we concluded that TEAD4 might play an important role in tumoral growth and have potential to be a therapeutic target in OSCCs. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. G1/S Inhibitors and the SWI/SNF Complex Control Cell-Cycle Exit during Muscle Differentiation.

    PubMed

    Ruijtenberg, Suzan; van den Heuvel, Sander

    2015-07-16

    The transition from proliferating precursor cells to post-mitotic differentiated cells is crucial for development, tissue homeostasis, and tumor suppression. To study cell-cycle exit during differentiation in vivo, we developed a conditional knockout and lineage-tracing system for Caenorhabditis elegans. Combined lineage-specific gene inactivation and genetic screening revealed extensive redundancies between previously identified cell-cycle inhibitors and the SWI/SNF chromatin-remodeling complex. Muscle precursor cells missing either SWI/SNF or G1/S inhibitor function could still arrest cell division, while simultaneous inactivation of these regulators caused continued proliferation and a C. elegans tumor phenotype. Further genetic analyses support that SWI/SNF acts in concert with hlh-1 MyoD, antagonizes Polycomb-mediated transcriptional repression, and suppresses cye-1 Cyclin E transcription to arrest cell division of muscle precursors. Thus, SWI/SNF and G1/S inhibitors provide alternative mechanisms to arrest cell-cycle progression during terminal differentiation, which offers insight into the frequent mutation of SWI/SNF genes in human cancers.

  9. Deletion of histone deacetylase 3 reveals critical roles in S phase progression and DNA damage control.

    PubMed

    Bhaskara, Srividya; Chyla, Brenda J; Amann, Joseph M; Knutson, Sarah K; Cortez, David; Sun, Zu-Wen; Hiebert, Scott W

    2008-04-11

    Histone deacetylases (HDACs) are enzymes that modify key residues in histones to regulate chromatin architecture, and they play a vital role in cell survival, cell-cycle progression, and tumorigenesis. To understand the function of Hdac3, a critical component of the N-CoR/SMRT repression complex, a conditional allele of Hdac3 was engineered. Cre-recombinase-mediated inactivation of Hdac3 led to a delay in cell-cycle progression, cell-cycle-dependent DNA damage, and apoptosis in mouse embryonic fibroblasts (MEFs). While no overt defects in mitosis were observed in Hdac3-/- MEFs, including normal H3Ser10 phosphorylation, DNA damage was observed in Hdac3-/- interphase cells, which appears to be associated with defective DNA double-strand break repair. Moreover, we noted that Hdac3-/- MEFs were protected from DNA damage when quiescent, which may provide a mechanistic basis for the action of HDAC inhibitors on cycling tumor cells.

  10. A circadian rhythm orchestrated by histone deacetylase 3 controls hepatic lipid metabolism.

    PubMed

    Feng, Dan; Liu, Tao; Sun, Zheng; Bugge, Anne; Mullican, Shannon E; Alenghat, Theresa; Liu, X Shirley; Lazar, Mitchell A

    2011-03-11

    Disruption of the circadian clock exacerbates metabolic diseases, including obesity and diabetes. We show that histone deacetylase 3 (HDAC3) recruitment to the genome displays a circadian rhythm in mouse liver. Histone acetylation is inversely related to HDAC3 binding, and this rhythm is lost when HDAC3 is absent. Although amounts of HDAC3 are constant, its genomic recruitment in liver corresponds to the expression pattern of the circadian nuclear receptor Rev-erbα. Rev-erbα colocalizes with HDAC3 near genes regulating lipid metabolism, and deletion of HDAC3 or Rev-erbα in mouse liver causes hepatic steatosis. Thus, genomic recruitment of HDAC3 by Rev-erbα directs a circadian rhythm of histone acetylation and gene expression required for normal hepatic lipid homeostasis.

  11. Non-sirtuin histone deacetylases in the control of cardiac aging.

    PubMed

    Ferguson, Bradley S; McKinsey, Timothy A

    2015-06-01

    Histone deacetylases (HDACs) catalyze the removal of acetyl-groups from lysine residues within nucelosomal histone tails and thousands of non-histone proteins. The 18 mammalian HDACs are grouped into four classes. Classes I, II and IV HDACs employ zinc as a co-factor for catalytic activity, while class III HDACs (also known as sirtuins) require NAD+ for enzymatic function. Small molecule inhibitors of zinc-dependent HDACs are efficacious in multiple pre-clinical models of pressure overload and ischemic cardiomyopathy, reducing pathological hypertrophy and fibrosis, and improving contractile function. Emerging data have revealed numerous mechanisms by which HDAC inhibitors benefit the heart, including suppression of oxidative stress and inflammation, inhibition of MAP kinase signaling, and enhancement of cardiac protein aggregate clearance and autophagic flux. Here, we summarize recent findings with zinc-dependent HDACs and HDAC inhibitors in the heart, focusing on newly described functions for distinct HDAC isoforms (e.g. HDAC2, HDAC3 and HDAC6). Potential for pharmacological HDAC inhibition as a means of treating age-related cardiac dysfunction is also discussed. This article is part of a Special Issue entitled: CV Aging.

  12. Geothermal district G1

    SciTech Connect

    Not Available

    1988-12-01

    Geothermal District G1 includes 37 northeastern California counties and six geothermal fields: Lake City, Susanville, Litchfield, Wendel, Amedee, and Casa Diablo. Electrical generation from geothermal resources occurs in three of the fields: Wendel, Amedee, and Casa Diablo. Low-temperature geothermal projects are underway throughout the district and are described in a road log format. The ten projects described are located at Big Bend, Glass Mountain, Bieber, Alturas, Cedarville, Lake City, Honey Lake Valley, Greenville, and in Sierra and Mono Counties.

  13. Histone deacetylase enzymes as drug targets for the control of the sheep blowfly, Lucilia cuprina

    PubMed Central

    Kotze, Andrew C.; Hines, Barney M.; Bagnall, Neil H.; Anstead, Clare A.; Gupta, Praveer; Reid, Robert C.; Ruffell, Angela P.; Fairlie, David P.

    2015-01-01

    The Australian sheep blowfly, Lucilia cuprina, is an ecto-parasite that causes significant economic losses in the sheep industry. Emerging resistance to insecticides used to protect sheep from this parasite is driving the search for new drugs that act via different mechanisms. Inhibitors of histone deacetylases (HDACs), enzymes essential for regulating eukaryotic gene transcription, are prospective new insecticides based on their capacity to kill human parasites. The blowfly genome was found here to contain five HDAC genes corresponding to human HDACs 1, 3, 4, 6 and 11. The catalytic domains of blowfly HDACs 1 and 3 have high sequence identity with corresponding human and other Dipteran insect HDACs (Musca domestica and Drosophila melanogaster). On the other hand, HDACs 4, 6 and 11 from the blowfly and the other Dipteran species showed up to 53% difference in catalytic domain amino acids from corresponding human sequences, suggesting the possibility of developing HDAC inhibitors specific for insects as desired for a commercial insecticide. Differences in transcription patterns for different blowfly HDACs through the life cycle, and between the sexes of adult flies, suggest different functions in regulating gene transcription within this organism and possibly different vulnerabilities. Data that supports HDACs as possible new insecticide targets is the finding that trichostatin A and suberoylanilide hydroxamic acid retarded growth of early instar blowfly larvae in vitro, and reduced the pupation rate. Trichostatin A was 8-fold less potent than the commercial insecticide cyromazine in inhibiting larval growth. Our results support further development of inhibitors of blowfly HDACs with selectivity over human and other mammalian HDACs as a new class of prospective insecticides for sheep blowfly. PMID:27120067

  14. The Hos2 Histone Deacetylase Controls Ustilago maydis Virulence through Direct Regulation of Mating-Type Genes

    PubMed Central

    Elías-Villalobos, Alberto; Fernández-Álvarez, Alfonso; Moreno-Sánchez, Ismael; Helmlinger, Dominique; Ibeas, José I.

    2015-01-01

    Morphological changes are critical for host colonisation in plant pathogenic fungi. These changes occur at specific stages of their pathogenic cycle in response to environmental signals and are mediated by transcription factors, which act as master regulators. Histone deacetylases (HDACs) play crucial roles in regulating gene expression, for example by locally modulating the accessibility of chromatin to transcriptional regulators. It has been reported that HDACs play important roles in the virulence of plant fungi. However, the specific environment-sensing pathways that control fungal virulence via HDACs remain poorly characterised. Here we address this question using the maize pathogen Ustilago maydis. We find that the HDAC Hos2 is required for the dimorphic switch and pathogenic development in U. maydis. The deletion of hos2 abolishes the cAMP-dependent expression of mating type genes. Moreover, ChIP experiments detect Hos2 binding to the gene bodies of mating-type genes, which increases in proportion to their expression level following cAMP addition. These observations suggest that Hos2 acts as a downstream component of the cAMP-PKA pathway to control the expression of mating-type genes. Interestingly, we found that Clr3, another HDAC present in U. maydis, also contributes to the cAMP-dependent regulation of mating-type gene expression, demonstrating that Hos2 is not the only HDAC involved in this control system. Overall, our results provide new insights into the role of HDACs in fungal phytopathogenesis. PMID:26317403

  15. Production of stable bispecific IgG1 by controlled Fab-arm exchange: scalability from bench to large-scale manufacturing by application of standard approaches.

    PubMed

    Gramer, Michael J; van den Bremer, Ewald T J; van Kampen, Muriel D; Kundu, Amitava; Kopfmann, Peter; Etter, Eric; Stinehelfer, David; Long, Justin; Lannom, Tom; Noordergraaf, Esther H; Gerritsen, Jolanda; Labrijn, Aran F; Schuurman, Janine; van Berkel, Patrick H C; Parren, Paul W H I

    2013-01-01

    The manufacturing of bispecific antibodies can be challenging for a variety of reasons. For example, protein expression problems, stability issues, or the use of non-standard approaches for manufacturing can result in poor yield or poor facility fit. In this paper, we demonstrate the use of standard antibody platforms for large-scale manufacturing of bispecific IgG1 by controlled Fab-arm exchange. Two parental antibodies that each contain a single matched point mutation in the CH3 region were separately expressed in Chinese hamster ovary cells and manufactured at 1000 L scale using a platform fed-batch and purification process that was designed for standard antibody production. The bispecific antibody was generated by mixing the two parental molecules under controlled reducing conditions, resulting in efficient Fab-arm exchange of>95% at kg scale. The reductant was removed via diafiltration, resulting in spontaneous reoxidation of interchain disulfide bonds. Aside from the bispecific nature of the molecule, extensive characterization demonstrated that the IgG1 structural integrity was maintained, including function and stability. These results demonstrate the suitability of this bispecific IgG1 format for commercial-scale manufacturing using standard antibody manufacturing techniques.

  16. The histone deacetylase HDA19 controls root cell elongation and modulates a subset of phosphate starvation responses in Arabidopsis

    PubMed Central

    Chen, Chun-Ying; Wu, Keqiang; Schmidt, Wolfgang

    2015-01-01

    The length of root epidermal cells and their patterning into files of hair-bearing and non-hair cells are genetically determined but respond with high plasticity to environmental cues. Limited phyto-availability of the essential mineral nutrient phosphate (Pi) increases the number of root hairs by longitudinal shortening of epidermal cells and by reprogramming the fate of cells in positions normally occupied by non-hair cells. Through analysis of the root morphology and transcriptional profiles from transgenic Arabidopsis lines with altered expression of the histone deacetylase HDA19, we show that in an intricate interplay of Pi availability and intrinsic factors, HDA19 controls the epidermal cell length, probably by altering the positional bias that dictates epidermal patterning. In addition, HDA19 regulates several Pi-responsive genes that encode proteins with important regulatory or metabolic roles in the acclimation to Pi deficiency. In particular, HDA19 affects genes encoding SPX (SYG1/Pho81/XPR) domain-containing proteins and genes involved in membrane lipid remodeling, a key response to Pi starvation that increases the free Pi in plants. Our data add a novel, non-transcriptionally regulated component of the Pi signaling network and emphasize the importance of reversible post-translational histone modification for the integration of external signals into intrinsic developmental and metabolic programs. PMID:26508133

  17. Chitin Deacetylases: Properties and Applications

    PubMed Central

    Zhao, Yong; Park, Ro-Dong; Muzzarelli, Riccardo A. A.

    2010-01-01

    Chitin deacetylases, occurring in marine bacteria, several fungi and a few insects, catalyze the deacetylation of chitin, a structural biopolymer found in countless forms of marine life, fungal cell and spore walls as well as insect cuticle and peritrophic matrices. The deacetylases recognize a sequence of four GlcNAc units in the substrate, one of which undergoes deacetylation: the resulting chitosan has a more regular deacetylation pattern than a chitosan treated with hot NaOH. Nevertheless plain chitin is a poor substrate, but glycolated, reprecipitated or depolymerized chitins are good ones. The marine Vibrio sp. colonize the chitin particles and decompose the chitin thanks to the concerted action of chitinases and deacetylases, otherwise they could not tolerate chitosan, a recognized antibacterial biopolymer. In fact, chitosan is used to prevent infections in fishes and crustaceans. Considering that chitin deacetylases play very important roles in the biological attack and defense systems, they may find applications for the biological control of fungal plant pathogens or insect pests in agriculture and for the biocontrol of opportunistic fungal human pathogens. PMID:20161969

  18. Chitin deacetylases: properties and applications.

    PubMed

    Zhao, Yong; Park, Ro-Dong; Muzzarelli, Riccardo A A

    2010-01-14

    Chitin deacetylases, occurring in marine bacteria, several fungi and a few insects, catalyze the deacetylation of chitin, a structural biopolymer found in countless forms of marine life, fungal cell and spore walls as well as insect cuticle and peritrophic matrices. The deacetylases recognize a sequence of four GlcNAc units in the substrate, one of which undergoes deacetylation: the resulting chitosan has a more regular deacetylation pattern than a chitosan treated with hot NaOH. Nevertheless plain chitin is a poor substrate, but glycolated, reprecipitated or depolymerized chitins are good ones. The marine Vibrio sp. colonize the chitin particles and decompose the chitin thanks to the concerted action of chitinases and deacetylases, otherwise they could not tolerate chitosan, a recognized antibacterial biopolymer. In fact, chitosan is used to prevent infections in fishes and crustaceans. Considering that chitin deacetylases play very important roles in the biological attack and defense systems, they may find applications for the biological control of fungal plant pathogens or insect pests in agriculture and for the biocontrol of opportunistic fungal human pathogens.

  19. Deregulated G1-S control and energy stress contribute to the synthetic-lethal interactions between inactivation of RB and TSC1 or TSC2.

    PubMed

    Gordon, Gabriel M; Zhang, Tianyi; Zhao, Jiong; Du, Wei

    2013-05-01

    Synthetic lethality is a potential strategy for cancer treatment by specifically promoting the death of cancer cells with particular defects such as the loss of the RB (RB1) tumor suppressor. We previously showed that inactivation of both RB and TSC2 induces synergistic apoptosis during the development of Drosophila melanogaster and in cancer cells. However, the in vivo mechanism of this synthetic-lethal interaction is not clear. Here, we show that synergistic cell death in tissues that have lost the RB and TSC orthologs rbf and dtsc1/gig, respectively, or overexpress Rheb and dE2F1, are correlated with synergistic defects in G1-S control, which causes cells to accumulate DNA damage. Coexpression of the G1-S inhibitor Dap, but not the G2-M inhibitor dWee1, decreases DNA damage and reduces cell death. In addition, we show that rbf and dtsc1 mutant cells are under energy stress, are sensitive to decreased energy levels and depend on the cellular energy stress-response pathway for survival. Decreasing mitochondrial ATP synthesis by inactivating cova or abrogating the energy-stress response by removing the metabolic regulator LKB1 both enhance the elimination of cells lacking either rbf or dtsc1. These observations, in conjunction with the finding that deregulation of TORC1 induces activation of JNK, indicate that multiple cellular stresses are induced and contribute to the synthetic-lethal interactions between RB and TSC1/TSC2 inactivation. The insights gained from this study suggest new approaches for targeting RB-deficient cancers.

  20. C/EBPα regulates CRL4Cdt2-mediated degradation of p21 in response to UVB-induced DNA damage to control the G1/S checkpoint

    PubMed Central

    Hall, Jonathan R; Bereman, Michael S; Nepomuceno, Angelito I; Thompson, Elizabeth A; Muddiman, David C; Smart, Robert C

    2014-01-01

    The bZIP transcription factor, C/EBPα is highly inducible by UVB and other DNA damaging agents in keratinocytes. C/EBPα-deficient keratinocytes fail to undergo cell cycle arrest in G1 in response to UVB-induced DNA damage and mice lacking epidermal C/EBPα are highly susceptible to UVB-induced skin cancer. The mechanism through which C/EBPα regulates the cell cycle checkpoint in response to DNA damage is unknown. Here we report untreated C/EBPα-deficient keratinocytes have normal levels of the cyclin-dependent kinase inhibitor, p21, however, UVB-treated C/EBPα-deficient keratinocytes fail to up-regulate nuclear p21 protein levels despite normal up-regulation of Cdkn1a mRNA levels. UVB-treated C/EBPα-deficient keratinocytes displayed a 4-fold decrease in nuclear p21 protein half-life due to the increased proteasomal degradation of p21 via the E3 ubiquitin ligase CRL4Cdt2. Cdt2 is the substrate recognition subunit of CRL4Cdt2 and Cdt2 mRNA and protein levels were up-regulated in UVB-treated C/EBPα-deficient keratinocytes. Knockdown of Cdt2 restored p21 protein levels in UVB-treated C/EBPα-deficient keratinocytes. Lastly, the failure to accumulate p21 in response to UVB in C/EBPα-deficient keratinocytes resulted in decreased p21 interactions with critical cell cycle regulatory proteins, increased CDK2 activity, and inappropriate entry into S-phase. These findings reveal C/EBPα regulates G1/S cell cycle arrest in response to DNA damage via the control of CRL4Cdt2 mediated degradation of p21. PMID:25483090

  1. Ablation of the Retinoblastoma gene family deregulates G1 control causing immortalization and increased cell turnover under growth-restricting conditions

    PubMed Central

    Dannenberg, Jan-Hermen; van Rossum, Agnes; Schuijff, Leontine; te Riele, Hein

    2000-01-01

    The retinoblastoma suppressor pRB belongs to the family of so-called pocket proteins, which also includes p107 and p130. These proteins may functionally overlap in cell cycle control and tumor suppression. We have generated an isogenic set of embryonic stem (ES) cell lines carrying single or compound loss-of-function mutations in the Rb gene family, including a cell line completely devoid of all three pocket proteins. None of the knockout combinations affected the growth characteristics of ES cells; however, concomitant ablation of all three pocket proteins strongly impaired their differentiation capacity. For the generated genotypes, primary mouse embryonic fibroblasts (MEFs) also were obtained. While inactivation of Rb alone did not alleviate the senescence response of MEFs, pRB/p107-deficient MEFs, after having adapted to in vitro culturing, continued to proliferate at modest rate. Additional ablation of p130 rendered MEFs completely insensitive to senescence-inducing signals and strongly increased their proliferation rate. Although triple-knockout MEFs retained anchorage dependence, they lacked proper G1 control and showed increased cell turnover under growth-inhibiting conditions. PMID:11114893

  2. Chitin deacetylase product inhibition.

    PubMed

    Jaworska, Malgorzata M

    2011-02-01

    Chitin deacetylase is the only known enzyme catalyzing the hydrolysis of the acetamino linkage in the N-acetylglucosamine units of chitin and chitosan. This reaction can play an important role in enzymatic production of chitosan from chitin, or in enzymatic modification of chitosan, which has applications in medicine, pharmacy or plant protection. It was previously shown that acetic acid, a product of the deacetylation process, may act as an inhibitor of chitin deacetylase. Here we show the mechanism of inhibition of chitin deacetylase isolated from Absidia orchidis vel coerulea by acetic acid released during the deacetylation process. The process follows competitive inhibition with respect to acetic acid with an inhibition constant of K(i) = 0.286 mmol/L. These results will help to find the optimal system to carry out the enzymatic deacetylation process for industrial applications.

  3. The Arabidopsis N-Acetylornithine Deacetylase Controls Ornithine Biosynthesis via a Linear Pathway with Downstream Effects on Polyamine Levels.

    PubMed

    Molesini, Barbara; Zanzoni, Serena; Mennella, Giuseppe; Francese, Gianluca; Losa, Alessia; L Rotino, Giuseppe; Pandolfini, Tiziana

    2017-01-01

    Arabidopsis thaliana At4g17830 codes for a protein showing sequence similarity with the Escherichia coli N-acetylornithine deacetylase (EcArgE), an enzyme implicated in the linear ornithine (Orn) biosynthetic pathway. In plants, N-acetylornithine deacetylase (NAOD) activity has yet to be demonstrated; however, At4g17830-silenced and mutant (atnaod) plants display an impaired reproductive phenotype and altered foliar levels of Orn and polyamines (PAs). Here, we showed the direct connection between At4g17830 function and Orn biosynthesis, demonstrating biochemically that At4g17830 codes for a NAOD. These results are the first experimental proof that Orn can be produced in Arabidopsis via a linear pathway. In this study, to identify the role of AtNAOD in reproductive organs, we carried out a transcriptomic analysis on atnaod mutant and wild-type flowers. In the atnaod mutant, the most relevant effects were the reduced expression of cysteine-rich peptide-coding genes, known to regulate male-female cross-talk during reproduction, and variation in the expression of genes involved in nitrogen:carbon (N:C) status. The atnaod mutant also exhibited increased levels of sucrose and altered sensitivity to glucose. We hypothesize that AtNAOD participates in Orn and PA homeostasis, contributing to maintain an optimal N:C balance during reproductive development. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  4. Novel Interaction of Class IIb Histone Deacetylase 6 (HDAC6) with Class IIa HDAC9 Controls Gonadotropin Releasing Hormone (GnRH) Neuronal Cell Survival and Movement.

    PubMed

    Salian-Mehta, Smita; Xu, Mei; McKinsey, Timothy A; Tobet, Stuart; Wierman, Margaret E

    2015-05-29

    The impact of histone deacetylases (HDACs) in the control of gonadotropin releasing hormone (GnRH) neuronal development is unknown. We identified an increase in many HDACs in GT1-7 (differentiated) compared with NLT (undifferentiated) GnRH neuronal cell lines. Increased HDAC9 mRNA and protein and specific deacetylase activity in GT1-7 cells suggested a functional role. Introduction of HDAC9 in NLT cells protected from serum withdrawal induced apoptosis and impaired basal neuronal cell movement. Conversely, silencing of endogenous HDAC9 in GT1-7 cells increased apoptosis and cell movement. Comparison of WT and mutant HDAC9 constructs demonstrated that the HDAC9 pro-survival effects required combined cytoplasmic and nuclear localization, whereas the effects on cell movement required a cytoplasmic site of action. Co-immunoprecipitation demonstrated a novel interaction of HDAC9 selectively with the Class IIb HDAC6. HDAC6 was also up-regulated at the mRNA and protein levels, and HDAC6 catalytic activity was significantly increased in GT1-7 compared with NLT cells. HDAC9 interacted with HDAC6 through its second catalytic domain. Silencing of HDAC6, HDAC9, or both, in GT1-7 cells augmented apoptosis compared with controls. HDAC6 and -9 had additive effects to promote cell survival via modulating the BAX/BCL2 pathway. Silencing of HDAC6 resulted in an activation of movement of GT1-7 cells with induction in acetylation of α-tubulin. Inhibition of HDAC6 and HDAC9 together resulted in an additive effect to increase cell movement but did not alter the acetylation of αtubulin. Together, these studies identify a novel interaction of Class IIa HDAC9 with Class IIb HDAC6 to modulate cell movement and survival in GnRH neurons. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Novel Interaction of Class IIb Histone Deacetylase 6 (HDAC6) with Class IIa HDAC9 Controls Gonadotropin Releasing Hormone (GnRH) Neuronal Cell Survival and Movement*

    PubMed Central

    Salian-Mehta, Smita; Xu, Mei; McKinsey, Timothy A.; Tobet, Stuart; Wierman, Margaret E.

    2015-01-01

    The impact of histone deacetylases (HDACs) in the control of gonadotropin releasing hormone (GnRH) neuronal development is unknown. We identified an increase in many HDACs in GT1-7 (differentiated) compared with NLT (undifferentiated) GnRH neuronal cell lines. Increased HDAC9 mRNA and protein and specific deacetylase activity in GT1-7 cells suggested a functional role. Introduction of HDAC9 in NLT cells protected from serum withdrawal induced apoptosis and impaired basal neuronal cell movement. Conversely, silencing of endogenous HDAC9 in GT1-7 cells increased apoptosis and cell movement. Comparison of WT and mutant HDAC9 constructs demonstrated that the HDAC9 pro-survival effects required combined cytoplasmic and nuclear localization, whereas the effects on cell movement required a cytoplasmic site of action. Co-immunoprecipitation demonstrated a novel interaction of HDAC9 selectively with the Class IIb HDAC6. HDAC6 was also up-regulated at the mRNA and protein levels, and HDAC6 catalytic activity was significantly increased in GT1-7 compared with NLT cells. HDAC9 interacted with HDAC6 through its second catalytic domain. Silencing of HDAC6, HDAC9, or both, in GT1-7 cells augmented apoptosis compared with controls. HDAC6 and -9 had additive effects to promote cell survival via modulating the BAX/BCL2 pathway. Silencing of HDAC6 resulted in an activation of movement of GT1-7 cells with induction in acetylation of α-tubulin. Inhibition of HDAC6 and HDAC9 together resulted in an additive effect to increase cell movement but did not alter the acetylation of αtubulin. Together, these studies identify a novel interaction of Class IIa HDAC9 with Class IIb HDAC6 to modulate cell movement and survival in GnRH neurons. PMID:25873389

  6. The Histone Deacetylase Sirt6 Controls Embryonic Stem Cell Fate Via Tet-Mediated Production of 5-Hydroxymethylcytosine

    PubMed Central

    Etchegaray, Jean-Pierre; Chavez, Lukas; Huang, Yun; Ross, Kenneth N.; Choi, Jiho; Martinez-Pastor, Barbara; Walsh, Ryan M.; Sommer, Cesar A.; Lienhard, Matthias; Kugel, Sita; Silberman, Dafne M.; Ramaswamy, Sridhar; Mostoslavsky, Gustavo; Hochedlinger, Konrad; Goren, Alon; Rao, Anjana; Mostoslavsky, Raul

    2015-01-01

    How embryonic stem cells (ESC) commit to specific cell lineages and ultimately yield all cell types of a fully formed organism remains a major question. ESC differentiation is accompanied by large-scale histone and DNA modifications, but the relations between these two categories of epigenetic changes are not understood. Here we demonstrate the hierarchical interplay between the histone deacetylase, sirtuin 6 (Sirt6), which targets acetylated histone H3 at lysines 9 and 56 (H3K9ac and H3K56ac), and the Tet (Ten-eleven translocation) enzymes, which convert 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC). ESCs derived from Sirt6 knockout (S6KO) mice are skewed towards neuroectoderm development. This phenotype is associated with derepression of Oct4, Sox2 and Nanog, which in turn causes an upregulation of Tet enzymes and elevated production of 5hmC. Genome-wide analysis revealed an upregulation of neuroectoderm genes marked with 5hmC in S6KO ESCs, thereby implicating Tet enzymes in the neuroectoderm-skewed differentiation phenotype of S6KO ESCs, which is fully rescued upon knockdown of Tets. We demonstrate a new role for Sirt6 as a chromatin regulator safeguarding the balance between pluripotency and differentiation through Tet-dependent regulation of 5hmC levels. PMID:25915124

  7. The histone deacetylase inhibitor SAHA acts in synergism with fenretinide and doxorubicin to control growth of rhabdoid tumor cells.

    PubMed

    Kerl, Kornelius; Ries, David; Unland, Rebecca; Borchert, Christiane; Moreno, Natalia; Hasselblatt, Martin; Jürgens, Heribert; Kool, Marcel; Görlich, Dennis; Eveslage, Maria; Jung, Manfred; Meisterernst, Michael; Frühwald, Michael

    2013-06-13

    Rhabdoid tumors are highly aggressive malignancies affecting infants and very young children. In many instances these tumors are resistant to conventional type chemotherapy necessitating alternative approaches. Proliferation assays (MTT), apoptosis (propidium iodide/annexin V) and cell cycle analysis (DAPI), RNA expression microarrays and western blots were used to identify synergism of the HDAC (histone deacetylase) inhibitor SAHA with fenretinide, tamoxifen and doxorubicin in rhabdoidtumor cell lines. HDAC1 and HDAC2 are overexpressed in primary rhabdoid tumors and rhabdoid tumor cell lines. Targeting HDACs in rhabdoid tumors induces cell cycle arrest and apoptosis. On the other hand HDAC inhibition induces deregulated gene programs (MYCC-, RB program and the stem cell program) in rhabdoid tumors. These programs are in general associated with cell cycle progression. Targeting these activated pro-proliferative genes by combined approaches of HDAC-inhibitors plus fenretinide, which inhibits cyclinD1, exhibit strong synergistic effects on induction of apoptosis. Furthermore, HDAC inhibition sensitizes rhabdoid tumor cell lines to cell death induced by chemotherapy. Our data demonstrate that HDAC inhibitor treatment in combination with fenretinide or conventional chemotherapy is a promising tool for the treatment of chemoresistant rhabdoid tumors.

  8. Plant-specific Histone Deacetylases HDT½ Regulate GIBBERELLIN 2-OXIDASE 2 Expression to Control Arabidopsis Root Meristem Cell Number.

    PubMed

    Li, Huchen; Torres-Garcia, Jesus; Latrasse, David; Benhamed, Moussa; Schilderink, Stefan; Zhou, Wenkun; Kulikova, Olga; Hirt, Heribert; Bisseling, Ton

    2017-08-30

    Root growth is modulated by environmental factors and depends on cell production in the root meristem (RM). New cells in the meristem are generated by stem cells and transit-amplifying cells, which together determine RM cell number. Transcription factors and chromatin-remodelling factors have been implicated in regulating the switch from stem cells to transit-amplifying cells. Here we show that two Arabidopsis thaliana paralogs encoding plant-specific histone deacetylases, HDT1 and HDT2, regulate a second switch from transit-amplifying cells to expanding cells. Knockdown of HDT½ (hdt1,2i) results in an earlier switch and causes a reduced RM cell number. Our data show that HDT½ negatively regulate the acetylation level of the C19-GIBBERELLIN 2-OXIDASE 2 (GA2ox2) locus and repress the expression of GA2ox2 in the RM and elongation zone. Overexpression of GA2ox2 in the RM phenocopies the hdt1,2i phenotype. Conversely, knockout of GA2ox2 partially rescues the root growth defect of hdt1,2i. These results suggest that by repressing the expression of GA2ox2, HDT½ likely fine-tune gibberellin metabolism and they are crucial for regulating the switch from cell division to expansion to determine RM cell number. We propose that HDT½ function as part of a mechanism that modulates root growth in response to environmental factors. © 2017 American Society of Plant Biologists. All rights reserved.

  9. Arabidopsis seed germination speed is controlled by SNL histone deacetylase-binding factor-mediated regulation of AUX1

    PubMed Central

    Wang, Zhi; Chen, Fengying; Li, Xiaoying; Cao, Hong; Ding, Meng; Zhang, Cun; Zuo, Jinghong; Xu, Chaonan; Xu, Jimei; Deng, Xin; Xiang, Yong; Soppe, Wim J. J.; Liu, Yongxiu

    2016-01-01

    Histone acetylation is known to affect the speed of seed germination, but the molecular regulatory basis of this remains ambiguous. Here we report that loss of function of two histone deacetylase-binding factors, SWI-INDEPENDENT3 (SIN3)-LIKE1 (SNL1) and SNL2, results in accelerated radicle protrusion and growth during seed germination. AUXIN RESISTANT 1 (AUX1) is identified as a key factor in this process, enhancing germination speed downstream of SNL1 and SNL2. AUX1 expression and histone H3 acetylation at lysines 9 and 18 is regulated by SNL1 and SNL2. The D-type cyclins encoding genes CYCD1;1 and CYCD4;1 display increased expression in AUX1 over-expression lines and the snl1snl2 double mutant. Accordingly, knockout of CYCD4;1 reduces seed germination speed of AUX1 over-expression lines and snl1snl2 suggesting the importance of cell cycling for radicle protrusion during seed germination. Together, our work identifies AUX1 as a link between histone acetylation mediated by SNL1 and SNL2, and radicle growth promoted by CYCD1;1 and CYCD4;1 during seed germination. PMID:27834370

  10. Valproic Acid, a Histone Deacetylase Inhibitor, in Combination with Paclitaxel for Anaplastic Thyroid Cancer: Results of a Multicenter Randomized Controlled Phase II/III Trial

    PubMed Central

    Pugliese, Mariateresa; Gallo, Marco; Brignardello, Enrico; Milla, Paola; Orlandi, Fabio; Limone, Paolo Piero; Arvat, Emanuela; Boccuzzi, Giuseppe; Piovesan, Alessandro

    2016-01-01

    Anaplastic thyroid cancer (ATC) has a median survival less than 5 months and, to date, no effective therapy exists. Taxanes have recently been stated as the main drug treatment for ATC, and the histone deacetylase inhibitor valproic acid efficiently potentiates the effects of paclitaxel in vitro. Based on these data, this trial assessed the efficacy and safety of the combination of paclitaxel and valproic acid for the treatment of ATC. This was a randomized, controlled phase II/III trial, performed on 25 ATC patients across 5 centers in northwest Italy. The experimental arm received the combination of paclitaxel (80 mg/m2/weekly) and valproic acid (1,000 mg/day); the control arm received paclitaxel alone. Overall survival and disease progression, evaluated in terms of progression-free survival, were the primary outcomes. The secondary outcome was the pharmacokinetics of paclitaxel. The coadministration of valproic acid did not influence the pharmacokinetics of paclitaxel. Neither median survival nor median time to progression was statistically different in the two arms. Median survival of operated-on patients was significantly better than that of patients who were not operated on. The present trial demonstrates that the addition of valproic acid to paclitaxel has no effect on overall survival and disease progression of ATC patients. This trial is registered with EudraCT 2008-005221-11. PMID:27766105

  11. Histone deacetylases and atherosclerosis.

    PubMed

    Zheng, Xia-xia; Zhou, Tian; Wang, Xin-An; Tong, Xiao-hong; Ding, Jia-wang

    2015-06-01

    Atherosclerosis is the most common pathological process that leads to cardiovascular diseases, a disease of large- and medium-sized arteries that is characterized by a formation of atherosclerotic plaques consisting of necrotic cores, calcified regions, accumulated modified lipids, smooth muscle cells (SMCs), endothelial cells, leukocytes, and foam cells. Recently, the question about how to suppress the occurrence of atherosclerosis and alleviate the progress of cardiovascular disease becomes the hot topic. Accumulating evidence suggests that histone deacetylases(HDACs) play crucial roles in arteriosclerosis. This review summarizes the effect of HDACs and HDAC inhibitors(HDACi) on the progress of atherosclerosis. Copyright © 2015. Published by Elsevier Ireland Ltd.

  12. Histone Deacetylases 6 and 9 and Sirtuin-1 Control Foxp3+ Regulatory T Cell Function Through Shared and Isotype-Specific Mechanisms

    PubMed Central

    Beier, Ulf H.; Wang, Liqing; Han, Rongxiang; Akimova, Tatiana; Liu, Yujie; Hancock, Wayne W.

    2013-01-01

    Therapeutic targeting of histone/protein deacetylase 6 (HDAC6), HDAC9, or the sirtuin-1 (Sirt1) augments the suppressive functions of regulatory T cells (Tregs) that contain the transcription factor Foxp3. However, it is unclear whether distinct mechanisms are involved or whether combined inhibition of these targets would be more beneficial. We compared the suppressive functions of Tregs from wild-type C57BL/6 mice with those from mice with either global (HDAC6−/−, HDAC9−/−, and HDAC6−/−HDAC9−/−), or conditional (fl-Sirt1/CD4-Cre or fl-Sirt1/Foxp3-Cre) HDAC deletion, as well as treatment with isoform-selective HDAC inhibitors. We found that the heat shock response was important for the improvement of Treg suppressive function mediated by HDAC6 inhibition, but not Sirt1 inhibition. Furthermore, although HDAC6, HDAC9, and Sirt1 all deacetylated Foxp3, each protein had diverse effects on transcription factors controlling Foxp3 gene expression. For example, loss of HDAC9 was associated with stabilization of the acetylation of signal transducer and activator of transcription 5 (STAT5) and of its transcriptional activity. Hence, targeting different HDACs increased Treg function by multiple and additive mechanisms, which indicates the therapeutic potential for combinations of HDAC inhibitors in the management of autoimmunity and organ transplantation. PMID:22715468

  13. Histone Deacetylases and Cardiometabolic Diseases

    PubMed Central

    Yiew, Kan Hui; Chatterjee, Tapan K.; Hui, David Y.; Weintraub, Neal L.

    2015-01-01

    Cardiometabolic disease, emerging as a worldwide epidemic, is a combination of metabolic derangements leading to type 2 diabetes and cardiovascular disease. Genetic and environmental factors are linked through epigenetic mechanisms to the pathogenesis of cardiometabolic disease. Post-translational modifications of histone tails, including acetylation and deacetylation, epigenetically alter chromatin structure and dictate cell-specific gene expression patterns. The histone deacetylase (HDAC) family is comprised of 18 members that regulate gene expression by altering the acetylation status of nucleosomal histones and by functioning as nuclear transcriptional co-repressors. HDACs regulate key aspects of metabolism, inflammation, and vascular function pertinent to cardiometabolic disease in a cell- and tissue-specific manner. HDACs also likely play a role in the “metabolic memory” of diabetes, an important clinical aspect of the disease. Understanding the molecular, cellular, and physiological functions of HDACs in cardiometabolic disease is expected to provide insight into disease pathogenesis, risk factor control, and therapeutic development. PMID:26183616

  14. Role of RhoA, mDia, and ROCK in cell shape-dependent control of the Skp2-p27kip1 pathway and the G1/S transition.

    PubMed

    Mammoto, Akiko; Huang, Sui; Moore, Kimberly; Oh, Philmo; Ingber, Donald E

    2004-06-18

    Cell shape-dependent control of cell-cycle progression underlies the spatial differentials of growth that drive tissue morphogenesis, yet little is known about how cell distortion impacts the biochemical signaling machinery that is responsible for growth control. Here we show that the Rho family GTPase, RhoA, conveys the "cell shape signal" to the cell-cycle machinery in human capillary endothelial cells. Cells accumulating p27(kip1) and arrested in mid G(1) phase when spreading were inhibited by restricted extracellular matrix adhesion, whereas constitutively active RhoA increased expression of the F-box protein Skp2 required for ubiquitination-dependent degradation of p27(kip1) and restored G(1) progression in these cells. Studies with dominant-negative and constitutively active forms of mDia1, a downstream effector of RhoA, and with a pharmacological inhibitor of ROCK, another RhoA target, revealed that RhoA promoted G(1) progression by altering the balance of activities between these two downstream effectors. These data indicate that signaling proteins such as mDia1 and ROCK, which are thought to be involved primarily in cytoskeletal remodeling, also mediate cell growth regulation by coupling cell shape to the cell-cycle machinery at the level of signal transduction.

  15. Butyrate Histone Deacetylase Inhibitors

    PubMed Central

    Boosalis, Michael S.; Perrine, Susan P.; Sangerman, José

    2012-01-01

    Abstract In addition to being a part of the metabolic fatty acid fuel cycle, butyrate is also capable of inducing growth arrest in a variety of normal cell types and senescence-like phenotypes in gynecological cancer cells, inhibiting DNA synthesis and cell growth in colonic tumor cell lines, suppressing hTERT mRNA expression and telomerase activity in human prostate cancer cells, and inducing stem cell differentiation and apoptosis by DNA fragmentation. It regulates gene expression by inhibiting histone deacetylases (HDACs), enhances memory recovery and formation in mice, stimulates neurogenesis in the ischemic brain, promotes osteoblast formation, selectively blocks cell replication in transformed cells (compared to healthy cells), and can prevent and treat diet-induced obesity and insulin resistance in mouse models of obesity, as well as stimulate fetal hemoglobin expression in individuals with hematologic diseases such as the thalassemias and sickle-cell disease, in addition to a multitude of other biochemical effects in vivo. However, efforts to exploit the potential of butyrate in the clinical treatment of cancer and other medical disorders are thwarted by its poor pharmacological properties (short half-life and first-pass hepatic clearance) and the multigram doses needed to achieve therapeutic concentrations in vivo. Herein, we review some of the methods used to overcome these difficulties with an emphasis on HDAC inhibition. PMID:23514803

  16. The extracellular constitutive production of chitin deacetylase in Metarhizium anisopliae: possible edge to entomopathogenic fungi in the biological control of insect pests.

    PubMed

    Nahar, Pallavi; Ghormade, Vandana; Deshpande, Mukund V

    2004-02-01

    The possible contribution of extracellular constitutively produced chitin deacetylase by Metarhizium anisopliae in the process of insect pathogenesis has been evaluated. Chitin deacetylase converts chitin, a beta-1,4-linked N-acetylglucosamine polymer, into its deacetylated form chitosan, a glucosamine polymer. When grown in a yeast extract-peptone medium, M. anisopliae constitutively produced the enzymes protease, lipase, and two chitin-metabolizing enzymes, viz. chitin deacetylase (CDA) and chitosanase. Chitinase activity was induced in chitin-containing medium. Staining of 7.5% native polyacrylamide gels at pH 8.9 revealed CDA activity in three bands. SDS-PAGE showed that the apparent molecular masses of the three isoforms were 70, 37, and 26 kDa, respectively. Solubilized melanin (10microg) inhibited chitinase activity, whereas CDA was unaffected. Following germination of M. anisopliae conidia on isolated Helicoverpa armigera, cuticle revealed the presence of chitosan by staining with 3-methyl-2-benzothiazoline hydrazone. Blue patches of chitosan were observed on cuticle, indicating conversion of chitin to chitosan. Hydrolysis of chitin with constitutively produced enzymes of M. anisopliae suggested that CDA along with chitosanase contributed significantly to chitin hydrolysis. Thus, chitin deacetylase was important in initiating pathogenesis of M. anisopliae softening the insect cuticle to aid mycelial penetration. Evaluation of CDA and chitinase activities in other isolates of Metarhizium showed that those strains had low chitinase activity but high CDA activity. Chemical assays of M. anisopliae cell wall composition revealed the presence of chitosan. CDA may have a dual role in modifying the insect cuticular chitin for easy penetration as well as for altering its own cell walls for defense from insect chitinase.

  17. 26 CFR 1.280G-1 - Golden parachute payments.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... control that is described in any of the following sections of the Internal Revenue Code: section 501(c... 26 Internal Revenue 3 2011-04-01 2011-04-01 false Golden parachute payments. 1.280G-1 Section 1.280G-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME...

  18. Regulation of small GTPase activity by G1 cyclins.

    PubMed

    Pedraza, Neus; Cemeli, Tània; Monserrat, Ma Ventura; Garí, Eloi; Ferrezuelo, Francisco

    2017-01-27

    Together with a cyclin-dependent kinase (CDK) partner G1 cyclins control cell cycle entry by phosphorylating a number of nuclear targets and releasing a transcriptional program at the end of G1 phase. Yeast G1 cyclins also operate on cytoplasmic targets involved in the polarization of the cytoskeleton and vesicle trafficking. These processes are mainly controlled by the small GTPase Cdc42, and G1 cyclins regulate the activity of this and other small GTPases through the modulation of their regulators and effectors. This regulation is key for different developmental outcomes in unicellular organisms. In mammalian cells cytoplasmic G1 cyclin D1 has been shown to promote the activity of Rac1 and Ral GTPases and to block RhoA. Regulation of these small GTPases by G1 cyclins may constitute a mechanism to coordinate proliferation with cell migration and morphogenesis, important processes not only during normal development and organogenesis but also for tumor formation and metastasis. Here we briefly review the evidence supporting a role of G1 cyclins and CDKs as regulators of the activity of small GTPases, emphasizing their functional relevance both in budding yeast and in mammalian cells.

  19. Rho-kinase signaling controls nucleocytoplasmic shuttling of class IIa histone deacetylase (HDAC7) and transcriptional activation of orphan nuclear receptor NR4A1.

    PubMed

    Compagnucci, Claudia; Barresi, Sabina; Petrini, Stefania; Bertini, Enrico; Zanni, Ginevra

    2015-04-03

    Rho-kinase (ROCK) has been well documented to play a key role in RhoA-induced actin remodeling. ROCK activation results in myosin light chain (MLC) phosphorylation either by direct action on MLC kinase (MLCK) or by inhibition of MLC phosphatase (MLCP), modulating actin-myosin contraction. We found that inhibition of the ROCK pathway in induced pluripotent stem cells, leads to nuclear export of HDAC7 and transcriptional activation of the orphan nuclear receptor NR4A1 while in cells with constitutive ROCK hyperactivity due to loss of function of the RhoGTPase activating protein Oligophrenin-1 (OPHN1), the orphan nuclear receptor NR4A1 is downregulated. Our study identify a new target of ROCK signaling via myosin phosphatase subunit (MYPT1) and Histone Deacetylase (HDAC7) at the nuclear level and provide new insights in the cellular functions of ROCK. Copyright © 2014 Elsevier Inc. All rights reserved.

  20. Saccharomyces cerevisiae TORC1 Controls Histone Acetylation by Signaling Through the Sit4/PP6 Phosphatase to Regulate Sirtuin Deacetylase Nuclear Accumulation

    PubMed Central

    Workman, Jason J.; Chen, Hongfeng; Laribee, R. Nicholas

    2016-01-01

    The epigenome responds to changes in the extracellular environment, yet how this information is transmitted to the epigenetic regulatory machinery is unclear. Using a Saccharomyces cerevisiae yeast model, we demonstrate that target of rapamycin complex 1 (TORC1) signaling, which is activated by nitrogen metabolism and amino acid availability, promotes site-specific acetylation of histone H3 and H4 N-terminal tails by opposing the activity of the sirtuin deacetylases Hst3 and Hst4. TORC1 does so through suppression of the Tap42-regulated Sit4 (PP6) phosphatase complex, as sit4Δ rescues histone acetylation under TORC1-repressive conditions. We further demonstrate that TORC1 inhibition, and subsequent PP6 activation, causes a selective, rapid, nuclear accumulation of Hst4, which correlates with decreased histone acetylation. This increased Hst4 nuclear localization precedes an elevation in Hst4 protein expression, which is attributed to reduced protein turnover, suggesting that nutrient signaling through TORC1 may limit Hst4 nuclear accumulation to facilitate Hst4 degradation and maintain histone acetylation. This pathway is functionally relevant to TORC1 signaling since the stress sensitivity of a nonessential TORC1 mutant (tco89Δ) to hydroxyurea and arsenic can be reversed by combining tco89Δ with either hst3Δ, hst4Δ, or sit4Δ. Surprisingly, while hst3Δ or hst4Δ rescues the sensitivity tco89Δ has to low concentrations of the TORC1 inhibitor rapamycin, sit4Δ fails to do so. These results suggest Sit4 provides an additional function necessary for TORC1-dependent cell growth and proliferation. Collectively, this study defines a novel mechanism by which TORC1 suppresses a PP6-regulated sirtuin deacetylase pathway to couple nutrient signaling to epigenetic regulation. PMID:27343235

  1. Pharmacogenomics and histone deacetylase inhibitors.

    PubMed

    Goey, Andrew Kl; Sissung, Tristan M; Peer, Cody J; Figg, William D

    2016-11-01

    The histone deacetylase inhibitor valproic acid (VPA) has been used for many decades in neurology and psychiatry. The more recent introduction of the histone deacetylase inhibitors (HDIs) belinostat, romidepsin and vorinostat for treatment of hematological malignancies indicates the increasing popularity of these agents. Belinostat, romidepsin and vorinostat are metabolized or transported by polymorphic enzymes or drug transporters. Thus, genotype-directed dosing could improve pharmacotherapy by reducing the risk of toxicities or preventing suboptimal treatment. This review provides an overview of clinical studies on the effects of polymorphisms on the pharmacokinetics, efficacy or toxicities of HDIs including belinostat, romidepsin, vorinostat, panobinostat, VPA and a number of novel compounds currently being tested in Phase I and II trials. Although pharmacogenomic studies for HDIs are scarce, available data indicate that therapy with belinostat (UGT1A1), romidepsin (ABCB1), vorinostat (UGT2B17) or VPA (UGT1A6) could be optimized by upfront genotyping.

  2. Cell Proliferation and Expression of Cell Cycle Regulatory Proteins that Control the G1/S Transition Are Age Dependent and Lobe Specific in the Brown Norway Rat Model of Prostatic Hyperplasia

    PubMed Central

    Yan, Jinchun; Brown, Terry R.

    2008-01-01

    Age-dependent epithelial cell hyperplasia in the dorsal and lateral lobes of Brown Norway rats is analogous to benign prostatic hyperplasia in aging men. A major question is whether differential lobe-specific and age-dependent proliferation of cells, rather than cell survival, contributes to the hyperplasia. Although serum testosterone (T) levels decline in aged rats, active cell proliferation was detected as Ki67-positive cells in the dorsal and lateral lobes. We determined whether androgens differentially affect cell proliferation and cell-cycle regulatory proteins in the prostate lobes of young and aged rats. Castrated rats were treated with different doses of T to restore serum levels to those of intact young or aged rats. Rates of cell proliferation, measured by 5-bromodeoxyuridine labeling, peaked after 3-d T treatment in all lobes. 5-bromodeoxyuridine-labeling indices were higher in the dorsal and lateral lobes of aged than of young rats with equivalent serum T levels. No age-dependent difference was seen in the ventral lobe. Cell proliferation was marked by increased levels of cyclins D1 and E and cyclin-dependent kinases 4 and 6, decreased p27 and increased phosphorylation of Rb. Levels of cyclins D1 and E were higher in the dorsal and lateral lobes of intact and T-treated aged than young rats. Confocal immunofluorescent microscopy documented changes in cyclin-dependent kinase 4 and cyclin D1 subcellular localization. Cyclin D1 nuclear localization correlated with the time frame for cell proliferation. In conclusion, rates of cell proliferation and levels of cell-cycle regulatory proteins that control the G1/S transition exhibit lobe-specific and age-dependent differences in response to androgens. PMID:17962342

  3. Cell proliferation and expression of cell cycle regulatory proteins that control the G1/S transition are age dependent and lobe specific in the Brown Norway rat model of prostatic hyperplasia.

    PubMed

    Yan, Jinchun; Brown, Terry R

    2008-01-01

    Age-dependent epithelial cell hyperplasia in the dorsal and lateral lobes of Brown Norway rats is analogous to benign prostatic hyperplasia in aging men. A major question is whether differential lobe-specific and age-dependent proliferation of cells, rather than cell survival, contributes to the hyperplasia. Although serum testosterone (T) levels decline in aged rats, active cell proliferation was detected as Ki67-positive cells in the dorsal and lateral lobes. We determined whether androgens differentially affect cell proliferation and cell-cycle regulatory proteins in the prostate lobes of young and aged rats. Castrated rats were treated with different doses of T to restore serum levels to those of intact young or aged rats. Rates of cell proliferation, measured by 5-bromodeoxyuridine labeling, peaked after 3-d T treatment in all lobes. 5-bromodeoxyuridine-labeling indices were higher in the dorsal and lateral lobes of aged than of young rats with equivalent serum T levels. No age-dependent difference was seen in the ventral lobe. Cell proliferation was marked by increased levels of cyclins D1 and E and cyclin-dependent kinases 4 and 6, decreased p27 and increased phosphorylation of Rb. Levels of cyclins D1 and E were higher in the dorsal and lateral lobes of intact and T-treated aged than young rats. Confocal immunofluorescent microscopy documented changes in cyclin-dependent kinase 4 and cyclin D1 subcellular localization. Cyclin D1 nuclear localization correlated with the time frame for cell proliferation. In conclusion, rates of cell proliferation and levels of cell-cycle regulatory proteins that control the G1/S transition exhibit lobe-specific and age-dependent differences in response to androgens.

  4. Knockdown of selenocysteine-specific elongation factor in Amblyomma maculatum alters the pathogen burden of Rickettsia parkeri with epigenetic control by the Sin3 histone deacetylase corepressor complex.

    PubMed

    Adamson, Steven W; Browning, Rebecca E; Budachetri, Khemraj; Ribeiro, José M C; Karim, Shahid

    2013-01-01

    Selenocysteine is the 21st naturally-occurring amino acid. Selenoproteins have diverse functions and many remain uncharacterized, but they are typically associated with antioxidant activity. The incorporation of selenocysteine into the nascent polypeptide chain recodes the TGA stop codon and this process depends upon a number of essential factors including the selenocysteine elongation factor (SEF). The transcriptional expression of SEF did not change significantly in tick midguts throughout the blood meal, but decreased in salivary glands to 20% at the end of the fast feeding phase. Since selenoprotein translation requires this specialized elongation factor, we targeted this gene for knockdown by RNAi to gain a global view of the role selenoproteins play in tick physiology. We found no significant differences in tick engorgement and embryogenesis but detected no antioxidant capacity in tick saliva. The transcriptional profile of selenoproteins in R. parkeri-infected Amblyomma maculatum revealed declined activity of selenoprotein M and catalase and increased activity of selenoprotein O, selenoprotein S, and selenoprotein T. Furthermore, the pathogen burden was significantly altered in SEF-knockdowns. We then determined the global impact of SEF-knockdown by RNA-seq, and mapped huge shifts in secretory gene expression that could be the result of downregulation of the Sin3 histone deacetylase corepressor complex.

  5. Knockdown of Selenocysteine-Specific Elongation Factor in Amblyomma maculatum Alters the Pathogen Burden of Rickettsia parkeri with Epigenetic Control by the Sin3 Histone Deacetylase Corepressor Complex

    PubMed Central

    Adamson, Steven W.; Browning, Rebecca E.; Budachetri, Khemraj; Ribeiro, José M. C.; Karim, Shahid

    2013-01-01

    Selenocysteine is the 21st naturally-occurring amino acid. Selenoproteins have diverse functions and many remain uncharacterized, but they are typically associated with antioxidant activity. The incorporation of selenocysteine into the nascent polypeptide chain recodes the TGA stop codon and this process depends upon a number of essential factors including the selenocysteine elongation factor (SEF). The transcriptional expression of SEF did not change significantly in tick midguts throughout the blood meal, but decreased in salivary glands to 20% at the end of the fast feeding phase. Since selenoprotein translation requires this specialized elongation factor, we targeted this gene for knockdown by RNAi to gain a global view of the role selenoproteins play in tick physiology. We found no significant differences in tick engorgement and embryogenesis but detected no antioxidant capacity in tick saliva. The transcriptional profile of selenoproteins in R. parkeri-infected Amblyomma maculatum revealed declined activity of selenoprotein M and catalase and increased activity of selenoprotein O, selenoprotein S, and selenoprotein T. Furthermore, the pathogen burden was significantly altered in SEF-knockdowns. We then determined the global impact of SEF-knockdown by RNA-seq, and mapped huge shifts in secretory gene expression that could be the result of downregulation of the Sin3 histone deacetylase corepressor complex. PMID:24282621

  6. Rho-kinase signaling controls nucleocytoplasmic shuttling of class IIa Histone Deacetylase (HDAC7) and transcriptional activation of orphan nuclear receptor NR4A1

    SciTech Connect

    Compagnucci, Claudia; Barresi, Sabina; Petrini, Stefania; Bertini, Enrico; Zanni, Ginevra

    2015-04-03

    Rho-kinase (ROCK) has been well documented to play a key role in RhoA-induced actin remodeling. ROCK activation results in myosin light chain (MLC) phosphorylation either by direct action on MLC kinase (MLCK) or by inhibition of MLC phosphatase (MLCP), modulating actin–myosin contraction. We found that inhibition of the ROCK pathway in induced pluripotent stem cells, leads to nuclear export of HDAC7 and transcriptional activation of the orphan nuclear receptor NR4A1 while in cells with constitutive ROCK hyperactivity due to loss of function of the RhoGTPase activating protein Oligophrenin-1 (OPHN1), the orphan nuclear receptor NR4A1 is downregulated. Our study identify a new target of ROCK signaling via myosin phosphatase subunit (MYPT1) and Histone Deacetylase (HDAC7) at the nuclear level and provide new insights in the cellular functions of ROCK. - Highlights: • ROCK regulates nucleocytoplasmic shuttling of HDAC7 via phosphorylation of MYPT1. • Nuclear export of HDAC7 and upregulation of NR4A1 occurs with low ROCK activity. • High levels of ROCK activity due to OPHN1 loss of function downregulate NR4A1.

  7. Chemical Phylogenetics of Histone Deacetylases

    PubMed Central

    Bradner, James E.; West, Nathan; Grachan, Melissa L.; Greenberg, Edward F.; Haggarty, Stephen J.; Warnow, Tandy; Mazitschek, Ralph

    2010-01-01

    The broad study of histone deacetylases in chemistry, biology and medicine relies on tool compounds to derive mechanistic insights. A phylogenetic analysis of Class I and II HDACs as targets of a comprehensive, structurally diverse panel of inhibitors revealed unexpected isoform selectivity even among compounds widely perceived as non-selective. The synthesis and study of a focused library of cinnamic hydroxamates allowed the identification of a first non-selective HDAC inhibitor. These data will guide a more informed use of HDAC inhibitors as chemical probes and therapeutic agents. PMID:20139990

  8. Properties of the yeast nuclear histone deacetylase.

    PubMed Central

    Sanchez del Pino, M M; Lopez-Rodas, G; Sendra, R; Tordera, V

    1994-01-01

    A nuclear histone deacetylase from yeast was partially purified and some of its characteristics were studied. Histone deacetylase activity was stimulated in vitro by high-mobility-group nonhistone chromatin proteins 1 and 2 and ubiquitin and inhibited by spermine and spermidine, whereas n-butyrate had no significant inhibitory effect. Like the mammalian enzyme, partially purified histone deacetylase from yeast was strongly inhibited by trichostatin A. However, in crude extract preparations the yeast enzyme was not inhibited and treatment with trichostatin in vivo did not show any effect, either on the histone acetylation level or on cell viability. At low ionic strength, the enzyme can be isolated as a complex of high molecular mass that is much less inhibited by trichostatin A than is partially purified histone deacetylase activity. Furthermore, radiolabelled oligonucleosomes were more efficiently deacetylated by the complex than by the low-molecular-mass form of the enzyme. The histone deacetylase activity was separated from a polyamine deacetylase activity and its specificity studied. Using h.p.l.c.-purified core histone species as substrate, histone deacetylase from yeast is able to deacetylate all core histones with a slight preference for H3. Our results support the idea that the yeast histone deacetylase may act as a high-molecular-mass complex in vivo. Images Figure 3 PMID:7980438

  9. Strategic Cell-Cycle Regulatory Features That Provide Mammalian Cells with Tunable G1 Length and Reversible G1 Arrest

    PubMed Central

    Pfeuty, Benjamin

    2012-01-01

    Transitions between consecutive phases of the eukaryotic cell cycle are driven by the catalytic activity of selected sets of cyclin-dependent kinases (Cdks). Yet, their occurrence and precise timing is tightly scheduled by a variety of means including Cdk association with inhibitory/adaptor proteins (CKIs). Here we focus on the regulation of G1-phase duration by the end of which cells of multicelled organisms must decide whether to enter S phase or halt, and eventually then, differentiate, senesce or die to obey the homeostatic rules of their host. In mammalian cells, entry in and progression through G1 phase involve sequential phosphorylation and inactivation of the retinoblastoma Rb proteins, first, by cyclin D-Cdk4,6 with the help of CKIs of the Cip/Kip family and, next, by the cyclin E-Cdk2 complexes that are negatively regulated by Cip/Kip proteins. Using a dynamical modeling approach, we show that the very way how the Rb and Cip/Kip regulatory modules interact differentially with cyclin D-Cdk4,6 and cyclin E-Cdk2 provides to mammalian cells a powerful means to achieve an exquisitely-sensitive control of G1-phase duration and fully reversible G1 arrests. Consistently, corruption of either one of these two modules precludes G1 phase elongation and is able to convert G1 arrests from reversible to irreversible. This study unveils fundamental design principles of mammalian G1-phase regulation that are likely to confer to mammalian cells the ability to faithfully control the occurrence and timing of their division process in various conditions. PMID:22558136

  10. Histone deacetylase inhibitor MS-275 augments expression of a subset of IFN-γ-regulated genes in Toxoplasma gondii-infected macrophages but does not improve parasite control.

    PubMed

    Sumpf, Kristina; Nast, Roswitha; Downie, Bryan; Salinas, Gabriela; Lüder, Carsten G K

    2017-09-01

    Toxoplasma gondii is a ubiquitous apicomplexan parasite of mammals and birds and an important pathogen of humans. IFN-γ is the major mediator of host resistance against T. gondii but intriguingly, parasite-infected host cells including macrophages are severely impaired to respond to IFN-γ due to defective transcriptional activation of target genes. Here, we tested the possibility that the impaired responsiveness of T. gondii-infected macrophages to IFN-γ can be restored by inhibiting histone deacetylases (HDACs) using the class I-specific inhibitor MS-275. Treatment of RAW264.7 cells with MS-275 indeed increased MHC class II surface expression in infected and non-infected cells and largely abolished the inhibition of IFN-γ-regulated MHC class II expression exerted by T. gondii. Genome-wide transcriptome profiling revealed that MS-275 increased mean mRNA levels of IFN-γ-regulated genes particularly in non-infected macrophages. Transcript levels of 33% of IFN-γ secondary response genes but only those of a few primary response genes were also increased by MS-275 in T. gondii-infected cells. Importantly, the unresponsiveness of parasite-infected cells to IFN-γ was however not abolished by MS-275. Furthermore, MS-275 also up-regulated several anti-inflammatory cytokines or signaling molecules in T. gondii-infected macrophages. It additionally regulated expression of more than 2500 genes in non-infected macrophages expression of which was surprisingly counteracted by prior infection with T. gondii. FACS analysis and immunofluorescence microscopy revealed that MS-275 did not considerably diminish the number of parasite-positive cells or the intracellular replication in macrophages stimulated or not with IFN-γ. Thus, a supportive therapy using MS-275 appears inappropriate for treatment of toxoplasmosis. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Comparison of early development in utero of cloned fetuses derived from bovine fetal fibroblasts at the G1 and G0/G1 phases.

    PubMed

    Ideta, Atsushi; Hayama, Koh; Urakawa, Manami; Tsuchiya, Kanami; Aoyagi, Yoshito; Saeki, Kazuhiro

    2010-06-01

    In bovine somatic cell nuclear transfer (NT), embryos are more likely to develop to full term when they are derived from fibroblasts at the G1 phase instead of cells at the G0/G1 phase. To better understand the reason for this difference, we examined morphological development in the early pregnancy of NT embryos using G1 phase cells (G1-NT embryos) and G0/G1 phase cells (G0/G1-NT embryos). Blastocysts derived from G1 and G0/G1-NT embryos were transferred to recipient heifers, and the conceptuses at day 50 of gestation were retrieved non-surgically using prostaglandin F(2alpha) and oxytocin. In vitro-fertilized (IVF), parthenogenetic and artificially inseminated (AI) embryos were used as controls. The percentages of embryos that developed to the blastocyst stage did not differ between G1 and G0/G1-NT embryos. Pregnancy rates at day 30 of recipient heifers carrying G1-NT, G0/G1-NT, IVF, parthenogenetic and AI embryos were similar (57-100%). Two recipient heifers carrying parthenogenetic embryos returned to estrus between days 30 and 50 of gestation, whereas all other pregnancies remained viable. Most fetuses at day 50 of gestation of all experimental groups (83%) were recovered non-surgically by several PGF(2alpha) and oxytocin treatments. Recovery rates of normal fetuses derived from G1-NT embryos (83%), IVF embryos (80%) and AI embryos (88%) were greater than those of G0/G1-NT embryos (33%) and parthenogenetic embryos (0%). Our results suggest that NT embryos reconstructed with cells at the G1 phase have a high developmental competence from the time of embryo transfer to day 50 of gestation. Copyright (c) 2010 Elsevier B.V. All rights reserved.

  12. Epigenetic targeting of histone deacetylase: therapeutic potential in Parkinson's disease?

    PubMed

    Harrison, Ian F; Dexter, David T

    2013-10-01

    Parkinson's disease (PD) is the most common movement disorder affecting more than 4million people worldwide. The primary motor symptoms of the disease are due to degeneration of dopaminergic nigrostriatal neurons. Dopamine replacement therapies have therefore revolutionised disease management by partially controlling these symptoms. However these drugs can produce debilitating side effects when used long term and do not protect degenerating neurons against death. Recent evidence has highlighted a pathological imbalance in PD between the acetylation and deacetylation of the histone proteins around which deoxyribonucleic acid (DNA) is coiled, in favour of excessive histone deacetylation. This mechanism of adding/removing acetyl groups to histone lysine residues is one of many epigenetic regulatory processes which control the expression of genes, many of which will be essential for neuronal survival. Hence, such epigenetic modifications may have a pathogenic role in PD. It has therefore been hypothesised that if this pathological imbalance can be corrected with the use of histone deacetylase inhibiting agents then neurodegeneration observed in PD can be ameliorated. This article will review the current literature with regard to epigenetic changes in PD and the use of histone deacetylase inhibitors (HDACIs) in PD: examining the evidence of the neuroprotective effects of numerous HDACIs in cellular and animal models of Parkinsonian cell death. Ultimately answering the question: does epigenetic targeting of histone deacetylases hold therapeutic potential in PD?

  13. Induction of a G1-S checkpoint in fission yeast.

    PubMed

    Bøe, Cathrine A; Krohn, Marit; Rødland, Gro Elise; Capiaghi, Christoph; Maillard, Olivier; Thoma, Fritz; Boye, Erik; Grallert, Beáta

    2012-06-19

    Entry into S phase is carefully regulated and, in most organisms, under the control of a G(1)-S checkpoint. We have previously described a G(1)-S checkpoint in fission yeast that delays formation of the prereplicative complex at chromosomal replication origins after exposure to UV light (UVC). This checkpoint absolutely depends on the Gcn2 kinase. Here, we explore the signal for activation of the Gcn2-dependent G(1)-S checkpoint in fission yeast. If some form of DNA damage can activate the checkpoint, deficient DNA repair should affect the length of the checkpoint-induced delay. We find that the cell-cycle delay differs in repair-deficient mutants from that in wild-type cells. However, the duration of the delay depends not only on the repair capacity of the cells, but also on the nature of the repair deficiency. First, the delay is abolished in cells that are deficient in the early steps of repair. Second, the delay is prolonged in repair mutants that fail to complete repair after the incision stage. We conclude that the G(1)-S delay depends on damage to the DNA and that the activating signal derives not from the initial DNA damage, but from a repair intermediate(s). Surprisingly, we find that activation of Gcn2 does not depend on the processing of DNA damage and that activated Gcn2 alone is not sufficient to delay entry into S phase in UVC-irradiated cells. Thus, the G(1)-S delay depends on at least two different inputs.

  14. Charge variants in IgG1

    PubMed Central

    Goswami, Sirj; Hutchinson, Ryan; Kwong, Zephania W; Yang, Jihong; Wang, Xiangdan; Yao, Zhenling; Sreedhara, Alavattam; Cano, Tony; Tesar, Devin; Nijem, Ihsan; Allison, David E; Wong, Pin Yee; Kao, Yung-Hsiang; Quan, Cynthia; Joshi, Amita; Harris, Reed J; Motchnik, Paul

    2010-01-01

    Antibody charge variants have gained considerable attention in the biotechnology industry due to their potential influence on stability and biological activity. Subtle differences in the relative proportions of charge variants are often observed during routine biomanufacture or process changes and pose a challenge to demonstrating product comparability. To gain further insights into the impact on biological activity and pharmacokinetics (PK) of monoclonal antibody (mAb) charge heterogeneity, we isolated the major charge forms of a recombinant humanized IgG1 and compared their in vitro properties and in vivo PK. The mAb starting material had a pI range of 8.7–9.1 and was composed of about 20% acidic variants, 12% basic variants and 68% main peak. Cation exchange displacement chromatography was used to isolate the acidic, basic and main peak fractions for animal studies. Detailed analyses were performed on the isolated fractions to identify specific chemical modification contributing to the charge differences and were also characterized for purity and in vitro potency prior to being administered either subcutaneously (SC) or intravenously (IV) in rats. All isolated materials had similar potency and rat FcRn binding relative to the starting material. Following IV or SC administration (10 mg/kg) in rats, no difference in serum PK was observed, indicating that physiochemical modifications and pI differences among charge variants were not sufficient to result in PK changes. Thus, these results provided meaningful information for the comparative evaluation of charge-related heterogeneity of mAbs and suggested that charge variants of IgGs do not affect the in vitro potency, FcRn binding affinity or the PK properties in rats. PMID:20818176

  15. Nonpeptide Macrocyclic Histone Deacetylase Inhibitors

    PubMed Central

    Oyelere, Adegboyega K.; Chen, Po C.; Guerrant, William; Mwakwari, Sandra C.; Hood, Rebecca; Zhang, Yunzhe; Fan, Yuhong

    2009-01-01

    Inhibition of Histone Deacetylases inhibitors (HDACi) hold great promise in cancer therapy due to their demonstrated ability to arrest proliferation of nearly all transformed cell types. Of the several structurally distinct small molecules HDACi reported, macrocyclic depsipeptides have the most complex recognition cap-group moieties and present an excellent opportunity for the modulation of the biological activities of HDACi. Unfortunately, the structure–activity relationship (SAR) studies for this class of compounds have been impaired largely because most macrocyclic HDACi known to date are comprised of complex peptide macrocycles. In addition to retaining the pharmacologically disadvantaged peptidyl-backbone, they offer only limited opportunity for side-chain modifications. Here we report the discovery of a new class of macrocyclic HDACi based on the macrolide antibiotics skeletons. SAR studies revealed that these compounds displayed both linker-length and macrolide-type dependent HDAC inhibition activities with IC50 in low nanomolar range. In addition, these nonpeptide macrocyclic HDACi are more selective against HDAC 1 and 2 relative to HDAC 8, another class I HDAC isoform, hence have sub-class HDAC isoform selectivity. PMID:19093884

  16. Resetting a functional G1 nucleus after mitosis.

    PubMed

    de Castro, Ines J; Gokhan, Ezgi; Vagnarelli, Paola

    2016-09-01

    The maintenance of the correct cellular information goes beyond the simple transmission of an intact genetic code from one generation to the next. Epigenetic changes, topological cues and correct protein-protein interactions need to be re-established after each cell division to allow the next cell cycle to resume in the correct regulated manner. This process begins with mitotic exit and re-sets all the changes that occurred during mitosis thus restoring a functional G1 nucleus in preparation for the next cell cycle. Mitotic exit is triggered by inactivation of mitotic kinases and the reversal of their phosphorylation activities on many cellular components, from nuclear lamina to transcription factors and chromatin itself. To reverse all these phosphorylations, phosphatases act during mitotic exit in a timely and spatially controlled manner directing the events that lead to a functional G1 nucleus. In this review, we will summarise the recent developments on the control of phosphatases and their known substrates during mitotic exit, and the key steps that control the restoration of chromatin status, nuclear envelope reassembly and nuclear body re-organisation. Although pivotal work has been conducted in this area in yeast, due to differences between the mitotic exit network between yeast and vertebrates, we will mainly concentrate on the vertebrate system.

  17. Cloning and expression of two chitin deacetylase genes of Saccharomyces cerevisiae.

    PubMed

    Mishra, C; Semino, C E; McCreath, K J; de la Vega, H; Jones, B J; Specht, C A; Robbins, P W

    1997-03-30

    Chitin deacetylase (EC 3.5.1.41), which hydrolyses the N-acetamido groups of N-acetyl-D-glucosamine residues in chitin, has been demonstrated in crude extracts from sporulating Saccharomyces cerevisiae. Two S. cerevisiae open reading frames (ORFs), identified by the Yeast Genome Project, have protein sequence homology to a chitin deacetylase from Mucor rouxii. Northern blot hybridizations show each ORF was transcribed in diploid cells after transfer to sporulation medium and prior to formation of asci. Each ORF was cloned in a vector under transcriptional control of the GAL 1, 10 promoter and introduced back into haploid strains of S. cerevisiae. Chitin deacetylase activity was detected by in vitro assays from vegetative cells grown in galactose. Chemical analysis of these cells also demonstrated the synthesis of chitosam in vivo. Both recombinant chitin deacetylases showed similar qualitative and quantitative activities toward chitooligosaccharides in vitro. A diploid strain deleted to both ORFs, when sporulated, did not show deacetylase activity. The mutant spores were hypersensitive to lytic enzymes (Glusulase or Zymolyase).

  18. Histone deacetylases in kidney development: implications for disease and therapy.

    PubMed

    Chen, Shaowei; El-Dahr, Samir S

    2013-05-01

    Histone deacetylases (HDACs) are an evolutionarily conserved group of enzymes that regulate a broad range of biological processes through removal of acetyl groups from histones as well as non-histone proteins. Recent studies using a variety of pharmacological inhibitors and genetic models of HDACs have revealed a central role of HDACs in control of kidney development. These findings provide new insights into the epigenetic mechanisms underlying congenital anomalies of the kidney and urinary tract (CAKUT) and implicate the potential of HDACs as therapeutic targets in kidney diseases, such as cystic kidney diseases and renal cell cancers. Determining the specific functions of individual HDAC members would be an important task of future research.

  19. Erasers of Histone Acetylation: The Histone Deacetylase Enzymes

    PubMed Central

    Seto, Edward; Yoshida, Minoru

    2014-01-01

    Histone deacetylases (HDACs) are enzymes that catalyze the removal of acetyl functional groups from the lysine residues of both histone and nonhistone proteins. In humans, there are 18 HDAC enzymes that use either zinc- or NAD+-dependent mechanisms to deacetylate acetyl lysine substrates. Although removal of histone acetyl epigenetic modification by HDACs regulates chromatin structure and transcription, deacetylation of nonhistones controls diverse cellular processes. HDAC inhibitors are already known potential anticancer agents and show promise for the treatment of many diseases. PMID:24691964

  20. 26 CFR 1.149(g)-1 - Hedge bonds.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 26 Internal Revenue 2 2012-04-01 2012-04-01 false Hedge bonds. 1.149(g)-1 Section 1.149(g)-1...) INCOME TAXES (CONTINUED) Tax Exemption Requirements for State and Local Bonds § 1.149(g)-1 Hedge bonds... of replacement proceeds (other than amounts in a bona fide debt service fund or a reasonably...

  1. 26 CFR 1.149(g)-1 - Hedge bonds.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 2 2011-04-01 2011-04-01 false Hedge bonds. 1.149(g)-1 Section 1.149(g)-1...) INCOME TAXES (CONTINUED) Tax Exemption Requirements for State and Local Bonds § 1.149(g)-1 Hedge bonds... of replacement proceeds (other than amounts in a bona fide debt service fund or a reasonably...

  2. 26 CFR 1.149(g)-1 - Hedge bonds.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 26 Internal Revenue 2 2014-04-01 2014-04-01 false Hedge bonds. 1.149(g)-1 Section 1.149(g)-1...) INCOME TAXES (CONTINUED) Tax Exemption Requirements for State and Local Bonds § 1.149(g)-1 Hedge bonds... of replacement proceeds (other than amounts in a bona fide debt service fund or a reasonably...

  3. 26 CFR 1.149(g)-1 - Hedge bonds.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 26 Internal Revenue 2 2013-04-01 2013-04-01 false Hedge bonds. 1.149(g)-1 Section 1.149(g)-1...) INCOME TAXES (CONTINUED) Tax Exemption Requirements for State and Local Bonds § 1.149(g)-1 Hedge bonds... of replacement proceeds (other than amounts in a bona fide debt service fund or a reasonably...

  4. 26 CFR 1.167(g)-1 - Basis for depreciation.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 2 2010-04-01 2010-04-01 false Basis for depreciation. 1.167(g)-1 Section 1.167(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES (CONTINUED) Itemized Deductions for Individuals and Corporations § 1.167(g)-1 Basis...

  5. 26 CFR 301.6503(g)-1 - Suspension pending correction.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 18 2010-04-01 2010-04-01 false Suspension pending correction. 301.6503(g)-1 Section 301.6503(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED... Collection § 301.6503(g)-1 Suspension pending correction. The running of the periods of limitations provided...

  6. 26 CFR 1.149(g)-1 - Hedge bonds.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 2 2010-04-01 2010-04-01 false Hedge bonds. 1.149(g)-1 Section 1.149(g)-1...) INCOME TAXES (CONTINUED) Tax Exemption Requirements for State and Local Bonds § 1.149(g)-1 Hedge bonds... for purposes of section 149(g) and this section. In addition, the following terms have the following...

  7. 26 CFR 31.3402(g)-1 - Supplemental wage payments.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 15 2010-04-01 2010-04-01 false Supplemental wage payments. 31.3402(g)-1 Section 31.3402(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED... SOURCE Collection of Income Tax at Source § 31.3402(g)-1 Supplemental wage payments. (a) In general...

  8. Caspase-mediated specific cleavage of human histone deacetylase 4.

    PubMed

    Liu, Fang; Dowling, Melissa; Yang, Xiang-Jiao; Kao, Gary D

    2004-08-13

    Histone deacetylase 4 (HDAC4) is a class II HDAC implicated in controlling gene expression important for diverse cellular functions, but little is known about how its expression and stability are regulated. We report here that this deacetylase is unusually unstable, with a half-life of less than 8 h. Consistent with the instability of HDAC4 protein, its mRNA was also highly unstable (with a half-life of less than 4 h). The degradation of HDAC4 could be accelerated by exposure of cells to ultraviolet irradiation. HDAC4 degradation was not dependent on proteasome or CRM1-mediated export activity but instead was caspase-dependent and was detectable in diverse human cancer lines. Of two potential caspase consensus motifs in HDAC4, both lying within a region containing proline-, glutamic acid-, serine-, and threonine-rich (PEST) sequences, we identified, by site-directed mutagenesis, Asp-289 as the prime cleavage site. Notably, this residue is not conserved among other class IIa members, HDAC5, -7, and -9. Finally, the induced expression of caspase-cleavable HDAC4 led to markedly increased apoptosis. These results therefore unexpectedly link the regulation of HDAC4 protein stability to caspases, enzymes that are important for controlling cell death and differentiation.

  9. Most of the G1 period in hamster cells is eliminated by lengthening the S period.

    PubMed Central

    Stancel, G M; Prescott, D M; Liskay, R M

    1981-01-01

    Two Chinese hamster cell lines, G1+-1 and CHO, have been grown in the presence of low concentrations of hydroxyurea to determine how a slowing DNA synthesis (i.e., a lengthening of the S period) affects the length of the G1 period. Hydroxyurea concentrations of approximately 10 microM do not alter the generation times of these cell lines but do cause increases in S with corresponding decreases in G1. In both cell lines, 10 microM hydroxyurea reduces G1 to an absolute value of 1 hr, which represents decreases of 70% (G1+-1) and 60% (CHO) from control values. Higher concentrations of hydroxyurea increase the generation times and lengths of S for both cell lines but do not reduce G1 below the minimum value of 1 hr. These observations indicate that the majority of G1 is expendable and most of G1 therefore cannot contain specific events required for the initiation of DNA synthesis. This result supports the hypothesis that G1 is a portion of the cell growth cycle but not of the chromosome cycle. PMID:6947230

  10. 26 CFR 1.475(g)-1 - Effective dates.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 6 2010-04-01 2010-04-01 false Effective dates. 1.475(g)-1 Section 1.475(g)-1...) INCOME TAXES Inventories § 1.475(g)-1 Effective dates. (a)-(b) (c) Section 1.475(a)-3 (concerning... effective date of § 1.1275-6). (g) (h) Section 1.475(b)-4 (concerning transitional issues relating...

  11. Targeting Histone Deacetylases in Diseases: Where Are We?

    PubMed Central

    Benedetti, Rosaria; Conte, Mariarosaria

    2015-01-01

    Abstract Significance: Epigenetic inactivation of pivotal genes involved in cell growth is a hallmark of human pathologies, in particular cancer. Histone acetylation balance obtained through opposing actions of histone deacetylases (HDACs) and histone acetyltransferases is one epigenetic mechanism controlling gene expression and is, thus, associated with disease etiology and progression. Interfering pharmacologically with HDAC activity can correct abnormalities in cell proliferation, migration, vascularization, and death. Recent Advances: Histone deacetylase inhibitors (HDACi) represent a new class of cytostatic agents that interfere with the function of HDACs and are able to increase gene expression by indirectly inducing histone acetylation. Several HDACi, alone or in combination with DNA-demethylating agents, chemopreventive, or classical chemotherapeutic drugs, are currently being used in clinical trials for solid and hematological malignancies, and are, thus, promising candidates for cancer therapy. Critical Issues: (i) Non-specific (off-target) HDACi effects due to activities unassociated with HDAC inhibition. (ii) Advantages/disadvantages of non-selective or isoform-directed HDACi. (iii) Limited number of response-predictive biomarkers. (iv) Toxicity leading to dysfunction of critical biological processes. Future Directions: Selective HDACi could achieve enhanced clinical utility by reducing or eliminating the serious side effects associated with current first-generation non-selective HDACi. Isoform-selective and pan-HDACi candidates might benefit from the identification of biomarkers, enabling better patient stratification and prediction of response to treatment. Antioxid. Redox Signal. 23, 99–126. PMID:24382114

  12. Structure of Prokaryotic Polyamine Deacetylase Reveals Evolutionary Functional Relationships with Eukaryotic Histone Deacetylases

    PubMed Central

    Lombardi, Patrick M.; Angell, Heather D.; Whittington, Douglas A.; Flynn, Erin F.; Rajashankar, Kanagalaghatta R.; Christianson, David W.

    2011-01-01

    Polyamines are a ubiquitous class of polycationic small molecules that can influence gene expression by binding to nucleic acids. Reversible polyamine acetylation regulates nucleic acid binding and is required for normal cell cycle progression and proliferation. Here, we report the structures of Mycoplana ramosa acetylpolyamine amidohydrolase (APAH) complexed with a transition state analogue and a hydroxamate inhibitor, and an inactive mutant complexed with two acetylpolyamine substrates. The structure of APAH is the first of a histone deacetylase-like oligomer and reveals that an 18-residue insert in the L2 loop promotes dimerization and the formation of an 18-Å long “L”-shaped active site tunnel at the dimer interface, accessible only to narrow and flexible substrates. The importance of dimerization for polyamine deacetylase function leads to the suggestion that a comparable dimeric or double-domain histone deacetylase could catalyze polyamine deacetylation reactions in eukaryotes. PMID:21268586

  13. Structure of Prokaryotic Polyamine Deacetylase Reveals Evolutionary Functional Relationships with Eukaryotic Histone Deacetylases

    SciTech Connect

    P Lombardi; H Angell; D Whittington; E Flynn; K Rajashankar; D Christianson

    2011-12-31

    Polyamines are a ubiquitous class of polycationic small molecules that can influence gene expression by binding to nucleic acids. Reversible polyamine acetylation regulates nucleic acid binding and is required for normal cell cycle progression and proliferation. Here, we report the structures of Mycoplana ramosa acetylpolyamine amidohydrolase (APAH) complexed with a transition state analogue and a hydroxamate inhibitor and an inactive mutant complexed with two acetylpolyamine substrates. The structure of APAH is the first of a histone deacetylase-like oligomer and reveals that an 18-residue insert in the L2 loop promotes dimerization and the formation of an 18 {angstrom} long 'L'-shaped active site tunnel at the dimer interface, accessible only to narrow and flexible substrates. The importance of dimerization for polyamine deacetylase function leads to the suggestion that a comparable dimeric or double-domain histone deacetylase could catalyze polyamine deacetylation reactions in eukaryotes.

  14. Inhibitors of Histone Deacetylases for Radiosensitization of Prostate Cancer

    DTIC Science & Technology

    2005-02-01

    AD Award Number: W81XWH-04-1-0170 TITLE: Inhibitors of Histone Deacetylases for Radiosensitization of Prostate Cancer PRINCIPAL INVESTIGATOR: Mira 0...of Histone Deacetylases for Radiosensitization W81XWH-04-1-0170 of Prostate Cancer 6. AUTHOR(S) Mira 0. Jung, Ph.D. 7. PERFORMING ORGANIZA TION NAME(S...cellular radiation sensitivity. 14. SUBJECT TERMS 15. NUMBER OF PA GES Radiation sensitivity, histone deacetylase , cytotoxicity 13 16. PRICE CODE 17

  15. 26 CFR 31.3121(g)-1 - Agricultural labor.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 15 2010-04-01 2010-04-01 false Agricultural labor. 31.3121(g)-1 Section 31....3121(g)-1 Agricultural labor. (a) In general. (1) The term “agricultural labor” as defined in section... agricultural labor and to the test for determining whether cash remuneration paid for agricultural...

  16. 26 CFR 31.3121(g)-1 - Agricultural labor.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 15 2011-04-01 2011-04-01 false Agricultural labor. 31.3121(g)-1 Section 31....3121(g)-1 Agricultural labor. (a) In general. (1) The term “agricultural labor” as defined in section... agricultural labor and to the test for determining whether cash remuneration paid for agricultural...

  17. 26 CFR 301.6503(g)-1 - Suspension pending correction.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 18 2011-04-01 2011-04-01 false Suspension pending correction. 301.6503(g)-1... Collection § 301.6503(g)-1 Suspension pending correction. The running of the periods of limitations provided... making correction under section 4963(e)(1)(B)....

  18. 26 CFR 1.280G-1 - Golden parachute payments.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 26 Internal Revenue 3 2014-04-01 2014-04-01 false Golden parachute payments. 1.280G-1 Section 1... (CONTINUED) INCOME TAXES (CONTINUED) Items Not Deductible § 1.280G-1 Golden parachute payments. The following questions and answers relate to the treatment of golden parachute payments under section 280G of the...

  19. 26 CFR 1.280G-1 - Golden parachute payments.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 26 Internal Revenue 3 2012-04-01 2012-04-01 false Golden parachute payments. 1.280G-1 Section 1... (CONTINUED) INCOME TAXES (CONTINUED) Items Not Deductible § 1.280G-1 Golden parachute payments. The following questions and answers relate to the treatment of golden parachute payments under section 280G of the...

  20. 26 CFR 1.280G-1 - Golden parachute payments.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 26 Internal Revenue 3 2013-04-01 2013-04-01 false Golden parachute payments. 1.280G-1 Section 1... (CONTINUED) INCOME TAXES (CONTINUED) Items Not Deductible § 1.280G-1 Golden parachute payments. The following questions and answers relate to the treatment of golden parachute payments under section 280G of the...

  1. NARSTO SOS99NASH G-1 AIR CHEMISTRY DATA

    Atmospheric Science Data Center

    2014-04-25

    NARSTO SOS99NASH G-1 AIR CHEMISTRY DATA Project Title:  NARSTO ... Carbon Monoxide Ultraviolet Radiation Atmospheric Pressure Atmospheric Temperature Dew Point Upper Level Winds ... Data Guide Documents:  SOS99Nash G-1 Air Guide Related Data:  Southern Oxidants Study ...

  2. Immunoglobulin G1 Fc domain motions: implications for Fc engineering

    PubMed Central

    Frank, Martin; Walker, Ross C.; Lanzilotta, William N.; Prestegard, James H.; Barb, Adam W.

    2014-01-01

    The fragment crystallizable (Fc) region links the key pathogen identification and destruction properties of immunoglobulin G(IgG). Pathogen opsonization positions Fcs to activate pro-inflammatory Fcγ receptors (FcγRs) on immune cells. The cellular response and committal to a damaging, though protective, immune response is tightly controlled at multiple levels. Control mechanisms are diverse and in many cases unclear, but one frequently suggested contribution originates in Fcγ receptor affinity being modulated through shifts in Fc conformational sampling. Here we report a previously unseen IgG1 Fc conformation. This observation motivated an extensive molecular dynamics (MD) investigation of polypeptide and glycan motions that revealed greater amplitude of motion for the N-terminal Cγ2 domains and N-glycan than previously observed. Residues in the Cγ2/Cγ3 interface and disulphide-bonded hinge were identified as influencing the Cγ2 motion. Our results are consistent with a model of Fc that is structurally dynamic. Conformational states that are competent to bind immune-stimulating FcγRs interconverted with Fc conformations distinct from those observed in FcγR complexes, which may represent a transient, nonbinding population. PMID:24522230

  3. Functional overlap among distinct G1/S inhibitory pathways allows robust G1 arrest by yeast mating pheromones.

    PubMed

    Pope, Patricia A; Pryciak, Peter M

    2013-12-01

    In budding yeast, mating pheromones arrest the cell cycle in G1 phase via a pheromone-activated Cdk-inhibitor (CKI) protein, Far1. Alternate pathways must also exist, however, because deleting the cyclin CLN2 restores pheromone arrest to far1 cells. Here we probe whether these alternate pathways require the G1/S transcriptional repressors Whi5 and Stb1 or the CKI protein Sic1, whose metazoan analogues (Rb or p27) antagonize cell cycle entry. Removing Whi5 and Stb1 allows partial escape from G1 arrest in far1 cln2 cells, along with partial derepression of G1/S genes, which implies a repressor-independent route for inhibiting G1/S transcription. This route likely involves pheromone-induced degradation of Tec1, a transcriptional activator of the cyclin CLN1, because Tec1 stabilization also causes partial G1 escape in far1 cln2 cells, and this is additive with Whi5/Stb1 removal. Deleting SIC1 alone strongly disrupts Far1-independent G1 arrest, revealing that inhibition of B-type cyclin-Cdk activity can empower weak arrest pathways. Of interest, although far1 cln2 sic1 cells escaped G1 arrest, they lost viability during pheromone exposure, indicating that G1 exit is deleterious if the arrest signal remains active. Overall our findings illustrate how multiple distinct G1/S-braking mechanisms help to prevent premature cell cycle commitment and ensure a robust signal-induced G1 arrest.

  4. Histone deacetylase homologs regulate epigenetic inheritance of transcriptional silencing and chromosome segregation in fission yeast.

    PubMed

    Grewal, S I; Bonaduce, M J; Klar, A J

    1998-10-01

    Position-effect control at the silent mat2-mat3 interval and at centromeres and telomeres in fission yeast is suggested to be mediated through the assembly of heterochromatin-like structures. Therefore, trans-acting genes that affect silencing may encode either chromatin proteins, factors that modify them, or factors that affect chromatin assembly. Here, we report the identification of an essential gene, clr6 (cryptic loci regulator), which encodes a putative histone deacetylase that when mutated affects epigenetically maintained repression at the mat2-mat3 region and at centromeres and reduces the fidelity of chromosome segregation. Furthermore, we show that the Clr3 protein, when mutated, alleviates recombination block at mat region as well as silencing at donor loci and at centromeres and telomeres, also shares strong homology to known histone deacetylases. Genetic analyses indicate that silencing might be regulated by at least two overlapping histone deacetylase activities. We also found that transient inhibition of histone deacetylase activity by trichostatin A results in the increased missegregation of chromosomes in subsequent generations and, remarkably, alters the imprint at the mat locus, causing the heritable conversion of the repressed epigenetic state to the expressed state. This work supports the model that the level of histone deacetylation has a role in the assembly of repressive heterochromatin and provides insight into the mechanism of epigenetic inheritance.

  5. Bla g 1 allergen levels in Zagreb area household dust.

    PubMed

    Prester, Ljerka; Macan, Jelena

    2011-03-01

    Cockroach allergy is a health problem in many parts of the world. In urban environments, indoor exposure to cockroach allergens involves a risk of asthma. The aim of this study was to measure the mass fraction of Bla g 1, a major allergen of the German cockroach (Blatella germanica) in 30 house samples, collected at random from Zagreb area households, Croatia. Dust samples were collected on cellulose filters by vacuuming living rooms floors. After extraction, Bla g 1 was detected using the commercial enzyme-linked immunosorbent assay (ELISA). Only four of the thirty households had detectable Bla g 1 levels, and only in one was its concentration higher than 2.0 U g(-1), the threshold associated with sensitisation. The Bla g 1 ELISA proved highly sensitive, with the detection limit of 0.12 U g(-1). The within- and between-assay imprecision was 8.9 % and 14.4 %, respectively, and accuracy 85 % to 120 %. Low Bla g 1 levels in the household dust support previously reported low prevalence of skin sensitisation to B. germanica among Zagreb residents. Further monitoring should reveal if there are differences in cockroach allergen exposure and sensitisation between households from other geographic areas in Croatia.

  6. The retinoblastoma protein: functions beyond the G1-S regulator.

    PubMed

    Uchida, Chiharu

    2012-12-01

    Retinoblastoma protein (pRB) is functionally inactivated in a large number of tumors including retinoblastoma, osteosarcoma, small-cell lung carcinoma, as well as bladder, breast and prostate cancers. The best known role of pRB in preventing cancer is inhibition of cell cycle progression by controlling the exit from the cell cycle into G0/G1. In addition, increasing evidence has suggested that pRB has important roles in DNA replication during S phase and G2/M transition. The tumor suppressor function of pRB has also been demonstrated by directly promoting differentiation via cell cycle exit with specific gene expression. Inactivation of pRB function during these cell cycle phases leads to dysregulated cell proliferation and/or chromosomal instability, which are strongly linked to cancer development. Thus pRB plays important roles through multiple functions in determining cell fate, i.e., normal growth/death and differentiation, or tumor formation. Therapeutic intervention by reactivation of pRB function would be expected to be an effective treatment against various cancers.

  7. Histone deacetylases: Targets for antifungal drug development

    PubMed Central

    Kmetzsch, Livia

    2015-01-01

    The interaction of pathogens and its hosts causes a drastic change in the transcriptional landscape in both cells. Among the several mechanisms of gene regulation, transcriptional initiation is probably the main point. In such scenario, the access of transcriptional machinery to promoter is highly regulated by post-translational modification of histones, such as acetylation, phosphorylation and others. Inhibition of histone deacetylases is able to reduce fungal pathogens fitness during infection and, therefore, is currently being considered for the development of new antifungal therapy strategies. PMID:26151486

  8. Histone deacetylase inhibitor for NUT midline carcinoma.

    PubMed

    Maher, Ossama M; Christensen, Anthony M; Yedururi, Sireesha; Bell, Diana; Tarek, Nidale

    2015-04-01

    NUT Midline carcinoma (NMC) is a rare and invariably fatal poorly differentiated carcinoma characterized by chromosomal rearrangement involving the nuclear protein of the testis (NUT) gene. Current approaches do not provide durable response. We report a case of widely metastatic NMC in a 17-year-old female who, following an initial response to combination chemotherapy developed rapid disease progression. Treatment with vorinostat, a histone deacetylase inhibitor (HDACi) resulted in an objective response, yet she died in less than one year from initial diagnosis. This report shows a potentially promising activity of HDACi in the treatment of NMC that needs further exploration. © 2015 Wiley Periodicals, Inc.

  9. Chitin oligosaccharide deacetylase from Shewanella baltica ATCC BAA-1091.

    PubMed

    Hirano, Takako; Shiraishi, Haruka; Ikejima, Masafumi; Uehara, Rie; Hakamata, Wataru; Nishio, Toshiyuki

    2017-03-01

    Chitin oligosaccharide deacetylase (COD) from bacteria that have been examined so far typically comprise two carbohydrate-binding domains (CBDs) and one polysaccharide deacetylase domain. In contrast, Shewanella baltica ATCC BAA-1091 COD (Sb-COD) has only one CBD, yet exhibits chitin-binding properties and substrate specificities similar to those of other CODs.

  10. Loss of Brap Results in Premature G1/S Phase Transition and Impeded Neural Progenitor Differentiation.

    PubMed

    Lanctot, Alison A; Guo, Yan; Le, Yicong; Edens, Brittany M; Nowakowski, Richard S; Feng, Yuanyi

    2017-08-01

    Cells initiate fate decisions during G1 phase by converting extracellular signals into distinctive cell cycle kinetics. The DNA replication timing is determined in G1 phase; lengthened G1 and hastened S phases correlate with increased neurogenic propensity of neural progenitor cells (NPCs), although the underlying molecular control remains elusive. Here, we report that proper G1 phase completion in NPCs requires Brap, a Ras-Erk signaling modulator with ubiquitin E3 ligase activity. We identified Skp2 and Skp2-associated SCF ubiquitin ligase as a key target of Brap-mediated polyubiquitination. Loss of Brap resulted in elevated Skp2, which increased p27(Kip1) destruction, leading to G1 phase truncation and premature S phase entry. The aberrantly executed G1 in Brap-mutant NPCs, followed by hindered S phase progression and increased G2 phase arrest, which together prolonged the cell cycle, impeded neuronal differentiation and culminated in microcephaly. These findings demonstrate that neuronal differentiation is potentiated during G1 phase by Brap-directed cascade of events in cell signaling and protein turnover. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  11. Heart failure: the pivotal role of histone deacetylases.

    PubMed

    Hewitson, Ruth; Dargan, James; Collis, David; Green, Aneta; Moorjani, Narain; Ohri, Sunil; Townsend, Paul A

    2013-02-01

    Heart failure, a state in which cardiac output is unable to meet the metabolic demands of the tissues, poses a significant health burden; following an initial hospital admission with heart failure, five-year mortality is close to 50%. Cardiac hypertrophy, characterised by increased cardiomyocyte size and protein synthesis, has deleterious effects when prolonged and contributes to heart failure. Cardiac hypertrophy itself increases risk of morbidity and mortality. Histone deacetylases are chromatin modifiers which deacetylate the N-terminal tails of histones and have been implicated in common cardiac pathologies associated with hypertrophy. There are 18 histone deacetylases separated into four classes. Class I histone deacetylases interact with heat shock proteins and are pro-hypertrophic, class IIa histone deacetylases repress hypertrophy by inhibiting the activity of transcription factors such as myocyte enhancer factor 2. Histone deacetylases present an exciting new target in combating cardiac hypertrophy and progression to heart failure.

  12. Characterization of api g 1.0201, a new member of the Api g 1 family of celery allergens.

    PubMed

    Hoffmann-Sommergruber, K; Ferris, R; Pec, M; Radauer, C; O'Riordain, G; Laimer Da Camara Machado, M; Scheiner, O; Breiteneder, H

    2000-06-01

    The association of pollinosis with allergy to plant foods occurs in up to 70% of tree pollen-allergic patients. In recent years, some of the relevant cross-reacting proteins have been characterized at the molecular and immunological level. Api g 1 has been identified as the celery homologue of the major birch pollen allergen, Bet v 1. Although a number of Bet v 1 isoforms have been characterized from birch pollen, little is known about isoforms of food allergens and their allergenic features. Api g 1.0201, an isoform of Api g 1, was isolated from a cDNA library, cloned and sequenced. The cDNA was expressed in Escherichia coli and the purified recombinant protein was tested in immunoblots. Api g 1.0201 displays 72% sequence similarity to the previously identified Api g 1.0101 and consists of 159 amino acid residues. The sequence of Api g 1.0201 has five additional amino acid residues at the carboxy-terminus as compared to Api g 1.0101. Purified recombinant Api g 1.0201 is recognized by IgE from the sera of celery-allergic patients, as well as by the murine monoclonal anti-Bet v 1 antibody. In general, this isoform displays a weaker IgE-binding capacity than Api g 1.0101, as concluded from immunoblotting experiments. Results from inhibition assays revealed that IgE-binding to Api g 1.0201 is only slightly reduced by preincubation with either purified recombinant Api g 1.0101 or purified recombinant Bet v 1a. Total inhibition was only achieved when using purified natural Bet v 1. At present, little is known about the IgE-binding capacity of isoforms of Bet v 1 homologues of food allergens. Identification and characterization of such isoforms may help to contribute to a better understanding of food allergy and the observed cross-reactivity to pollen allergy. Copyright 2000 S. Karger AG, Basel.

  13. Final checkup of neoplastic DNA replication: evidence for failure in decision-making at the mitotic cell cycle checkpoint G(1)/S.

    PubMed

    Prindull, Gregor

    2008-11-01

    Processing of epigenomic transcriptional information by cell cycle phase G(1) and decision-making at checkpoint G(1)/S are the final organizational steps preceding gene replication in transcriptional reorientation programs (i.e., switches from proliferation to cycle arrest and neoplastic transformation). Further analyses of cycle progression will open up new approaches in antineoplastic therapy. The following bibliographic databases were consulted: Central Medical Library Cologne, PubMed (English), the last search was done on April 23,2008 and key words searched were: cell cycle, cell memory, DNA methylation, embryonal/neoplastic stem cells, enzyme-modulated chromatin, G(1)-G(1)/S checkpoint, genomic/epigenomics, genomic viral DNA, histones, telomere/telomerases, transcription factors, neoplastic transformation, senescence. Gene transcription and epigenomic surveillance form a functional entity. In proliferation programs, transcriptional information is mediated by chromatin and DNA methylation, analyzed and processed in G(1) phase, and converged on the parental checkpoint G(1)/S for final decision-making on DNA replication. Genomic reorientation appears to be associated with transcriptional instability, which normally is corrected, possibly during the G(2)/M phase, to new levels of epigenomic equilibria. We speculate that daughter stem cells inherit persistent neoplasm-specific transcriptional instabilities through failure of the parental G(1)/S checkpoint. Foreign, silenced, potentially oncogenic DNA sequences, i.e. regular components of the human genome such as endogenous retroviruses, could conceivably be activated for expression in neoplastic transformation by epigenomic histone deacetylase/acetyl transferase/histone methyltransferase-mixed lineage leukemia deregulations. Failure of cell cycle G(1)/S decision-making for DNA replication is the final and possibly a major cause in neoplastic transformation. Therefore, further analysis of the dynamics of G(1)-G(1

  14. Deuteron Spin Structure function g1 at low Q^2

    NASA Astrophysics Data System (ADS)

    Adhikari, Krishna; Kuhn, Sebastian

    2012-10-01

    The spin structure function g1(x,Q^2) and its moments provide crucial information on the internal structure of the nucleon. At low momentum transfer Q^2, one can study the transition from partonic (quark-gluon) to hadronic (nucleonic) degrees of freedom and test effective theories based on QCD, for instance Chiral Perturbation Theory (ChPT). As Q^2 goes to zero, the first moment of g1 is constrained by the GDH sum rule and its ChPT extensions, which makes measurements of g1 in this region uniquely interesting. As part of the large program of spin structure function measurements with CLAS at Jefferson Lab, the EG4 experiment measured the cross section difference between electron beam and proton/deuteron target spins parallel and antiparallel to each other (and the beam direction) down to small scattering angles (approx. 7 degrees). From these differences, g1 can be extracted, with minimal model uncertainties, down to Q^2 as low as 0.01 GeV^2. We will give a brief overview of the experiment and its analysis, and present first preliminary results on the deuteron spin structure function g1d(x,Q^2).

  15. Histone deacetylase 10 structure and molecular function as a polyamine deacetylase

    NASA Astrophysics Data System (ADS)

    Hai, Yang; Shinsky, Stephen A.; Porter, Nicholas J.; Christianson, David W.

    2017-05-01

    Cationic polyamines such as spermidine and spermine are critical in all forms of life, as they regulate the function of biological macromolecules. Intracellular polyamine metabolism is regulated by reversible acetylation and dysregulated polyamine metabolism is associated with neoplastic diseases such as colon cancer, prostate cancer and neuroblastoma. Here we report that histone deacetylase 10 (HDAC10) is a robust polyamine deacetylase, using recombinant enzymes from Homo sapiens (human) and Danio rerio (zebrafish). The 2.85 Å-resolution crystal structure of zebrafish HDAC10 complexed with a transition-state analogue inhibitor reveals that a glutamate gatekeeper and a sterically constricted active site confer specificity for N8-acetylspermidine hydrolysis and disfavour acetyllysine hydrolysis. Both HDAC10 and spermidine are known to promote cellular survival through autophagy. Accordingly, this work sets a foundation for studying the chemical biology of autophagy through the structure-based design of inhibitors that may also serve as new leads for cancer chemotherapy.

  16. Selenium-containing histone deacetylase inhibitors for melanoma management

    PubMed Central

    Gowda, Raghavendra; Madhunapantula, SubbaRao V.; Desai, Dhimant; Amin, Shantu; Robertson, Gavin P.

    2012-01-01

    Melanoma incidence and mortality rates continue to increase each year. Lack of clinically viable agents, drug combinations, effective targeted delivery approaches and success inhibiting targets in tumor tissue have made this disease one of the most difficult to treat, which makes prevention an important option for decreasing disease incidence and mortality rates. Inhibiting histone deacetylases (HDAC) is an approach currently being explored to more effectively treat melanoma but use for prevention has not been explored. In this study, novel selenium containing derivatives of the FDA approved HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) called 5-phenylcarbamoylpentyl selenocyanide (PCP-SeCN) and Bis{5-phenylcarbamoylpentyl} diselenide (B(PCP)-2Se) were created and efficacy tested for preventing early melanocytic lesion development in skin. Topical application of PCP-SeCN and B(PCP)-2Se inhibited melanocytic lesion development in laboratory-generated skin by up to 87% with negligible toxicological effect. Mechanistically, PCP-SeCN and B(PCP)-2Se inhibited HDAC activity and had new inhibitory properties by moderating Akt activity to induce cellular apoptosis as demonstrated by an increase in the sub-G0-G1 cell population, and cleaved caspase-3 as well as PARP levels. Furthermore, PCP-SeCN and B(PCP)-2Se inhibited cell proliferation by inhibiting cyclin D1 expression and increasing p21 levels. Thus, PCP-SeCN and B(PCP)-2Se are potential melanoma chemopreventive agents with enhanced efficacy compared with SAHA due to new PI3 kinase pathway inhibitory properties. PMID:22669577

  17. Rapid biodegradation of organophosphorus pesticides by Stenotrophomonas sp. G1.

    PubMed

    Deng, Shuyan; Chen, Yao; Wang, Daosheng; Shi, Taozhong; Wu, Xiangwei; Ma, Xin; Li, Xiangqiong; Hua, Rimao; Tang, Xinyun; Li, Qing X

    2015-10-30

    Organophosphorus insecticides have been widely used, which are highly poisonous and cause serious concerns over food safety and environmental pollution. A bacterial strain being capable of degrading O,O-dialkyl phosphorothioate and O,O-dialkyl phosphate insecticides, designated as G1, was isolated from sludge collected at the drain outlet of a chlorpyrifos manufacture plant. Physiological and biochemical characteristics and 16S rDNA gene sequence analysis suggested that strain G1 belongs to the genus Stenotrophomonas. At an initial concentration of 50 mg/L, strain G1 degraded 100% of methyl parathion, methyl paraoxon, diazinon, and phoxim, 95% of parathion, 63% of chlorpyrifos, 38% of profenofos, and 34% of triazophos in 24 h. Orthogonal experiments showed that the optimum conditions were an inoculum volume of 20% (v/v), a substrate concentration of 50 mg/L, and an incubation temperature in 40 °C. p-Nitrophenol was detected as the metabolite of methyl parathion, for which intracellular methyl parathion hydrolase was responsible. Strain G1 can efficiently degrade eight organophosphorus pesticides (OPs) and is a very excellent candidate for applications in OP pollution remediation.

  18. Deuteron Spin Structure function g1 at low Q2

    NASA Astrophysics Data System (ADS)

    Adhikari, Krishna; Kuhn, Sebastian

    2013-10-01

    The spin structure function g1 (x ,Q2) and its moments provide crucial information on the internal structure of the nucleon. At low momentum transfer Q2, one can study the transition from partonic (quark-gluon) to hadronic (nucleonic) degrees of freedom and test effective theories based on QCD, such as Chiral Perturbation Theory (χPT). As Q2 goes to zero, the first moment of g1 is constrained by the GDH sum rule and its χPT extensions, which makes measurements of g1 in this region uniquely interesting. As part of a large program of spin structure function measurements with CLAS at Jefferson Lab, the EG4 experiment measured the polarized cross section difference (between the cases of longitudinally polarized electron beam and proton/deuteron target having parallel and antiparallel spins) down to about 7 degrees in the scattering angles. From these differences, g1 can be extracted, with minimal model uncertainties, down to Q2 as low as 0.01 GeV2. We will discuss the experiment and the status of its analysis and present preliminary results. Supported by DOE grant DEFG0296ER40960.

  19. Human pancreatic β-cell G1/S molecule cell cycle atlas.

    PubMed

    Fiaschi-Taesch, Nathalie M; Kleinberger, Jeffrey W; Salim, Fatimah G; Troxell, Ronnie; Wills, Rachel; Tanwir, Mansoor; Casinelli, Gabriella; Cox, Amy E; Takane, Karen K; Scott, Donald K; Stewart, Andrew F

    2013-07-01

    Expansion of pancreatic β-cells is a key goal of diabetes research, yet induction of adult human β-cell replication has proven frustratingly difficult. In part, this reflects a lack of understanding of cell cycle control in the human β-cell. Here, we provide a comprehensive immunocytochemical "atlas" of G1/S control molecules in the human β-cell. This atlas reveals that the majority of these molecules, previously known to be present in islets, are actually present in the β-cell. More importantly, and in contrast to anticipated results, the human β-cell G1/S atlas reveals that almost all of the critical G1/S cell cycle control molecules are located in the cytoplasm of the quiescent human β-cell. Indeed, the only nuclear G1/S molecules are the cell cycle inhibitors, pRb, p57, and variably, p21: none of the cyclins or cdks necessary to drive human β-cell proliferation are present in the nuclear compartment. This observation may provide an explanation for the refractoriness of human β-cells to proliferation. Thus, in addition to known obstacles to human β-cell proliferation, restriction of G1/S molecules to the cytoplasm of the human β-cell represents an unanticipated obstacle to therapeutic human β-cell expansion.

  20. Human Pancreatic β-Cell G1/S Molecule Cell Cycle Atlas

    PubMed Central

    Fiaschi-Taesch, Nathalie M.; Kleinberger, Jeffrey W.; Salim, Fatimah G.; Troxell, Ronnie; Wills, Rachel; Tanwir, Mansoor; Casinelli, Gabriella; Cox, Amy E.; Takane, Karen K.; Scott, Donald K.; Stewart, Andrew F.

    2013-01-01

    Expansion of pancreatic β-cells is a key goal of diabetes research, yet induction of adult human β-cell replication has proven frustratingly difficult. In part, this reflects a lack of understanding of cell cycle control in the human β-cell. Here, we provide a comprehensive immunocytochemical “atlas” of G1/S control molecules in the human β-cell. This atlas reveals that the majority of these molecules, previously known to be present in islets, are actually present in the β-cell. More importantly, and in contrast to anticipated results, the human β-cell G1/S atlas reveals that almost all of the critical G1/S cell cycle control molecules are located in the cytoplasm of the quiescent human β-cell. Indeed, the only nuclear G1/S molecules are the cell cycle inhibitors, pRb, p57, and variably, p21: none of the cyclins or cdks necessary to drive human β-cell proliferation are present in the nuclear compartment. This observation may provide an explanation for the refractoriness of human β-cells to proliferation. Thus, in addition to known obstacles to human β-cell proliferation, restriction of G1/S molecules to the cytoplasm of the human β-cell represents an unanticipated obstacle to therapeutic human β-cell expansion. PMID:23493570

  1. Structure of the histone deacetylase SIRT2.

    PubMed

    Finnin, M S; Donigian, J R; Pavletich, N P

    2001-07-01

    Sir2 is an NAD-dependent histone deacetylase that mediates transcriptional silencing at mating-type loci, telomeres and ribosomal gene clusters, and has a critical role in the determination of life span in yeast and Caenorhabditis elegans. The 1.7 A crystal structure of the 323 amino acid catalytic core of human SIRT2, a homolog of yeast Sir2, reveals an NAD-binding domain, which is a variant of the Rossmann fold, and a smaller domain composed of a helical module and a zinc-binding module. A conserved large groove at the interface of the two domains is the likely site of catalysis based on mutagenesis. Intersecting this large groove, there is a pocket formed by the helical module. The pocket is lined with hydrophobic residues conserved within each of the five Sir2 classes, suggesting that it is a class-specific protein-binding site.

  2. Histone deacetylases play distinct roles in telomeric VSG expression site silencing in African trypanosomes.

    PubMed

    Wang, Qiao-Ping; Kawahara, Taemi; Horn, David

    2010-09-01

    African trypanosomes evade the host immune response through antigenic variation, which is achieved by periodically expressing different variant surface glycoproteins (VSGs). VSG expression is monoallelic such that only one of approximately 15 telomeric VSG expression sites (ESs) is transcribed at a time. Epigenetic regulation is involved in VSG control but our understanding of the mechanisms involved remains incomplete. Histone deacetylases are potential drug targets for diseases caused by protozoan parasites. Here, using recombinant expression we show that the essential Trypanosoma brucei deacetylases, DAC1 (class I) and DAC3 (class II) display histone deacetylase activity. Both DAC1 and DAC3 are nuclear proteins in the bloodstream stage parasite, while only DAC3 remains concentrated in the nucleus in insect-stage cells. Consistent with developmentally regulated localization, DAC1 antagonizes SIR2rp1-dependent telomeric silencing only in the bloodstream form, indicating a conserved role in the control of silent chromatin domains. In contrast, DAC3 is specifically required for silencing at VSG ES promoters in both bloodstream and insect-stage cells. We conclude that DAC1 and DAC3 play distinct roles in subtelomeric gene silencing and that DAC3 represents the first readily druggable target linked to VSG ES control in the African trypanosome.

  3. IgG1 Is Pathogenic in Leishmania mexicana Infection

    PubMed Central

    Chu, Niansheng; Thomas, Bolaji N.; Patel, Supriya R.; Buxbaum, Laurence U.

    2010-01-01

    There are over 2 million new cases of leishmaniasis annually, and no effective vaccine has been developed to prevent infection. In murine infection, Leishmania mexicana, which lives intracellularly in host macrophages, has developed pathways to hijack host IgG to induce a suppressive IL-10 response through FcγRs, the cell-surface receptors for IgG. To guide vaccine development away from detrimental Ab responses, which can accompany attempts to induce cell-mediated immunity, it is crucial to know which isotypes of IgG are pathogenic in this infection. We have found that IgG1 and IgG2a/c induce IL-10 from macrophages in vitro equally well but through different FcγR subtypes: IgG1 through FcγRIII, and IgG2a/c through FcγRI primarily, but also through FcγRIII. In sharp contrast, mice lacking IgG1 develop earlier and stronger IgG2a/c, IgG3, and IgM responses to L. mexicana infection and yet are more resistant to the infection. Thus, IgG1, but not IgG2a/c or IgG3, is pathogenic in vivo, in agreement with prior studies indicating that FcγRIII is required for chronic disease. This calls into question the assumption that macrophages, which should secrete IL-10 in response to both IgG1 and IgG2a/c immune complexes, are the most important source of IL-10 generated by IgG-FcγR engagement in L. mexicana infection. Further investigations are required to better determine the cell type responsible for this immunosuppressive FcγRIII-induced IL-10 pathway and whether IgG2a/c is protective. PMID:21037092

  4. The Histone Deacetylase SIRT6 Regulates Glucose Homeostasis via Hif1α

    PubMed Central

    Zhong, Lei; D'Urso, Agustina; Toiber, Debra; Sebastian, Carlos; Henry, Ryan E.; Vadysirisack, Douangsone D.; Guimaraes, Alexander; Marinelli, Brett; Wikstrom, Jakob D.; Nir, Tomer; Clish, Clary B.; Vaitheesvaran, Bhavapriya; Iliopoulos, Othon; Kurland, Irwin; Dor, Yuval; Weissleder, Ralph; Shirihai, Orian S.; Ellisen, Leif W.; Espinosa, Joaquin M.; Mostoslavsky, Raul

    2010-01-01

    Summary SIRT6 is a member of a highly conserved family of NAD+-dependent deacetylases with various roles in metabolism, stress resistance and lifespan. SIRT6 deficient mice develop normally but succumb to a lethal hypoglycemia early in life; however, the mechanism underlying this hypoglycemia remained unclear. Here, we demonstrate that SIRT6 functions as a histone H3K9 deacetylase to control the expression of multiple glycolytic genes. Specifically, SIRT6 appears to function as a co-repressor of the transcription factor Hif1α, a critical regulator of nutrient stress responses. Consistent with this notion, SIRT6 deficient cells exhibit increased Hif1α activity and show increased glucose uptake with up-regulation of glycolysis and diminished mitochondrial respiration. Our studies uncover a novel role for the chromatin factor SIRT6 as a master regulator of glucose homeostasis, and may provide the basis for novel therapeutic approaches against metabolic diseases, such as diabetes and obesity. PMID:20141841

  5. Characterization and cloning of chitin deacetylases from Rhizopus circinans.

    PubMed

    Gauthier, Carole; Clerisse, Fabienne; Dommes, Jacques; Jaspar-Versali, Marie-France

    2008-05-01

    Chitin deacetylase catalyzes hydrolysis of the acetamido groups of N-acetylglucosamine of chitin in fungal cell walls. Here a chitin deacetylase secreted by Rhizopus circinans was purified to homogeneity and partially characterized. The enzyme exhibits an apparent molecular weight of approximately 75kDa. At 37 degrees C it shows optimal activity at pH 5.5-6. Its pH stability and thermal stability are good. Mn(2+) and Mg(2+) slightly enhance the activity of the enzyme and Cu(2+) strongly inhibits it. An R. circinans cDNA library was constructed and screened with a homologous probe synthesized by RT-PCR or with synthetic primers derived from the N-terminal amino-acid sequence of the native purified chitin deacetylase. Three chitin deacetylase cDNAs (RC, D2, and I3/2) were isolated from the cDNA library and sequenced. These cDNAs exhibit features characteristic of chitin deacetylase sequences: the presence of a polysaccharide deacetylase domain, a metal-binding triad, the conserved catalytic residues, and high homology with various chitin deacetylase genes. The cDNAs were cloned in a Pichia pastoris expression system and produced as polyhistidine-tagged proteins. Only one recombinant enzyme (called RC) was active under the tested conditions. It was purified to homogeneity in a single step and further characterized. The protein showed an apparent molecular mass of approximately 75kDa and, like the native enzyme, showed optimal activity at pH 5.5-6 at 37 degrees C. It was strongly inhibited by Cu(2+). The isolation of several chitin deacetylase cDNAs from the same microorganism is discussed.

  6. Crack azimuths on Europa: The G1 lineament sequence revisited

    USGS Publications Warehouse

    Sarid, A.R.; Greenberg, R.; Hoppa, G.V.; Brown, D.M.; Geissler, P.

    2005-01-01

    The tectonic sequence in the anti-jovian area covered by regional mapping images from Galileo's orbit E15 is determined from a study of cross-cutting relationships among lineament features. The sequence is used to test earlier results from orbit G1, based on lower resolution images, which appeared to display a progressive change in azimuthal orientation over about 90?? in a clockwise sense. Such a progression is consistent with expected stress variations that would accompany plausible non-synchronous rotation. The more recent data provide a more complete record than the G1 data did. We find that to fit the sequence into a continual clockwise change of orientation would require at least 1000?? (> 5 cycles) of azimuthal rotation. If due to non-synchronous rotation of Europa, this result implies that we are seeing back further into the tectonic record than the G1 results had suggested. The three sets of orientations found by Geissler et al. now appear to have been spaced out over several cycles, not during a fraction of one cycle. While our more complete sequence of lineament formation is consistent with non-synchronous rotation, a statistical test shows that it cannot be construed as independent evidence. Other lines of evidence do support non-synchronous rotation, but azimuths of crack sequences do not show it, probably because only a couple of cracks form in a given region in any given non-synchronous rotation period. ?? 2004 Elsevier Inc. All rights reserved.

  7. Human recombinant antimannan immunoglobulin G1 antibody confers resistance to hematogenously disseminated candidiasis in mice.

    PubMed

    Zhang, Mason X; Bohlman, M Charlotte; Itatani, Carol; Burton, Dennis R; Parren, Paul W H I; St Jeor, Stephen C; Kozel, Thomas R

    2006-01-01

    Mannan is a major cell wall component found in Candida species. Natural antimannan antibody is present in sera from most normal adults, but its role in host resistance to hematogenously disseminated candidiasis is unknown. The purpose of this study was to develop recombinant human antimannan antibody and to study its protective function. A phage Fab display combinatorial library containing Fab genes from bone marrow lymphocytes was screened with Candida albicans yeast cells and chemically purified mannan. One antimannan Fab, termed M1, was converted to a full-length immunoglobulin G1 antibody, M1g1, and M1g1 was produced in CHO cells. The M1g1 epitope was found in C. albicans serotypes A and B, Candida tropicalis, Candida guilliermondii, Candida glabrata, and Candida parapsilosis. Its expression was active at both 23 degrees C and 37 degrees C and uniform over the cell surface. BALB/c mice passively immunized with M1g1 were more resistant than control mice to a lethal hematogenous infection by C. albicans, as evidenced by extension of survival in an M1g1 dose-dependent manner (P, 0.08 to <0.001) and by reduction in number of infection foci and their size in the kidney. In vitro studies found that M1g1 promoted phagocytosis and phagocytic killing of C. albicans yeast cells by mouse peritoneal macrophages and was required for activation of the mouse complement cascade. Thus, human antimannan antibody may have a protective role in host resistance to systemic candidiasis.

  8. Human Hydatid Disease in Peru Is Basically Restricted to Echinococcus granulosus Genotype G1

    PubMed Central

    Santivañez, Saul J.; Gutierrez, Ariana M.; Rosenzvit, Mara C.; Muzulin, Patricia M.; Rodriguez, Mary L.; Vasquez, Julio C.; Rodriguez, Silvia; Gonzalez, Armando E.; Gilman, Robert H.; Garcia, Hector H.

    2009-01-01

    A molecular PCR study using DNA from 21 hydatid cysts was performed to determine which strain type is responsible for human infection in Peru. The mitochondrial cytochrome c oxidase subunit 1 (CO1) gene was amplified in 20 out of 21 samples, revealing that all but 1 sample (19/20, 95%) belonged to the common sheep strain (G1). The remaining samples belonged to the camel strain (G6). The G1 genotype was most frequently found in human cases of cystic hydatid disease (CHD) in Peru. Local control measures should focus primarily on decreasing dog and sheep infection rather than intermediate reservoirs. PMID:18606769

  9. Inhibition of Histone Deacetylases Induces Bovine Leukemia Virus Expression In Vitro and In Vivo

    PubMed Central

    Merezak, C.; Reichert, M.; Van Lint, C.; Kerkhofs, P.; Portetelle, D.; Willems, L.; Kettmann, R.

    2002-01-01

    Packaging into nucleosomes results in a global transcriptional repression as a consequence of exclusion of sequence-specific factors. This inhibition can be relieved by using inhibitors of histone deacetylases, acetylation being a major characteristic of transcriptionally active chromatin. Paradoxically, the expression of only ∼2% of the total cellular genes is modulated by histone hyperacetylation. To unravel the potential role of this transcriptional control on BLV expression, we tested the effect of two highly specific inhibitors of deacetylases, trichostatin A (TSA) and trapoxin (TPX). Our results demonstrate that treatment with TSA efficiently enhanced long terminal repeat-directed gene expression of integrated reporter constructs in heterologous D17 stable cell lines. To further examine the biological relevance of these observations made in vitro, we analyzed ex vivo-isolated peripheral blood mononuclear cells (PBMCs) from bovine leukemia virus (BLV)-infected sheep. TSA deacetylase inhibitor induced a drastic increase in viral expression at levels comparable to those induced by treatment with phorbol-12-myristate 13-acetate and ionomycin, the most efficient activators of BLV expression known to date. TSA acted directly on BLV-infected B lymphocytes to increase viral expression and does not seem to require T-cell cooperation. Inhibition of deacetylation after treatment with TSA or TPX also significantly increased viral expression in PBMCs from cattle, the natural host for BLV. Together, our results show that BLV gene expression is, like that of a very small fraction of cellular genes, also regulated by deacetylation. PMID:11967319

  10. A novel histone deacetylase inhibitor Chidamide induces apoptosis of human colon cancer cells

    SciTech Connect

    Liu, Lin; Chen, Baoan; Qin, Shukui; Li, Suyi; He, Xiangming; Qiu, Shaomin; Zhao, Wei; Zhao, Hong

    2010-02-05

    Many studies have demonstrated that histone deacetylase (HDAC) inhibitors induce various tumor cells to undergo apoptosis, and such inhibitors have been used in different clinical trials against different human cancers. In this study, we designed and synthesized a novel HDAC inhibitor, Chidamide. We showed that Chidamide was able to increase the acetylation levels of histone H3 and to inhibit the PI3K/Akt and MAPK/Ras signaling pathways, which resulted in arresting colon cancer cells at the G1 phase of the cell cycle and promoting apoptosis. As a result, the proliferation of colon cancer cells was suppressed in vitro. Our data support the potential application of Chidamide as an anticancer agent in treating colon cancer. Future studies are needed to demonstrate its in vivo efficacy.

  11. Characterization of the N-deacetylase domain from the heparan sulfate N-deacetylase/N-sulfotransferase 2

    SciTech Connect

    Duncan, Michael B.; Liu, May; Fox, Courtney; Liu, Jian . E-mail: jian_liu@unc.edu

    2006-01-27

    Heparin and heparan sulfate are linear sulfated polysaccharides that exert a multitude of biological functions. Heparan sulfate glucosaminyl N-deacetylase/N-sulfotransferase isoform 2 (NDST-2), a key enzyme in the biosynthesis of heparin, contains two distinct activities. This bifunctional enzyme removes the acetyl group from N-acetylated glucosamine (N-deacetylase activity) and transfers a sulfuryl group to the unsubstituted amino position (N-sulfotransferase activity). The N-sulfotransferase activity of NDST has been unambiguously localized to the C-terminal domain of NDST. Here, we report that the N-terminal domain of NDST-2 retains N-deacetylase activity. The N-terminal domain (A66-P604) of human NDST-2, designated as N-deacetylase (NDase), was cloned as a (His){sub 6}-fusion protein, and protein expression was carried out in Escherichia coli. Heparosan treated with NDase contains N-unsubstituted glucosamine and is highly susceptible to N-sulfation by N-sulfotransferase. Our results conclude that the N-terminal domain of NDST-2 contains functional N-deacetylase activity. This finding helps further elucidate the mechanism of action of heparan sulfate N-deacetylase/N-sulfotransferases and the biosynthesis of heparan sulfate in general.

  12. Dual Inhibitors Against Topoisomerases and Histone Deacetylases

    PubMed Central

    Seo, Young Ho

    2015-01-01

    Topoisomerases and histone deacetylases (HDACs) are considered as important therapeutic targets for a wide range of cancers, due to their association with the initiation, proliferation and survival of cancer cells. Topoisomerases are involved in the cleavage and religation processes of DNA, while HDACs regulate a dynamic epigenetic modification of the lysine amino acid on various proteins. Extensive studies have been undertaken to discover small molecule inhibitor of each protein and thereby, several drugs have been transpired from this effort and successfully approved for clinical use. However, the inherent heterogeneity and multiple genetic abnormalities of cancers challenge the clinical application of these single targeted drugs. In order to overcome the limitations of a single target approach, a novel approach, simultaneously targeting topoisomerases and HDACs with a single molecule has been recently employed and attracted much attention of medicinal chemists in drug discovery. This review highlights the current studies on the discovery of dual inhibitors against topoisomerases and HDACs, provides their pharmacological aspects and advantages, and discusses the challenges and promise of the dual inhibitors. PMID:26151040

  13. Dual Inhibitors Against Topoisomerases and Histone Deacetylases.

    PubMed

    Seo, Young Ho

    2015-06-01

    Topoisomerases and histone deacetylases (HDACs) are considered as important therapeutic targets for a wide range of cancers, due to their association with the initiation, proliferation and survival of cancer cells. Topoisomerases are involved in the cleavage and religation processes of DNA, while HDACs regulate a dynamic epigenetic modification of the lysine amino acid on various proteins. Extensive studies have been undertaken to discover small molecule inhibitor of each protein and thereby, several drugs have been transpired from this effort and successfully approved for clinical use. However, the inherent heterogeneity and multiple genetic abnormalities of cancers challenge the clinical application of these single targeted drugs. In order to overcome the limitations of a single target approach, a novel approach, simultaneously targeting topoisomerases and HDACs with a single molecule has been recently employed and attracted much attention of medicinal chemists in drug discovery. This review highlights the current studies on the discovery of dual inhibitors against topoisomerases and HDACs, provides their pharmacological aspects and advantages, and discusses the challenges and promise of the dual inhibitors.

  14. Targeting tumor angiogenesis with Histone Deacetylase Inhibitors

    PubMed Central

    Ellis, Leigh; Hammers, Hans; Pili, Roberto

    2010-01-01

    Solid tumor malignancies including breast, lung and prostate carcinomas are considered to be angiogenesis dependent. Tumor angiogenesis is often mediated by hypoxia secondary to tumor growth or by increased oncogenic signaling. Both mechanisms result in increased hypoxia-inducible factor-1 alpha (HIF-1α) signaling and its transcriptional target vascular endothelial growth factor (VEGF). Critical to HIF-1α signaling are post translational modifications including acetylation mediated by histone acetyltransferases (HATS) and deacetylation by histone deacetylases (HDACs). More recently, HDACs were shown to be up-regulated in response to hypoxia mediating increased HIF-1α signaling. HDAC inhibitors represent a new class of anti-cancer therapeutics which show great promise at inhibiting angiogenesis in pre-clinical animal models and early phase clinical trials. This review will discuss the role of HIF-1α and VEGF influence on tumor angiogenesis and how HDACs play a critical role in HIF-1α transcriptional activity. Furthermore it will also be discussed how targeting HDACs via their inhibition create new avenues in treating solid malignancies by increasing the activity of established and novel therapeutic applications. PMID:19111391

  15. Histone Deacetylases and Their Inhibition in Candida Species

    PubMed Central

    Garnaud, Cécile; Champleboux, Morgane; Maubon, Danièle; Cornet, Muriel; Govin, Jérôme

    2016-01-01

    Fungi are generally benign members of the human mucosal flora or live as saprophytes in the environment. However, they can become pathogenic, leading to invasive and life threatening infections in vulnerable patients. These invasive fungal infections are regarded as a major public health problem on a similar scale to tuberculosis or malaria. Current treatment for these infections is based on only four available drug classes. This limited therapeutic arsenal and the emergence of drug-resistant strains are a matter of concern due to the growing number of patients to be treated, and new therapeutic strategies are urgently needed. Adaptation of fungi to drug pressure involves transcriptional regulation, in which chromatin dynamics and histone modifications play a major role. Histone deacetylases (HDACs) remove acetyl groups from histones and actively participate in controlling stress responses. HDAC inhibition has been shown to limit fungal development, virulence, biofilm formation, and dissemination in the infected host, while also improving the efficacy of existing antifungal drugs toward Candida spp. In this article, we review the functional roles of HDACs and the biological effects of HDAC inhibitors on Candida spp., highlighting the correlations between their pathogenic effects in vitro and in vivo. We focus on how HDAC inhibitors could be used to treat invasive candidiasis while also reviewing recent developments in their clinical evaluation. PMID:27547205

  16. Hypothalamic leptin action is mediated by histone deacetylase 5

    PubMed Central

    Kabra, Dhiraj G.; Pfuhlmann, Katrin; García-Cáceres, Cristina; Schriever, Sonja C.; Casquero García, Veronica; Kebede, Adam Fiseha; Fuente-Martin, Esther; Trivedi, Chitrang; Heppner, Kristy; Uhlenhaut, N. Henriette; Legutko, Beata; Kabra, Uma D.; Gao, Yuanqing; Yi, Chun-Xia; Quarta, Carmelo; Clemmensen, Christoffer; Finan, Brian; Müller, Timo D.; Meyer, Carola W.; Paez-Pereda, Marcelo; Stemmer, Kerstin; Woods, Stephen C.; Perez-Tilve, Diego; Schneider, Robert; Olson, Eric N.; Tschöp, Matthias H.; Pfluger, Paul T.

    2016-01-01

    Hypothalamic leptin signalling has a key role in food intake and energy-balance control and is often impaired in obese individuals. Here we identify histone deacetylase 5 (HDAC5) as a regulator of leptin signalling and organismal energy balance. Global HDAC5 KO mice have increased food intake and greater diet-induced obesity when fed high-fat diet. Pharmacological and genetic inhibition of HDAC5 activity in the mediobasal hypothalamus increases food intake and modulates pathways implicated in leptin signalling. We show HDAC5 directly regulates STAT3 localization and transcriptional activity via reciprocal STAT3 deacetylation at Lys685 and phosphorylation at Tyr705. In vivo, leptin sensitivity is substantially impaired in HDAC5 loss-of-function mice. Hypothalamic HDAC5 overexpression improves leptin action and partially protects against HFD-induced leptin resistance and obesity. Overall, our data suggest that hypothalamic HDAC5 activity is a regulator of leptin signalling that adapts food intake and body weight to our dietary environment. PMID:26923837

  17. Inhibition of histone deacetylase as a new mechanism of teratogenesis.

    PubMed

    Menegola, Elena; Di Renzo, Francesca; Broccia, Maria Luisa; Giavini, Erminio

    2006-12-01

    Histone deacetylases (HDACs) are nuclear and cytoplasmic enzymes that deacetylate a number of substrates, of which histones are the best known and described in the literature. HDACs are present in eukaryotic and bacteria cells, and are fundamental for a number of cellular functions, including correct gene expression. Surprisingly, only up to 20% of the whole genome is controlled by HDACs, but key processes for survival, proliferation, and differentiation have been strictly linked to HDAC enzyme functioning. The use of HDAC inhibitors (HDACi) has been proposed for the treatment of neoplastic diseases. Their effectiveness has been suggested for a number of liquid and solid tumors, particularly acute promyelocytic leukemia (APL). The role of HDACs in embryo development is currently under investigation. Published data indicate knockout phenotype analysis to be of particular interest, in which a number of HDACs play a key role during development. Little data have been published on the effects of HDACi on embryonic development, although for valproic acid (VPA), literature from the 1980s described its teratogenic effects in experimental animals and humans. To date, all tested HDACi have shown teratogenic effects similar to those described for VPA when tested in zebrafish, Xenopus laevis, and mice. HDACs were also able to alter embryo development in invertebrates and plants. A model, similar to that proposed in APL, involving retinoic acid receptors (RAR) and tissue specific Hox gene expression, is suggested to explain the HDAC effects on embryo development.

  18. Sirtuin 1 deacetylase: a key regulator of hepatic lipid metabolism.

    PubMed

    Kemper, Jongsook Kim; Choi, Sung-E; Kim, Dong Hyun

    2013-01-01

    Obesity is a serious medical problem worldwide and disruption of metabolic/energy homeostasis plays a pivotal role in this global epidemic. In obese people, fatty liver (steatosis) develops, which increases the risk for diabetes, cardiovascular disease, and even, liver cancer. Sirtuin 1 (SIRT1) is a NAD+-dependent deacetylase that functions as a key metabolic/energy sensor and mediates homeostatic responses to nutrient availability. Accumulating evidence indicates that SIRT1 is a master regulator of the transcriptional networks that control hepatic lipid metabolism. During energy-deprived conditions, SIRT1 deacetylates and alters the expression and activities of key transcriptional regulators involved in hepatic lipogenesis, fatty acid β-oxidation, and cholesterol/bile acid metabolism. This review will discuss the latest advances in this field, focusing on beneficial roles of SIRT1 in hepatic lipid metabolism including its potential as a therapeutic target for treatment of steatosis and other obesity-related metabolic diseases. Copyright © 2013 Elsevier Inc. All rights reserved.

  19. Enzymatic production of defined chitosan oligomers with a specific pattern of acetylation using a combination of chitin oligosaccharide deacetylases

    PubMed Central

    Hamer, Stefanie Nicole; Cord-Landwehr, Stefan; Biarnés, Xevi; Planas, Antoni; Waegeman, Hendrik; Moerschbacher, Bruno Maria; Kolkenbrock, Stephan

    2015-01-01

    Chitin and chitosan oligomers have diverse biological activities with potentially valuable applications in fields like medicine, cosmetics, or agriculture. These properties may depend not only on the degrees of polymerization and acetylation, but also on a specific pattern of acetylation (PA) that cannot be controlled when the oligomers are produced by chemical hydrolysis. To determine the influence of the PA on the biological activities, defined chitosan oligomers in sufficient amounts are needed. Chitosan oligomers with specific PA can be produced by enzymatic deacetylation of chitin oligomers, but the diversity is limited by the low number of chitin deacetylases available. We have produced specific chitosan oligomers which are deacetylated at the first two units starting from the non-reducing end by the combined use of two different chitin deacetylases, namely NodB from Rhizobium sp. GRH2 that deacetylates the first unit and COD from Vibrio cholerae that deacetylates the second unit starting from the non-reducing end. Both chitin deacetylases accept the product of each other resulting in production of chitosan oligomers with a novel and defined PA. When extended to further chitin deacetylases, this approach has the potential to yield a large range of novel chitosan oligomers with a fully defined architecture. PMID:25732514

  20. Enzymatic production of defined chitosan oligomers with a specific pattern of acetylation using a combination of chitin oligosaccharide deacetylases

    NASA Astrophysics Data System (ADS)

    Hamer, Stefanie Nicole; Cord-Landwehr, Stefan; Biarnés, Xevi; Planas, Antoni; Waegeman, Hendrik; Moerschbacher, Bruno Maria; Kolkenbrock, Stephan

    2015-03-01

    Chitin and chitosan oligomers have diverse biological activities with potentially valuable applications in fields like medicine, cosmetics, or agriculture. These properties may depend not only on the degrees of polymerization and acetylation, but also on a specific pattern of acetylation (PA) that cannot be controlled when the oligomers are produced by chemical hydrolysis. To determine the influence of the PA on the biological activities, defined chitosan oligomers in sufficient amounts are needed. Chitosan oligomers with specific PA can be produced by enzymatic deacetylation of chitin oligomers, but the diversity is limited by the low number of chitin deacetylases available. We have produced specific chitosan oligomers which are deacetylated at the first two units starting from the non-reducing end by the combined use of two different chitin deacetylases, namely NodB from Rhizobium sp. GRH2 that deacetylates the first unit and COD from Vibrio cholerae that deacetylates the second unit starting from the non-reducing end. Both chitin deacetylases accept the product of each other resulting in production of chitosan oligomers with a novel and defined PA. When extended to further chitin deacetylases, this approach has the potential to yield a large range of novel chitosan oligomers with a fully defined architecture.

  1. Enzymatic production of defined chitosan oligomers with a specific pattern of acetylation using a combination of chitin oligosaccharide deacetylases.

    PubMed

    Hamer, Stefanie Nicole; Cord-Landwehr, Stefan; Biarnés, Xevi; Planas, Antoni; Waegeman, Hendrik; Moerschbacher, Bruno Maria; Kolkenbrock, Stephan

    2015-03-03

    Chitin and chitosan oligomers have diverse biological activities with potentially valuable applications in fields like medicine, cosmetics, or agriculture. These properties may depend not only on the degrees of polymerization and acetylation, but also on a specific pattern of acetylation (PA) that cannot be controlled when the oligomers are produced by chemical hydrolysis. To determine the influence of the PA on the biological activities, defined chitosan oligomers in sufficient amounts are needed. Chitosan oligomers with specific PA can be produced by enzymatic deacetylation of chitin oligomers, but the diversity is limited by the low number of chitin deacetylases available. We have produced specific chitosan oligomers which are deacetylated at the first two units starting from the non-reducing end by the combined use of two different chitin deacetylases, namely NodB from Rhizobium sp. GRH2 that deacetylates the first unit and COD from Vibrio cholerae that deacetylates the second unit starting from the non-reducing end. Both chitin deacetylases accept the product of each other resulting in production of chitosan oligomers with a novel and defined PA. When extended to further chitin deacetylases, this approach has the potential to yield a large range of novel chitosan oligomers with a fully defined architecture.

  2. CUDC-907 Promotes Bone Marrow Adipocytic Differentiation Through Inhibition of Histone Deacetylase and Regulation of Cell Cycle.

    PubMed

    Ali, Dalia; Alshammari, Hassan; Vishnubalaji, Radhakrishnan; Chalisserry, Elna Paul; Hamam, Rimi; Alfayez, Musaad; Kassem, Moustapha; Aldahmash, Abdullah; Alajez, Nehad M

    2017-03-01

    The role of bone marrow adipocytes (BMAs) in overall energy metabolism and their effects on bone mass are currently areas of intensive investigation. BMAs differentiate from bone marrow stromal cells (BMSCs); however, the molecular mechanisms regulating BMA differentiation are not fully understood. In this study, we investigated the effect of CUDC-907, identified by screening an epigenetic small-molecule library, on adipocytic differentiation of human BMSCs (hBMSCs) and determined its molecular mechanism of action. Human bone marrow stromal cells exposed to CUDC-907 (500 nM) exhibited enhanced adipocytic differentiation (∼2.9-fold increase, P < 0.005) compared with that of control cells. Global gene expression and signaling pathway analyses of differentially expressed genes revealed a strong enrichment of genes involved in adipogenesis, cell cycle, and DNA replication. Chromatin immune precipitation combined with quantitative polymerase chain reaction showed significant increase in H3K9ac epigenetic marker in the promoter regions of AdipoQ, FABP4, PPARγ, KLF15, and CEBPA in CUDC-907-treated hBMSCs. Follow-up experiments corroborated that the inhibition of histone deacetylase (HDAC) activity enhanced adipocytic differentiation, while the inhibition of PI3K decreased adipocytic differentiation. In addition, CUDC-907 arrested hBMSCs in the G0-G1 phase of the cell cycle and reduced the number of S-phase cells. Our data reveal that HDAC, PI3K, and cell cycle genes are important regulators of BMA formation and demonstrate that adipocyte differentiation of hBMSCs is associated with complex changes in a number of epigenetic and genetic pathways, which can be targeted to regulate BMA formation.

  3. ATP6V1G1 — EDRN Public Portal

    Cancer.gov

    ATP6V1G1 is a subunit of vacuolar ATPase (V-ATPase), a multisubunit enzyme. V-ATPase is an enzyme transporter that functions to acidify intracellular compartments in eukaryotic cells. This acidification process is necessary for such intracellular processes as protein sorting, zymogen activation, receptor-mediated endocytosis, and synaptic vesicle proton gradient generation. V-ATPase is ubiquitously expressed and is present in endomembrane organelles such as vacuoles, lysosomes, endosomes, the Golgi apparatus, chromaffin granules and coated vesicles, as well as in the plasma membrane. V-ATPase is composed of a cytosolic V1 domain and a transmembrane V0 domain. The V1 domain consists of three A, three B, and two G subunits, as well as a C, D, E, F, and H subunit. The V1 domain contains the ATP catalytic site.

  4. Histone deacetylase inhibitors (HDACIs): multitargeted anticancer agents

    PubMed Central

    Ververis, Katherine; Hiong, Alison; Karagiannis, Tom C; Licciardi, Paul V

    2013-01-01

    Histone deacetylase (HDAC) inhibitors are an emerging class of therapeutics with potential as anticancer drugs. The rationale for developing HDAC inhibitors (and other chromatin-modifying agents) as anticancer therapies arose from the understanding that in addition to genetic mutations, epigenetic changes such as dysregulation of HDAC enzymes can alter phenotype and gene expression, disturb homeostasis, and contribute to neoplastic growth. The family of HDAC inhibitors is large and diverse. It includes a range of naturally occurring and synthetic compounds that differ in terms of structure, function, and specificity. HDAC inhibitors have multiple cell type-specific effects in vitro and in vivo, such as growth arrest, cell differentiation, and apoptosis in malignant cells. HDAC inhibitors have the potential to be used as monotherapies or in combination with other anticancer therapies. Currently, there are two HDAC inhibitors that have received approval from the US FDA for the treatment of cutaneous T-cell lymphoma: vorinostat (suberoylanilide hydroxamic acid, Zolinza) and depsipeptide (romidepsin, Istodax). More recently, depsipeptide has also gained FDA approval for the treatment of peripheral T-cell lymphoma. Many more clinical trials assessing the effects of various HDAC inhibitors on hematological and solid malignancies are currently being conducted. Despite the proven anticancer effects of particular HDAC inhibitors against certain cancers, many aspects of HDAC enzymes and HDAC inhibitors are still not fully understood. Increasing our understanding of the effects of HDAC inhibitors, their targets and mechanisms of action will be critical for the advancement of these drugs, especially to facilitate the rational design of HDAC inhibitors that are effective as antineoplastic agents. This review will discuss the use of HDAC inhibitors as multitargeted therapies for malignancy. Further, we outline the pharmacology and mechanisms of action of HDAC inhibitors while

  5. A mechanism for the suppression of homologous recombination in G1 cells.

    PubMed

    Orthwein, Alexandre; Noordermeer, Sylvie M; Wilson, Marcus D; Landry, Sébastien; Enchev, Radoslav I; Sherker, Alana; Munro, Meagan; Pinder, Jordan; Salsman, Jayme; Dellaire, Graham; Xia, Bing; Peter, Matthias; Durocher, Daniel

    2015-12-17

    DNA repair by homologous recombination is highly suppressed in G1 cells to ensure that mitotic recombination occurs solely between sister chromatids. Although many homologous recombination factors are cell-cycle regulated, the identity of the events that are both necessary and sufficient to suppress recombination in G1 cells is unknown. Here we report that the cell cycle controls the interaction of BRCA1 with PALB2-BRCA2 to constrain BRCA2 function to the S/G2 phases in human cells. We found that the BRCA1-interaction site on PALB2 is targeted by an E3 ubiquitin ligase composed of KEAP1, a PALB2-interacting protein, in complex with cullin-3 (CUL3)-RBX1 (ref. 6). PALB2 ubiquitylation suppresses its interaction with BRCA1 and is counteracted by the deubiquitylase USP11, which is itself under cell cycle control. Restoration of the BRCA1-PALB2 interaction combined with the activation of DNA-end resection is sufficient to induce homologous recombination in G1, as measured by RAD51 recruitment, unscheduled DNA synthesis and a CRISPR-Cas9-based gene-targeting assay. We conclude that the mechanism prohibiting homologous recombination in G1 minimally consists of the suppression of DNA-end resection coupled with a multi-step block of the recruitment of BRCA2 to DNA damage sites that involves the inhibition of BRCA1-PALB2-BRCA2 complex assembly. We speculate that the ability to induce homologous recombination in G1 cells with defined factors could spur the development of gene-targeting applications in non-dividing cells.

  6. A mechanism for the suppression of homologous recombination in G1 cells

    PubMed Central

    Orthwein, Alexandre; Noordermeer, Sylvie M.; Wilson, Marcus D.; Landry, Sébastien; Enchev, Radoslav I.; Sherker, Alana; Munro, Meagan; Pinder, Jordan; Salsman, Jayme; Dellaire, Graham; Xia, Bing; Peter, Matthias; Durocher, Daniel

    2016-01-01

    DNA repair by homologous recombination (HR)1 is highly suppressed in G1 cells2,3 to ensure that mitotic recombination occurs solely between sister chromatids4. Although many HR factors are cell cycle-regulated, the identity of the events that are both necessary and sufficient to suppress recombination in G1 cells is unknown. Here we report that the cell cycle controls the interaction of BRCA1 with PALB2-BRCA2 in order to constrain BRCA2 function to the S/G2 phases. We found that the BRCA1-interaction site on PALB2 is targeted by an E3 ubiquitin ligase composed of KEAP1, a PALB2-interacting protein5, in complex with CUL3-RBX16. PALB2 ubiquitylation suppresses its interaction with BRCA1 and is counteracted by the deubiquitylase USP11, which is itself under cell cycle control. Restoration of the BRCA1-PALB2 interaction combined with the activation of DNA end resection is sufficient to induce HR in G1, as measured by RAD51 recruitment, unscheduled DNA synthesis and a CRISPR/Cas9-based gene targeting assay. We conclude that the mechanism prohibiting HR in G1 minimally consists of the suppression of DNA end resection coupled to a multi-step block to BRCA2 recruitment to DNA damage sites that involves the inhibition of BRCA1-PALB2-BRCA2 complex assembly. We speculate that the ability to induce HR in G1 cells with defined factors could spur the development of gene targeting applications in non-dividing cells. PMID:26649820

  7. Attenuation of G1 checkpoint function by the non-genotoxic carcinogen phenobarbital.

    PubMed

    Gonzales, A J; Christensen, J G; Preston, R J; Goldsworthy, T L; Tlsty, T D; Fox, T R

    1998-07-01

    Non-genotoxic chemical carcinogens are capable of inducing tumors in rodents without interacting with or directly altering the genetic material. Since a preponderance of evidence suggests that cancer results from the accumulation of genetic alterations, the mechanisms by which many non-genotoxic carcinogens induce genotoxic events remain unclear. The present study investigated whether the mitogenic, non-genotoxic carcinogen phenobarbital (PB) could alter cell-cycle checkpoint controls, thereby indirectly leading to the accumulation of genetic damage. Initial studies involved characterizing cell-cycle checkpoint responses to DNA damage in freshly isolated B6C3F1 mouse hepatocytes. These cells responded to bleomycin-induced DNA damage by arresting in G1 and G2. Cell-cycle arrest was coupled with p53 protein induction; however, p21WAF1 protein levels remained unchanged. Studies that utilized hepatocytes isolated from C57BL p53-/- mice showed that the DNA damage-induced G1 cell-cycle arrest was dependent on p53 function, but cell-cycle arrest in G2 was not affected by loss of p53. PB was able to delay and attenuate the G1 checkpoint response without altering G2 checkpoint function. A reduction in p53 protein, but not transcript levels, was observed in hepatocytes exposed to PB. Additionally, PB delayed and attenuated p53 protein induction during DNA damage, which suggests that changes in the p53 protein may be contributing to the attenuated G1 checkpoint response caused by PB. Altered G1 checkpoint function represents an epigenetic mechanism by which phenobarbital may prevent the detection and repair of DNA damage and indirectly increase the frequency of genotoxic events above that occurring spontaneously. Abrogation of checkpoint controls may, thus, play an important mechanistic role in mitogenic, non-genotoxic chemical carcinogenesis.

  8. Production and application of high quality stable isotope-labeled human immunoglobulin G1 for mass spectrometry analysis.

    PubMed

    Phillip, Amsler; Thierry, Wolf; Christian, Lanshoeft; Anja, Bettighofer; Jochen, Eisfeld; Thomas, Moenius; Claudia, Probst; Coralie, Etter; Olivier, Heudi

    2017-03-01

    Here, we describe the production of stable isotope-labeled human immunoglobulin G1 ([(13) C]-hIgG1) using [(13) C]-L-lysine/arginine-labeled hIgG1. The fermentation process was run in shake flasks containing labeled arginine and lysinethat were incorporated into the produced recombinant hIgG1. The [(13) C]-hIgG1 was purified, and label incorporation was determined to be >99% at all lysine and arginine moieties. Sequence coverage was confirmed by peptide mapping. [(13) C]-hIgG1 was then used as an internal standard (IS) for the development of a liquid chromatography-tandem mass spectrometry method applicable to the quantitative analysis of all human types of hIgG1 in rat serum. Four conserved peptides, namely, GPSVFPLAPSSK, TTPPVLDSDGSFFLYSK, VVSVLTVLHQDWLNGK, and FNWYVDGVEVHNAK, originating from different parts of the fraction crystallizable region of hIgG1, were used for quantitation of hIgG1 in rat serum. The calibration curves with a coefficient of determination (r(2) ) between 0.9950 and 0.9962 resulting from the peak area ratio of each peptide to its respective labeled IS were reproducible. A mean bias within ±20.0% of the nominal values and a precision of ≤20.0 % were obtained for the calibration standards and quality control samples for each peptide. [(13) C]-hIgG1 was shown as a suitable IS for quantitative hIgG1 analysis in preclinical species by LC-MS/MS. Copyright © 2016 John Wiley & Sons, Ltd.

  9. Phosphate-Activated Cyclin-Dependent Kinase Stabilizes G1 Cyclin To Trigger Cell Cycle Entry

    PubMed Central

    Menoyo, S.; Ricco, N.; Bru, S.; Hernández-Ortega, S.; Escoté, X.; Aldea, M.

    2013-01-01

    G1 cyclins, in association with a cyclin-dependent kinase (CDK), are universal activators of the transcriptional G1-S machinery during entry into the cell cycle. Regulation of cyclin degradation is crucial for coordinating progression through the cell cycle, but the mechanisms that modulate cyclin stability to control cell cycle entry are still unknown. Here, we show that a lack of phosphate downregulates Cln3 cyclin and leads to G1 arrest in Saccharomyces cerevisiae. The stability of Cln3 protein is diminished in strains with low activity of Pho85, a phosphate-sensing CDK. Cln3 is an in vitro substrate of Pho85, and both proteins interact in vivo. More interestingly, cells that carry a CLN3 allele encoding aspartic acid substitutions at the sites of Pho85 phosphorylation maintain high levels of Cln3 independently of Pho85 activity. Moreover, these cells do not properly arrest in G1 in the absence of phosphate and they die prematurely. Finally, the activity of Pho85 is essential for accumulating Cln3 and for reentering the cell cycle after phosphate refeeding. Taken together, our data indicate that Cln3 is a molecular target of the Pho85 kinase that is required to modulate cell cycle entry in response to environmental changes in nutrient availability. PMID:23339867

  10. Phosphate-activated cyclin-dependent kinase stabilizes G1 cyclin to trigger cell cycle entry.

    PubMed

    Menoyo, S; Ricco, N; Bru, S; Hernández-Ortega, S; Escoté, X; Aldea, M; Clotet, J

    2013-04-01

    G1 cyclins, in association with a cyclin-dependent kinase (CDK), are universal activators of the transcriptional G1-S machinery during entry into the cell cycle. Regulation of cyclin degradation is crucial for coordinating progression through the cell cycle, but the mechanisms that modulate cyclin stability to control cell cycle entry are still unknown. Here, we show that a lack of phosphate downregulates Cln3 cyclin and leads to G1 arrest in Saccharomyces cerevisiae. The stability of Cln3 protein is diminished in strains with low activity of Pho85, a phosphate-sensing CDK. Cln3 is an in vitro substrate of Pho85, and both proteins interact in vivo. More interestingly, cells that carry a CLN3 allele encoding aspartic acid substitutions at the sites of Pho85 phosphorylation maintain high levels of Cln3 independently of Pho85 activity. Moreover, these cells do not properly arrest in G1 in the absence of phosphate and they die prematurely. Finally, the activity of Pho85 is essential for accumulating Cln3 and for reentering the cell cycle after phosphate refeeding. Taken together, our data indicate that Cln3 is a molecular target of the Pho85 kinase that is required to modulate cell cycle entry in response to environmental changes in nutrient availability.

  11. Tau--an inhibitor of deacetylase HDAC6 function.

    PubMed

    Perez, Mar; Santa-Maria, Ismael; Gomez de Barreda, Elena; Zhu, Xiongwei; Cuadros, Raquel; Cabrero, Jose Roman; Sanchez-Madrid, Francisco; Dawson, Hana N; Vitek, Michael P; Perry, George; Smith, Mark A; Avila, Jesus

    2009-06-01

    Analysis of brain microtubule protein from patients with Alzheimer's disease showed decreased alpha tubulin levels along with increased acetylation of the alpha tubulin subunit, mainly in those microtubules from neurons containing neurofibrillary tau pathology. To determine the relationship of tau protein and increased tubulin acetylation, we studied the effect of tau on the acetylation-deacetylation of tubulin. Our results indicate that tau binds to the tubulin-deacetylase, histone deacetylase 6 (HDAC6), decreasing its activity with a consequent increase in tubulin acetylation. As expected, increased acetylation was also found in tubulin from wild-type mice compared with tubulin from mice lacking tau because of the tau-mediated inhibition of the deacetylase. In addition, we found that an excess of tau protein, as a HDAC6 inhibitor, prevents induction of autophagy by inhibiting proteasome function.

  12. Suppression of caspase-11 expression by histone deacetylase inhibitors

    SciTech Connect

    Heo, Hyejung; Yoo, Lang; Shin, Ki Soon; Kang, Shin Jung

    2009-01-02

    It has been well documented that histone deacetylase inhibitors suppress inflammatory gene expression. Therefore, we investigated whether histone deacetylase inhibitors modulate the expression of caspase-11 that is known as an inducible caspase regulating both inflammation and apoptosis. In the present study, we show that sodium butyrate and trichostatin A, two structurally unrelated inhibitors of histone deacetylase (HDAC), effectively suppressed the induction of caspase-11 in mouse embryonic fibroblasts stimulated with lipopolysaccharides. Sodium butyrate inhibited the activation of upstream signaling events for the caspase-11 induction such as activation of p38 mitogen-activated protein kinase and c-Jun N-terminal kinase, degradation of inhibitor of {kappa}B, and activation of nuclear factor-{kappa}B. These results suggest that the HDAC inhibitor suppressed cytosolic signaling events for the induction of caspase-11 by inhibiting the deacetylation of non-histone proteins.

  13. Inhibition of histone deacetylases in cancer therapy: lessons from leukaemia.

    PubMed

    Ceccacci, Elena; Minucci, Saverio

    2016-03-15

    Histone deacetylases (HDACs) are a key component of the epigenetic machinery regulating gene expression, and behave as oncogenes in several cancer types, spurring the development of HDAC inhibitors (HDACi) as anticancer drugs. This review discusses new results regarding the role of HDACs in cancer and the effect of HDACi on tumour cells, focusing on haematological malignancies, particularly acute myeloid leukaemia. Histone deacetylases may have opposite roles at different stages of tumour progression and in different tumour cell sub-populations (cancer stem cells), highlighting the importance of investigating these aspects for further improving the clinical use of HDACi in treating cancer.

  14. G0/G1 Switch Gene 2 Regulates Cardiac Lipolysis*

    PubMed Central

    Heier, Christoph; Radner, Franz P. W.; Moustafa, Tarek; Schreiber, Renate; Grond, Susanne; Eichmann, Thomas O.; Schweiger, Martina; Schmidt, Albrecht; Cerk, Ines K.; Oberer, Monika; Theussl, H.-Christian; Wojciechowski, Jacek; Penninger, Josef M.; Zimmermann, Robert; Zechner, Rudolf

    2015-01-01

    The anabolism and catabolism of myocardial triacylglycerol (TAG) stores are important processes for normal cardiac function. TAG synthesis detoxifies and stockpiles fatty acids to prevent lipotoxicity, whereas TAG hydrolysis (lipolysis) remobilizes fatty acids from endogenous storage pools as energy substrates, signaling molecules, or precursors for complex lipids. This study focused on the role of G0/G1 switch 2 (G0S2) protein, which was previously shown to inhibit the principal TAG hydrolase adipose triglyceride lipase (ATGL), in the regulation of cardiac lipolysis. Using wild-type and mutant mice, we show the following: (i) G0S2 is expressed in the heart and regulated by the nutritional status with highest expression levels after re-feeding. (ii) Cardiac-specific overexpression of G0S2 inhibits cardiac lipolysis by direct protein-protein interaction with ATGL. This leads to severe cardiac steatosis. The steatotic hearts caused by G0S2 overexpression are less prone to fibrotic remodeling or cardiac dysfunction than hearts with a lipolytic defect due to ATGL deficiency. (iii) Conversely to the phenotype of transgenic mice, G0S2 deficiency results in a de-repression of cardiac lipolysis and decreased cardiac TAG content. We conclude that G0S2 acts as a potent ATGL inhibitor in the heart modulating cardiac substrate utilization by regulating cardiac lipolysis. PMID:26350455

  15. Enhancement of memory consolidation by the histone deacetylase inhibitor sodium butyrate in aged rats.

    PubMed

    Blank, Martina; Werenicz, Aline; Velho, Luciana Azevedo; Pinto, Diana F; Fedi, Ana Cláudia; Lopes, Mark William; Peres, Tanara Vieira; Leal, Rodrigo Bainy; Dornelles, Arethuza S; Roesler, Rafael

    2015-05-06

    Here we show that a systemic injection of the histone deacetylase inhibitor (HDACi) sodium butyrate (NaB) immediately after training in a step-down inhibitory avoidance task produced an enhancement of memory consolidation that persisted across consecutive retention tests during 14 days in aged rats, while it did not significantly affect memory in young adults. Control aged and young adult rats showed comparable basal levels of memory retention. Our results suggest that HDACis can display memory-enhancing effects specific for aged animals, even in the absence of age-related memory impairment.

  16. [Effect of phenylbutyrate, a histone deacetylase inhibitor, on differentiation and apoptosis of Kasumi-1 cells].

    PubMed

    Hao, Chang-lai; Tang, Ke-jing; Tian, Zheng; Xing, Hai-yan; Wang, Min; Wang, Jian-xiang

    2003-05-01

    To explore the blockade effect of phenylbutyrate (PB), a histone deacetylase inhibitor, on the in vitro biological function of AML1/ETO to reverse its transcription repression and induce Kasumi-1 cells to differentiate and apoptosis. Kasumi-1 cells were treated with PB at different concentrations in suspension culture. Cell proliferation was analysed by MTT assay, morphological changes by light and electron microscopy, expression of myeloid-specific differentiation antigen and cell cycle by flow cytometry, cell apoptosis by annexin V staining, agarose gel electrophoresis and flow cytometry. PB treatment caused a dose-dependent inhibition of the cell proliferation. The IC(50) was about 2.3 mmol/L. PB treatment led to a progressive decline in the fraction of S-phase cells and increase in G(0)/G(1) cells. PB induced a time- and dose-dependent increase in expression of myeloid cell surface protein CD(11b) and CD(13). A dose-dependent increase in early apoptosis for 2 days treatment, late apoptosis for 3 days treatment. The DNA ladder of apoptosis was observed on agarose gel electrophoresis for 5 days treatment. Morphological features of monocytoid differentiation and apoptosis were seen on Wright-Giemsa staining smears. PB treatment could inhibit proliferation of Kasumi-1 cells, induce partial differentiation, apoptosis and accumulation of cells in G(0)/G(1) phase.

  17. The existence of inflection points for generalized log-aesthetic curves satisfying G1 data

    NASA Astrophysics Data System (ADS)

    Karpagavalli, R.; Gobithaasan, R. U.; Miura, K. T.; Shanmugavel, Madhavan

    2015-12-01

    Log-Aesthetic (LA) curves have been implemented in a CAD/CAM system for various design feats. LA curves possess linear Logarithmic Curvature Graph (LCG) with gradient (shape parameter) denoted as α. In 2009, a generalized form of LA curves called Generalized Log-Aesthetic Curves (GLAC) has been proposed which has an extra shape parameter as ν compared to LA curves. Recently, G1 continuous GLAC algorithm has been proposed which utilizes the extra shape parameter using four control points. This paper discusses on the existence of inflection points in a GLAC segment satisfying G1 Hermite data and the effect of inflection point on convex hull property. It is found that the existence of inflection point can be avoided by manipulating the value of α. Numerical experiments show that the increase of α may remove the inflection point (if any) in a GLAC segment.

  18. The interplay between G protein-coupled receptor kinase 2 (GRK2) and histone deacetylase 6 (HDAC6) at the crossroads of epithelial cell motility

    PubMed Central

    Lafarga, Vanesa; Mayor, Jr, Federico; Penela, Petronila

    2012-01-01

    G protein-coupled receptor kinase 2 (GRK2) is emerging as a key integrative node in cell migration control. In addition to its canonical role in the desensitization of G protein-coupled receptors involved in chemotaxis, novel recently identified GRK2 substrates and interacting partners appear to mediate the GRK2-dependent modulation of diverse molecular processes involved in motility, such as gradient sensing, cell polarity or cytoskeletal reorganization. We have recently identified an interaction between GRK2 and histone deacetylase 6 (HDAC6), a major cytoplasmic α-tubulin deacetylase involved in cell motility and adhesion. GRK2 dynamically associates with and phosphorylates HDAC6 to stimulate its α-tubulin deacetylase activity at specific cellular localizations such as the leading edge of migrating cells, thus promoting local tubulin deacetylation and enhanced motility. This GRK2-HDAC6 functional interaction may have important implications in pathological contexts related to aberrant epithelial cell migration. PMID:23076141

  19. Dux4 induces cell cycle arrest at G1 phase through upregulation of p21 expression

    SciTech Connect

    Xu, Hongliang; Wang, Zhaoxia; Jin, Suqin; Hao, Hongjun; Zheng, Lemin; Zhou, Boda; Zhang, Wei; Lv, He; Yuan, Yun

    2014-03-28

    Highlights: • Dux4 induced TE671 cell proliferation defect and G1 phase arrest. • Dux4 upregulated p21 expression without activating p53. • Silencing p21 rescued Dux4 mediated proliferation defect and cell cycle arrest. • Sp1 binding site was required for Dux4-induced p21 promoter activation. - Abstract: It has been implicated that Dux4 plays crucial roles in development of facioscapulohumeral dystrophy. But the underlying myopathic mechanisms and related down-stream events of this retrogene were far from clear. Here, we reported that overexpression of Dux4 in a cell model TE671 reduced cell proliferation rate, and increased G1 phase accumulation. We also determined the impact of Dux4 on p53/p21 signal pathway, which controls the checkpoint in cell cycle progression. Overexpression of Dux4 increased p21 mRNA and protein level, while expression of p53, phospho-p53 remained unchanged. Silencing p21 rescued Dux4 mediated proliferation defect and cell cycle arrest. Furthermore, we demonstrated that enhanced Dux4 expression increased p21 promoter activity and elevated expression of Sp1 transcription factor. Mutation of Sp1 binding site decreased dux4 induced p21 promoter activation. Chromatin immunoprecipitation (ChIP) assays confirmed the Dux4-induced binding of Sp1 to p21 promoter in vivo. These results suggest that Dux4 might induce proliferation inhibition and G1 phase arrest through upregulation of p21.

  20. The SIT4 protein phosphatase functions in late G1 for progression into S phase.

    PubMed Central

    Sutton, A; Immanuel, D; Arndt, K T

    1991-01-01

    Saccharomyces cerevisiae strains containing temperature-sensitive mutations in the SIT4 protein phosphatase arrest in late G1 at the nonpermissive temperature. Order-of-function analysis shows that SIT4 is required in late G1 for progression into S phase. While the levels of SIT4 do not change in the cell cycle, SIT4 associates with two high-molecular-weight phosphoproteins in a cell-cycle-dependent fashion. In addition, we have identified a polymorphic gene, SSD1, that in some versions can suppress the lethality due to a deletion of SIT4 and can also partially suppress the phenotypic defects due to a null mutation in BCY1. The SSD1 protein is implicated in G1 control and has a region of similarity to the dis3 protein of Schizosaccharomyces pombe. We have also identified a gene, PPH2alpha, that in high copy number can partially suppress the growth defect of sit4 strains. The PPH2 alpha gene encodes a predicted protein that is 80% identical to the catalytic domain of mammalian type 2A protein phosphatases but also has an acidic amino-terminal extension not present in other phosphatases. Images PMID:1848673

  1. Balanced Production of Ribosome Components Is Required for Proper G1/S Transition in Saccharomyces cerevisiae *

    PubMed Central

    Gómez-Herreros, Fernando; Rodríguez-Galán, Olga; Morillo-Huesca, Macarena; Maya, Douglas; Arista-Romero, María; de la Cruz, Jesús; Chávez, Sebastián; Muñoz-Centeno, Mari Cruz

    2013-01-01

    Cell cycle regulation is a very accurate process that ensures cell viability and the genomic integrity of daughter cells. A fundamental part of this regulation consists in the arrest of the cycle at particular points to ensure the completion of a previous event, to repair cellular damage, or to avoid progression in potentially risky situations. In this work, we demonstrate that a reduction in nucleotide levels or the depletion of RNA polymerase I or III subunits generates a cell cycle delay at the G1/S transition in Saccharomyces cerevisiae. This delay is concomitant with an imbalance between ribosomal RNAs and proteins which, among others, provokes an accumulation of free ribosomal protein L5. Consistently with a direct impact of free L5 on the G1/S transition, rrs1 mutants, which weaken the assembly of L5 and L11 on pre-60S ribosomal particles, enhance both the G1/S delay and the accumulation of free ribosomal protein L5. We propose the existence of a surveillance mechanism that couples the balanced production of yeast ribosomal components and cell cycle progression through the accumulation of free ribosomal proteins. This regulatory pathway resembles the p53-dependent nucleolar-stress checkpoint response described in human cells, which indicates that this is a general control strategy extended throughout eukaryotes. PMID:24043628

  2. 26 CFR 301.6223(g)-1 - Responsibilities of the tax matters partner.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    .... 301.6223(g)-1 Section 301.6223(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE....6223(g)-1 Responsibilities of the tax matters partner. (a) Notices described in section 6223(a)—(1... beginning on or after October 4, 2001. For years beginning prior to October 4, 2001, see § 301.6223(g)-1T...

  3. Preferential synthesis of the G1m(1) allotype of IgG1 in the central nervous system of multiple sclerosis patients.

    PubMed

    Salier, J P; Goust, J M; Pandey, J P; Fudenberg, H H

    1981-09-18

    Quantitations of the G1m(1) and G1m(3) allotypic determinants of human immunoglobulin G were performed by radioimmunoassay on cerebrospinal fluid and serum samples from patients with multiple sclerosis and from patients with other neurological disorders. In multiple sclerosis patients that were heterozygous for these determinants, G1m(1) concentration in the cerebrospinal fluid was greatly increased-reflected by an increased ratio of G1m(1)-in comparison with that of patients with other neurological disorders. These results suggest that in the heterozygous multiple sclerosis patients, most of the plasma cells in the central nervous system that secrete oligoclonal immunoglobulin G preferentially synthesize G1m(1) IgG1 molecules.

  4. FACT Prevents the Accumulation of Free Histones Evicted from Transcribed Chromatin and a Subsequent Cell Cycle Delay in G1

    PubMed Central

    Muñoz-Centeno, Mari Cruz; Singh, Rakesh Kumar; Oreal, Vincent; Reddy, Gajjalaiahvari Ugander; Liang, Dun; Géli, Vincent; Gunjan, Akash; Chávez, Sebastián

    2010-01-01

    The FACT complex participates in chromatin assembly and disassembly during transcription elongation. The yeast mutants affected in the SPT16 gene, which encodes one of the FACT subunits, alter the expression of G1 cyclins and exhibit defects in the G1/S transition. Here we show that the dysfunction of chromatin reassembly factors, like FACT or Spt6, down-regulates the expression of the gene encoding the cyclin that modulates the G1 length (CLN3) in START by specifically triggering the repression of its promoter. The G1 delay undergone by spt16 mutants is not mediated by the DNA–damage checkpoint, although the mutation of RAD53, which is otherwise involved in histone degradation, enhances the cell-cycle defects of spt16-197. We reveal how FACT dysfunction triggers an accumulation of free histones evicted from transcribed chromatin. This accumulation is enhanced in a rad53 background and leads to a delay in G1. Consistently, we show that the overexpression of histones in wild-type cells down-regulates CLN3 in START and causes a delay in G1. Our work shows that chromatin reassembly factors are essential players in controlling the free histones potentially released from transcribed chromatin and describes a new cell cycle phenomenon that allows cells to respond to excess histones before starting DNA replication. PMID:20502685

  5. Ischemic preconditioning, retinal neuroprotection and histone deacetylase activities.

    PubMed

    Fan, Jie; Alsarraf, Oday; Chou, C James; Yates, Phillip W; Goodwin, Nicole C; Rice, Dennis S; Crosson, Craig E

    2016-05-01

    Increased histone deacetylase (HDAC) activity and the resulting dysregulation of protein acetylation is an integral event in retinal degenerations associated with ischemia and ocular hypertension. This study investigates the role of preconditioning on the process of acetylation in ischemic retinal injury. Rat eyes were unilaterally subjected to retinal injury by 45 min of acute ischemia, and retinal neuroprotection induced by 5 min of an ischemic preconditioning (IPC) event. HDAC activity was evaluated by a fluorometric enzymatic assay with selective isoform inhibitors. Retinal localization of acetylated histone-H3 was determined by immunohistochemistry on retina cross sections. Cleaved caspase-3 level was evaluated by Western blots. Electroretinogram (ERG) analyses were used to assess differences in retinal function seven days following ischemic injury. In control eyes, analysis of HDAC isoforms demonstrated that HDAC1/2 accounted for 28.4 ± 1.6%, HDAC3 for 42.4 ± 1.5% and HDAC6 activity 27.3 ± 3.5% of total activity. Following ischemia, total Class-I HDAC activity increased by 21.2 ± 6.2%, and this increase resulted solely from a rise in HDAC1/2 activity. No change in HDAC3 activity was measured. Activity of Class-II HDACs and HDAC8 was negligible. IPC stimulus prior to ischemic injury also suppressed the rise in Class-I HDAC activity, cleaved caspase-3 levels, and increased acetylated histone-H3 in the retina. In control animals 7 days post ischemia, ERG a- and b-wave amplitudes were significantly reduced by 34.9 ± 3.1% and 42.4 ± 6.3%, respectively. In rats receiving an IPC stimulus, the ischemia-induced decline in ERG a- and b-wave amplitudes was blocked. Although multiple HDACs were detected in the retina, these studies provide evidence that hypoacetylation associated with ischemic injury results from the selective rise in HDAC1/2 activity and that neuroprotection induced by IPC is mediated in part by suppressing HDAC activity. Copyright

  6. Phosphorus-based SAHA analogues as histone deacetylase inhibitors.

    PubMed

    Kapustin, Galina V; Fejér, György; Gronlund, Jennifer L; McCafferty, Dewey G; Seto, Edward; Etzkorn, Felicia A

    2003-08-21

    [structure: see text] Three analogues of suberoyl anilide hydroxamic acid (SAHA) with phosphorus metal-chelating functionalities were synthesized as inhibitors of histone deacetylases (HDACs). The compounds showed weak activity for HeLa nuclear extracts (IC(50) = 0.57-6.1 mM), HDAC8 (IC(50) = 0.28-0.41 mM), and histone-deacetylase-like protein (HDLP, IC(50) = 0.33-1.9 mM), suggesting that the transition state of HDAC is not analogous to zinc proteases. Antiproliferative activity against A2780 cancer cells (IC(50) = 0.11-0.12 mM), comparable to SAHA (0.15 mM), was observed.

  7. F-box protein specificity for g1 cyclins is dictated by subcellular localization.

    PubMed

    Landry, Benjamin D; Doyle, John P; Toczyski, David P; Benanti, Jennifer A

    2012-01-01

    Levels of G1 cyclins fluctuate in response to environmental cues and couple mitotic signaling to cell cycle entry. The G1 cyclin Cln3 is a key regulator of cell size and cell cycle entry in budding yeast. Cln3 degradation is essential for proper cell cycle control; however, the mechanisms that control Cln3 degradation are largely unknown. Here we show that two SCF ubiquitin ligases, SCF(Cdc4) and SCF(Grr1), redundantly target Cln3 for degradation. While the F-box proteins (FBPs) Cdc4 and Grr1 were previously thought to target non-overlapping sets of substrates, we find that Cdc4 and Grr1 each bind to all 3 G1 cyclins in cell extracts, yet only Cln3 is redundantly targeted in vivo, due in part to its nuclear localization. The related cyclin Cln2 is cytoplasmic and exclusively targeted by Grr1. However, Cdc4 can interact with Cdk-phosphorylated Cln2 and target it for degradation when cytoplasmic Cdc4 localization is forced in vivo. These findings suggest that Cdc4 and Grr1 may share additional redundant targets and, consistent with this possibility, grr1Δ cdc4-1 cells demonstrate a CLN3-independent synergistic growth defect. Our findings demonstrate that structurally distinct FBPs are capable of interacting with some of the same substrates; however, in vivo specificity is achieved in part by subcellular localization. Additionally, the FBPs Cdc4 and Grr1 are partially redundant for proliferation and viability, likely sharing additional redundant substrates whose degradation is important for cell cycle progression.

  8. F-Box Protein Specificity for G1 Cyclins Is Dictated by Subcellular Localization

    PubMed Central

    Landry, Benjamin D.; Doyle, John P.; Toczyski, David P.; Benanti, Jennifer A.

    2012-01-01

    Levels of G1 cyclins fluctuate in response to environmental cues and couple mitotic signaling to cell cycle entry. The G1 cyclin Cln3 is a key regulator of cell size and cell cycle entry in budding yeast. Cln3 degradation is essential for proper cell cycle control; however, the mechanisms that control Cln3 degradation are largely unknown. Here we show that two SCF ubiquitin ligases, SCFCdc4 and SCFGrr1, redundantly target Cln3 for degradation. While the F-box proteins (FBPs) Cdc4 and Grr1 were previously thought to target non-overlapping sets of substrates, we find that Cdc4 and Grr1 each bind to all 3 G1 cyclins in cell extracts, yet only Cln3 is redundantly targeted in vivo, due in part to its nuclear localization. The related cyclin Cln2 is cytoplasmic and exclusively targeted by Grr1. However, Cdc4 can interact with Cdk-phosphorylated Cln2 and target it for degradation when cytoplasmic Cdc4 localization is forced in vivo. These findings suggest that Cdc4 and Grr1 may share additional redundant targets and, consistent with this possibility, grr1Δ cdc4-1 cells demonstrate a CLN3-independent synergistic growth defect. Our findings demonstrate that structurally distinct FBPs are capable of interacting with some of the same substrates; however, in vivo specificity is achieved in part by subcellular localization. Additionally, the FBPs Cdc4 and Grr1 are partially redundant for proliferation and viability, likely sharing additional redundant substrates whose degradation is important for cell cycle progression. PMID:22844257

  9. Histone Deacetylase 6 Regulates Bladder Architecture and Host Susceptibility to Uropathogenic Escherichia coli

    PubMed Central

    Lewis, Adam J.; Dhakal, Bijaya K.; Liu, Ting; Mulvey, Matthew A.

    2016-01-01

    Histone deacetylase 6 (HDAC6) is a non-canonical, mostly cytosolic histone deacetylase that has a variety of interacting partners and substrates. Previous work using cell-culture based assays coupled with pharmacological inhibitors and gene-silencing approaches indicated that HDAC6 promotes the actin- and microtubule-dependent invasion of host cells by uropathogenic Escherichia coli (UPEC). These facultative intracellular pathogens are the major cause of urinary tract infections. Here, we examined the involvement of HDAC6 in bladder colonization by UPEC using HDAC6 knockout mice. Though UPEC was unable to invade HDAC6−/− cells in culture, the bacteria had an enhanced ability to colonize the bladders of mice that lacked HDAC6. This effect was transient, and by six hours post-inoculation bacterial titers in the HDAC6−/− mice were reduced to levels seen in wild type control animals. Subsequent analyses revealed that the mutant mice had greater bladder volume capacity and fluid retention, along with much higher levels of acetylated α-tubulin. In addition, infiltrating neutrophils recovered from the HDAC6−/− bladder harbored significantly more viable bacteria than their wild type counterparts. Cumulatively, these changes may negate any inhibitory effects that the lack of HDAC6 has on UPEC entry into individual host cells, and suggest roles for HDAC6 in other urological disorders such as urinary retention. PMID:26907353

  10. Functional characterization of Candida albicans Hos2 histone deacetylase

    PubMed Central

    Karthikeyan, G; Paul-Satyaseela, Maneesh; Dhatchana Moorthy, Nachiappan; Gopalaswamy, Radha; Narayanan, Shridhar

    2014-01-01

    Candida albicans is a mucosal commensal organism capable of causing superficial (oral and vaginal thrush) infections in immune normal hosts, but is a major pathogen causing systemic and mucosal infections in immunocompromised individuals. Azoles have been very effective anti-fungal agents and the mainstay in treating opportunistic mold and yeast infections. Azole resistant strains have emerged compromising the utility of this class of drugs. It has been shown that azole resistance can be reversed by the co-administration of a histone deacetylase (HDAC) inhibitor, suggesting that resistance is mediated by epigenetic mechanisms possibly involving Hos2, a fungal deacetylase. We report here the cloning and functional characterization of  HOS2 (High Osmolarity  Sensitive) , a gene coding for fungal histone deacetylase from  C. albicans. Inhibition studies showed that Hos2 is susceptible to pan inhibitors such as trichostatin A (TSA) and suberoylanilide hydroxamic acid (SAHA), but is not inhibited by class I inhibitors such as MS-275. This  in  vitro enzymatic assay, which is amenable to high throughput could be used for screening potent fungal Hos2 inhibitors that could be a potential anti-fungal adjuvant. Purified Hos2 protein consistently deacetylated tubulins, rather than histones from TSA-treated cells. Hos2 has been reported to be a putative NAD+ dependent histone deacetylase, a feature of sirtuins. We assayed for sirtuin activation with resveratrol and purified Hos2 protein and did not find any sirtuin activity. PMID:25110576

  11. Divergent Specificity Development of IgG1 and IgG4 Autoantibodies in Endemic Pemphigus Foliaceus (Fogo Selvagem).

    PubMed

    Maldonado, Mike; Diaz, Luis A; Prisayanh, Phillip; Yang, Jinsheng; Qaqish, Bahjat F; Aoki, Valeria; Hans-Filho, Gunter; Rivitti, Evandro A; Culton, Donna A; Qian, Ye

    2017-08-01

    We have shown that although the IgG response in fogo selvagem (FS) is mainly restricted to desmoglein (Dsg) 1, other keratinocyte cadherins are also targeted by FS patients and healthy control subjects living in the endemic region of Limão Verde, Brazil (endemic controls). Evaluating nonpathogenic IgG1 and pathogenic IgG4 subclass responses to desmosomal proteins may reveal important differences between pathogenic and nonpathogenic responses, and how these differences relate to the pathogenic IgG4 response and resultant FS. In this study, we tested by ELISA >100 sera from each FS patient, endemic control, and nonendemic control for IgG1 and IgG4 autoantibodies to keratinocyte cadherins besides Dsg1. IgG1 and IgG4 subclass responses in endemic controls are highly correlated between Dsg1 and other keratinocyte cadherins. This correlation persists in the IgG1 response among FS patients, but diminishes in IgG4 response, suggesting that IgG1 binds highly conserved linear epitopes among cadherins, whereas IgG4 binds mainly specific conformational epitopes on Dsg1. A confirmatory test comparing serum samples of 11 individuals before and after their FS onset substantiated our findings that IgG1 recognizes primarily linear epitopes on Dsg1 both before and after disease onset, whereas IgG4 recognizes primarily linear epitopes before disease onset, but recognizes more conformational epitopes on Dsg1 after the onset of disease. This study may provide a mechanism by which a specificity convergence of the IgG4 response to unique Dsg1 epitopes, most likely conformational pathogenic epitopes, leads to the onset of FS disease.

  12. Divergent Specificity Development of IgG1 and IgG4 Autoantibodies in Endemic Pemphigus Foliaceus (Fogo Selvagem)

    PubMed Central

    Maldonado, Mike; Diaz, Luis A.; Prisayanh, Phillip; Yang, Jinsheng; Qaqish, Bahjat F.; Aoki, Valeria; Hans-Filho, Gunter; Rivitti, Evandro A.; Culton, Donna A.; Qian, Ye

    2017-01-01

    We have shown that although the IgG response in fogo selvagem (FS) is mainly restricted to desmoglein (Dsg) 1, other keratinocyte cadherins are also targeted by FS patients and healthy control subjects living in the endemic region of Limão Verde, Brazil (endemic controls). Evaluating nonpathogenic IgG1 and pathogenic IgG4 subclass responses to desmosomal proteins may reveal important differences between pathogenic and nonpathogenic responses, and how these differences relate to the pathogenic IgG4 response and resultant FS. In this study, we tested by ELISA >100 sera from each FS patient, endemic control, and nonendemic control for IgG1 and IgG4 autoantibodies to keratinocyte cadherins besides Dsg1. IgG1 and IgG4 subclass responses in endemic controls are highly correlated between Dsg1 and other keratinocyte cadherins. This correlation persists in the IgG1 response among FS patients, but diminishes in IgG4 response, suggesting that IgG1 binds highly conserved linear epitopes among cadherins, whereas IgG4 binds mainly specific conformational epitopes on Dsg1. A confirmatory test comparing serum samples of 11 individuals before and after their FS onset substantiated our findings that IgG1 recognizes primarily linear epitopes on Dsg1 both before and after disease onset, whereas IgG4 recognizes primarily linear epitopes before disease onset, but recognizes more conformational epitopes on Dsg1 after the onset of disease. This study may provide a mechanism by which a specificity convergence of the IgG4 response to unique Dsg1 epitopes, most likely conformational pathogenic epitopes, leads to the onset of FS disease. PMID:28868524

  13. Chemical origins of isoform selectivity in histone deacetylase inhibitors.

    PubMed

    Butler, Kyle V; Kozikowski, Alan P

    2008-01-01

    Histones undergo extensive posttranslational modifications that affect gene expression. Acetylation is a key histone modification that is primarily regulated by two enzymes, one of which is histone deacetylase (HDAC). The activity of HDAC causes transcriptional silencing of DNA. Eleven distinct zinc-dependent histone deacetylase isoforms have been identified in humans. Each isoform has a unique structure and function, and regulates a unique set of genes. HDAC is responsible for the regulation of many genes involved in cancer cell proliferation, and it has been implicated in the pathogenesis of many neurological conditions. HDAC inhibitors are known to be very effective anti-cancer agents, and research has shown them to be potential treatments for many other conditions. Histone deacetylase inhibitors modify the expression of many genes, and it is possible that inhibition of one isoform could cause epigenetic changes that are beneficial to treatment of a disease, while inhibition of another isoform could cause contradictory changes. Selective HDAC inhibitors will be better able to avoid these types of situations than non-specific inhibitors, and may also be less toxic than pan-HDAC inhibitors. Many potent pan-HDAC inhibitors have already been developed, leaving the development of selective inhibitors at the forefront of HDAC drug development. Certain structural moieties may be added to HDAC inhibitors to give isoform selectivity, and these will be discussed in this review. This review will focus on the applications of selective HDAC inhibitors, inhibitors reported to show selectivity, and the relationship between inhibitor structure and selectivity.

  14. Activation of p53 Transcriptional Activity by SMRT: a Histone Deacetylase 3-Independent Function of a Transcriptional Corepressor

    PubMed Central

    Adikesavan, Anbu Karani; Karmakar, Sudipan; Pardo, Patricia; Wang, Liguo; Liu, Shuang; Li, Wei

    2014-01-01

    The silencing mediator of retinoic acid and thyroid hormone receptors (SMRT) is an established histone deacetylase 3 (HDAC3)-dependent transcriptional corepressor. Microarray analyses of MCF-7 cells transfected with control or SMRT small interfering RNA revealed SMRT regulation of genes involved in DNA damage responses, and the levels of the DNA damage marker γH2AX as well as poly(ADP-ribose) polymerase cleavage were elevated in SMRT-depleted cells treated with doxorubicin. A number of these genes are established p53 targets. SMRT knockdown decreased the activity of two p53-dependent reporter genes as well as the expression of p53 target genes, such as CDKN1A (which encodes p21). SMRT bound directly to p53 and was recruited to p53 binding sites within the p21 promoter. Depletion of GPS2 and TBL1, components of the SMRT corepressor complex, but not histone deacetylase 3 (HDAC3) decreased p21-luciferase activity. p53 bound to the SMRT deacetylase activation domain (DAD), which mediates HDAC3 binding and activation, and HDAC3 could attenuate p53 binding to the DAD region of SMRT. Moreover, an HDAC3 binding-deficient SMRT DAD mutant coactivated p53 transcriptional activity. Collectively, these data highlight a biological role for SMRT in mediating DNA damage responses and suggest a model where p53 binding to the DAD limits HDAC3 interaction with this coregulator, thereby facilitating SMRT coactivation of p53-dependent gene expression. PMID:24449765

  15. High Dub3 expression in mouse ESCs couples the G1/S checkpoint to pluripotency.

    PubMed

    van der Laan, Siem; Tsanov, Nikolay; Crozet, Carole; Maiorano, Domenico

    2013-11-07

    The molecular mechanism underlying G1/S checkpoint bypass in mouse embryonic stem cells (ESCs) remains unknown. DNA damage blocks S phase entry by inhibiting the CDK2 kinase through destruction of its activator, the Cdc25A phosphatase. We observed high Cdc25A levels in G1 that persist even after DNA damage in mouse ESCs. We also found higher expression of Dub3, a deubiquitylase that controls Cdc25A protein abundance. Moreover, we demonstrate that the Dub3 gene is a direct target of Esrrb, a key transcription factor of the self-renewal machinery. We show that Dub3 expression is strongly downregulated during neural conversion and precedes Cdc25A destabilization, while forced Dub3 expression in ESCs becomes lethal upon differentiation, concomitant to cell-cycle remodeling and lineage commitment. Finally, knockdown of either Dub3 or Cdc25A induced spontaneous differentiation of ESCs. Altogether, these findings couple the self-renewal machinery to cell-cycle control through a deubiquitylase in ESCs.

  16. Prostate Cancer Prevention by Sulforaphane, a Novel Dietary Histone Deacetylase Inhibitor

    DTIC Science & Technology

    2008-01-01

    sulforaphane , a novel dietary histone deacetylase inhibitor PRINCIPAL INVESTIGATOR: Yu Zhen CONTRACTING ORGANIZATION: Oregon State...ANNUAL 3. DATES COVERED 1 JAN 2007 - 31 DEC 2007 4. TITLE AND SUBTITLE Prostate cancer prevention by sulforaphane , a novel dietary histone deacetylase...Prostate cancer is the second leading cause of cancer related death in men. To test Sulforaphane (SFN) as a novel histone deacetylases (HDAC) inhibitor

  17. The G1 restriction point as critical regulator of neocortical neuronogenesis

    NASA Technical Reports Server (NTRS)

    Caviness, V. S. Jr; Takahashi, T.; Nowakowski, R. S.

    1999-01-01

    Neuronogenesis in the pseudostratified ventricular epithelium is the initial process in a succession of histogenetic events which give rise to the laminate neocortex. Here we review experimental findings in mouse which support the thesis that the restriction point of the G1 phase of the cell cycle is the critical point of regulation of the overall neuronogenetic process. The neuronogenetic interval in mouse spans 6 days. In the course of these 6 days the founder population and its progeny execute 11 cell cycles. With each successive cycle there is an increase in the fraction of postmitotic cells which leaves the cycle (the Q fraction) and also an increase in the length of the cell cycle due to an increase in the length of the G1 phase of the cycle. Q corresponds to the probability that postmitotic cells will exit the cycle at the restriction point of the G1 phase of the cell cycle. Q increases non-linearly, but the rate of change of Q with cycle (i.e., the first derivative) over the course of the neuronogenetic interval is a constant, k, which appears to be set principally by cell internal mechanisms which are species specific. Q also seems to be modulated, but at low amplitude, by a balance of mitogenic and antimitogenic influences acting from without the cell. We suggest that intracellular signal transduction systems control a general advance of Q during development and thereby determine the general developmental plan (i.e., cell number and laminar composition) of the neocortex and that external mitogens and anti-mitogens modulate this advance regionally and temporally and thereby produce regional modifications of the general plan.

  18. The G1 restriction point as critical regulator of neocortical neuronogenesis

    NASA Technical Reports Server (NTRS)

    Caviness, V. S. Jr; Takahashi, T.; Nowakowski, R. S.

    1999-01-01

    Neuronogenesis in the pseudostratified ventricular epithelium is the initial process in a succession of histogenetic events which give rise to the laminate neocortex. Here we review experimental findings in mouse which support the thesis that the restriction point of the G1 phase of the cell cycle is the critical point of regulation of the overall neuronogenetic process. The neuronogenetic interval in mouse spans 6 days. In the course of these 6 days the founder population and its progeny execute 11 cell cycles. With each successive cycle there is an increase in the fraction of postmitotic cells which leaves the cycle (the Q fraction) and also an increase in the length of the cell cycle due to an increase in the length of the G1 phase of the cycle. Q corresponds to the probability that postmitotic cells will exit the cycle at the restriction point of the G1 phase of the cell cycle. Q increases non-linearly, but the rate of change of Q with cycle (i.e., the first derivative) over the course of the neuronogenetic interval is a constant, k, which appears to be set principally by cell internal mechanisms which are species specific. Q also seems to be modulated, but at low amplitude, by a balance of mitogenic and antimitogenic influences acting from without the cell. We suggest that intracellular signal transduction systems control a general advance of Q during development and thereby determine the general developmental plan (i.e., cell number and laminar composition) of the neocortex and that external mitogens and anti-mitogens modulate this advance regionally and temporally and thereby produce regional modifications of the general plan.

  19. Recruitment of Cdc28 by Whi3 restricts nuclear accumulation of the G1 cyclin–Cdk complex to late G1

    PubMed Central

    Wang, Hongyin; Garí, Eloi; Vergés, Emili; Gallego, Carme; Aldea, Martí

    2004-01-01

    The G1 cyclin Cln3 is a key activator of cell-cycle entry in budding yeast. Here we show that Whi3, a negative G1 regulator of Cln3, interacts in vivo with the cyclin-dependent kinase Cdc28 and regulates its localization in the cell. Efficient interaction with Cdc28 depends on an N-terminal domain of Whi3 that is also required for cytoplasmic localization of Cdc28, and for proper regulation of G1 length and filamentous growth. On the other hand, nuclear accumulation of Cdc28 requires the nuclear localization signal of Cln3, which is also found in Whi3 complexes. Both Cln3 and Cdc28 are mainly cytoplasmic during early G1, and become nuclear in late G1. However, Whi3-deficient cells show a distinct nuclear accumulation of Cln3 and Cdc28 already in early G1. We propose that Whi3 constitutes a cytoplasmic retention device for Cln3–Cdc28 complexes, thus defining a key G1 event in yeast cells. PMID:14685274

  20. Recruitment of Cdc28 by Whi3 restricts nuclear accumulation of the G1 cyclin-Cdk complex to late G1.

    PubMed

    Wang, Hongyin; Garí, Eloi; Vergés, Emili; Gallego, Carme; Aldea, Martí

    2004-01-14

    The G1 cyclin Cln3 is a key activator of cell-cycle entry in budding yeast. Here we show that Whi3, a negative G1 regulator of Cln3, interacts in vivo with the cyclin-dependent kinase Cdc28 and regulates its localization in the cell. Efficient interaction with Cdc28 depends on an N-terminal domain of Whi3 that is also required for cytoplasmic localization of Cdc28, and for proper regulation of G1 length and filamentous growth. On the other hand, nuclear accumulation of Cdc28 requires the nuclear localization signal of Cln3, which is also found in Whi3 complexes. Both Cln3 and Cdc28 are mainly cytoplasmic during early G1, and become nuclear in late G1. However, Whi3-deficient cells show a distinct nuclear accumulation of Cln3 and Cdc28 already in early G1. We propose that Whi3 constitutes a cytoplasmic retention device for Cln3-Cdc28 complexes, thus defining a key G1 event in yeast cells.

  1. Analysis of the G1 arrest position of senescent WI38 cells by quinacrine dihydrochloride nuclear fluorescence: evidence for a late G1 arrest

    SciTech Connect

    Gorman, S.D.; Cristofalo, V.J.

    1986-11-01

    Senescence of the human diploid fibroblast-like cell line, W138, is characterized by a loss of proliferative activity and an arrest of cells with a 2C DNA content (G1 or G0). To examine the specific region within G1 in which senescent cells arrest, senescent cells were stained with quinacrine dihydrochloride (QDH) and their nuclear fluorescence was compared with that of young cultures arrested in early and late G1 by serum deprivation and hydroxyurea exposure, respectively. Release of these G1-arrested young cultures from their blocking conditions and timing the kinetics of their entry into the S phase by autoradiographic detection of (/sup 3/H)thymidine incorporation revealed that serum-deprived cells entered the S phase within 15-18h, whereas hydroxyurea-exposed cells entered the S phase within 1.5h, thus confirming their relative G1-arrest positions. QDH-stained, serum-deprived and hydroxyurea-exposed young cells exhibited relative nuclear fluorescence intensities of 51.7 and 23.9, respectively. Senescent cells exhibited a relative nuclear fluorescence intensity of 17.4, closely resembling the fluorescence of young cultures arrested in late G1 by hydroxyurea exposure. These data support the concept that senescent cells are arrested from further progression in the cell cycle in late G1.

  2. 26 CFR 1.904(g)-1 - Overall domestic loss and the overall domestic loss account.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 9 2011-04-01 2011-04-01 false Overall domestic loss and the overall domestic loss account. 1.904(g)-1 Section 1.904(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF... the United States § 1.904(g)-1 Overall domestic loss and the overall domestic loss account....

  3. 26 CFR 1.904(g)-1 - Overall domestic loss and the overall domestic loss account.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 9 2010-04-01 2010-04-01 false Overall domestic loss and the overall domestic loss account. 1.904(g)-1 Section 1.904(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF... States § 1.904(g)-1 Overall domestic loss and the overall domestic loss account. For further...

  4. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 16 Commercial Practices 1 2014-01-01 2014-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305... RULEâ) Appendix G1 to Part 305—Furnaces—Gas Furnace type Range of annual fuel utilization efficiencies (AFUEs) Low High Gas Furnaces Manufactured Before the Compliance Date of DOE Regional...

  5. 26 CFR 1.665(g)-1A - Capital gain distribution.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 8 2010-04-01 2010-04-01 false Capital gain distribution. 1.665(g)-1A Section 1.665(g)-1A Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX... Or After January 1, 1969 § 1.665(g)-1A Capital gain distribution. For any taxable year of a trust...

  6. 26 CFR 1.430(g)-1 - Valuation date and valuation of plan assets.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ....430(g)-1 Section 1.430(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Certain Stock Options § 1.430(g)-1 Valuation date and... valuation date and the valuation of a plan's assets for a plan year under section 430(g). Section 430 and...

  7. 26 CFR 1.402(g)-1 - Limitation on exclusion for elective deferrals.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    .... 1.402(g)-1 Section 1.402(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Pension, Profit-Sharing, Stock Bonus Plans, Etc. § 1.402(g)-1... section 402(a)(8) (before applying the limits of section 402(g) or this section). (2) Any employer...

  8. 26 CFR 25.2523(g)-1 - Special rule for charitable remainder trusts.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    .... 25.2523(g)-1 Section 25.2523(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE....2523(g)-1 Special rule for charitable remainder trusts. (a) In general. (1) With respect to gifts made... passing to the spouse qualifies for a marital deduction under section 2523(g) and the value of the...

  9. 26 CFR 1.143(g)-1 - Requirements related to arbitrage.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 2 2010-04-01 2010-04-01 false Requirements related to arbitrage. 1.143(g)-1 Section 1.143(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED....143(g)-1 Requirements related to arbitrage. (a) In general. Under section 143, for an issue to be an...

  10. 26 CFR 1.404(g)-1 - Deduction of employer liability payments.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ...(g)-1 Section 1.404(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Pension, Profit-Sharing, Stock Bonus Plans, Etc. § 1.404(g)-1... employer. (c) Limitations, etc.—(1) Permissible expenses. A payment shall be deductible under section 404(g...

  11. 26 CFR 1.414(g)-1 - Definition of plan administrator.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 5 2010-04-01 2010-04-01 false Definition of plan administrator. 1.414(g)-1 Section 1.414(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Pension, Profit-Sharing, Stock Bonus Plans, Etc. § 1.414(g)-1...

  12. 26 CFR 1.415(g)-1 - Disqualification of plans and trusts.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 5 2010-04-01 2010-04-01 false Disqualification of plans and trusts. 1.415(g)-1 Section 1.415(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Pension, Profit-Sharing, Stock Bonus Plans, Etc. § 1.415(g)-1...

  13. 26 CFR 1.642(g)-1 - Disallowance of double deductions; in general.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ....642(g)-1 Section 1.642(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Estates, Trusts, and Beneficiaries § 1.642(g)-1 Disallowance..., after a statement is filed under section 642(g) with respect to a particular item or portion of an item...

  14. 26 CFR 301.6323(g)-1 - Refiling of notice of tax lien.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 18 2010-04-01 2010-04-01 false Refiling of notice of tax lien. 301.6323(g)-1 Section 301.6323(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) PROCEDURE AND ADMINISTRATION PROCEDURE AND ADMINISTRATION Collection General Provisions § 301.6323(g)-1...

  15. 26 CFR 301.6511(g)-1 - Special rule for partnership items of federally registered partnerships.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... registered partnerships. 301.6511(g)-1 Section 301.6511(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... Limitations on Assessment and Collection § 301.6511(g)-1 Special rule for partnership items of federally...(g) must also be taken into account in applying the various special periods of limitation...

  16. 26 CFR 301.6511(g)-1 - Special rule for partnership items of federally registered partnerships.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 18 2011-04-01 2011-04-01 false Special rule for partnership items of federally registered partnerships. 301.6511(g)-1 Section 301.6511(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... Limitations on Assessment and Collection § 301.6511(g)-1 Special rule for partnership items of...

  17. 26 CFR 301.6511(g)-1 - Special rule for partnership items of federally registered partnerships.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 26 Internal Revenue 18 2012-04-01 2012-04-01 false Special rule for partnership items of federally registered partnerships. 301.6511(g)-1 Section 301.6511(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... Limitations on Assessment and Collection § 301.6511(g)-1 Special rule for partnership items of...

  18. 26 CFR 301.6511(g)-1 - Special rule for partnership items of federally registered partnerships.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 26 Internal Revenue 18 2013-04-01 2013-04-01 false Special rule for partnership items of federally registered partnerships. 301.6511(g)-1 Section 301.6511(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... Limitations on Assessment and Collection § 301.6511(g)-1 Special rule for partnership items of...

  19. 26 CFR 301.6511(g)-1 - Special rule for partnership items of federally registered partnerships.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 26 Internal Revenue 18 2014-04-01 2014-04-01 false Special rule for partnership items of federally registered partnerships. 301.6511(g)-1 Section 301.6511(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... Limitations on Assessment and Collection § 301.6511(g)-1 Special rule for partnership items of...

  20. Mitotic phosphorylation of eukaryotic initiation factor 4G1 (eIF4G1) at Ser1232 by Cdk1:cyclin B inhibits eIF4A helicase complex binding with RNA.

    PubMed

    Dobrikov, Mikhail I; Shveygert, Mayya; Brown, Michael C; Gromeier, Matthias

    2014-02-01

    During mitosis, global translation is suppressed, while synthesis of proteins with vital mitotic roles must go on. Prior evidence suggests that the mitotic translation shift involves control of initiation. Yet, no signals specifically targeting translation initiation factors during mitosis have been identified. We used phosphoproteomics to investigate the central translation initiation scaffold and "ribosome adaptor," eukaryotic initiation factor 4G1 (eIF4G1) in interphase or nocodazole-arrested mitotic cells. This approach and kinase inhibition assays, in vitro phosphorylation with recombinant kinase, and kinase depletion-reconstitution experiments revealed that Ser1232 in eIF4G1 is phosphorylated by cyclin-dependent kinase 1 (Cdk1):cyclin B during mitosis. Ser1232 is located in an unstructured region of the C-terminal portion of eIF4G1 that coordinates assembly of the eIF4G/-4A/-4B helicase complex and binding of the mitogen-activated protein kinase (MAPK) signal-integrating kinase, Mnk. Intense phosphorylation of Ser1232 in mitosis strongly enhanced the interactions of eIF4A with HEAT domain 2 of eIF4G and decreased association of eIF4G/-4A with RNA. Our findings implicate phosphorylation of eIF4G1(Ser1232) by Cdk1:cyclin B and its inhibitory effects on eIF4A helicase activity in the mitotic translation initiation shift.

  1. Fucose depletion from human IgG1 oligosaccharide enhances binding enthalpy and association rate between IgG1 and FcgammaRIIIa.

    PubMed

    Okazaki, Akira; Shoji-Hosaka, Emi; Nakamura, Kazuyasu; Wakitani, Masako; Uchida, Kazuhisa; Kakita, Shingo; Tsumoto, Kouhei; Kumagai, Izumi; Shitara, Kenya

    2004-03-05

    Depletion of fucose from human IgG1 oligosaccharide improves its affinity for Fcgamma receptor IIIa (FcgammaRIIIa). This is the first case where a glycoform modification is shown to improve glycoprotein affinity for the receptors without carbohydrate-binding capacity, suggesting a novel glyco-engineering strategy to improve ligand-receptor binding. To address the mechanisms of affinity improvement by the fucose depletion, we used isothermal titration calorimetry (ITC) and biosensor analysis with surface plasmon resonance. ITC demonstrated that IgG1-FcgammaRIIIa binding was driven by favorable binding enthalpy (DeltaH) but opposed by unfavorable binding entropy change (DeltaS). Fucose depletion from IgG1 enhanced the favorable DeltaH, leading to the increase in the binding constant of IgG1 for the receptor by a factor of 20-30. The increase in the affinity was mainly attributed to an enhanced association rate. A triple amino acid substitution in IgG1, S298A/E333A/K334A, is also known to improve IgG1 affinity for FcgammaRIIIa. ITC demonstrated that the amino acid substitution attenuated the unfavorable DeltaS resulting in a three- to fourfold increase in the binding constant. The affinity enhancement by the amino acid substitution was due to a reduced dissociation rate. These results indicate that the mechanism of affinity improvement by the fucose depletion is quite distinct from that by the amino acid substitution. Defucosylated IgG1 exhibited higher antibody-dependent cellular cytotoxicity (ADCC) than S298A/E333A/K334A-IgG1, showing a correlation between IgG1 affinity for FcgammaRIIIa and ADCC. We also examined the effect of FcgammaRIIIa polymorphism (Val158/Phe158) on IgG1-FcgammaRIIIa binding. The Phe to Val substitution increased FcgammaRIIIa affinity for IgG1 in an enthalpy-driven manner with the reduced dissociation rate. These results together highlight the distinctive functional improvement of affinity by IgG1 defucosylation and suggest that engineering of

  2. Revised direct radiocarbon dating of the Vindija G1 Upper Paleolithic Neandertals.

    PubMed

    Higham, Tom; Ramsey, Christopher Bronk; Karavanić, Ivor; Smith, Fred H; Trinkaus, Erik

    2006-01-17

    The 1998/1999 direct dating of two Neandertal specimens from level G(1) of Vindija Cave in Croatia to approximately 28,000 and approximately 29,000 radiocarbon ((14)C) years ago has led to interpretations concerning the late survival of Neandertals in south-central Europe, patterns of interaction between Neandertals and in-dispersing early modern humans in Europe, and complex biocultural scenarios for the earlier phases of the Upper Paleolithic. Given improvements, particularly in sample pretreatment techniques for bone radiocarbon samples, especially ultrafiltration of collagen samples, these Vindija G(1) Neandertal fossils are redated to approximately 32,000-33,000 (14)C years ago and possibly earlier. These results and the recent redating of a number of purportedly old modern human skeletal remains in Europe to younger time periods highlight the importance of fine chronological control when studying this biocultural time period and the tenuous nature of monolithic scenarios for the establishment of modern humans and earlier phases of the Upper Paleolithic in Europe.

  3. Revised direct radiocarbon dating of the Vindija G1 Upper Paleolithic Neandertals

    PubMed Central

    Higham, Tom; Ramsey, Christopher Bronk; Karavanić, Ivor; Smith, Fred H.; Trinkaus, Erik

    2006-01-01

    The 1998/1999 direct dating of two Neandertal specimens from level G1 of Vindija Cave in Croatia to ≈28,000 and ≈29,000 radiocarbon (14C) years ago has led to interpretations concerning the late survival of Neandertals in south-central Europe, patterns of interaction between Neandertals and in-dispersing early modern humans in Europe, and complex biocultural scenarios for the earlier phases of the Upper Paleolithic. Given improvements, particularly in sample pretreatment techniques for bone radiocarbon samples, especially ultrafiltration of collagen samples, these Vindija G1 Neandertal fossils are redated to ≈32,000–33,000 14C years ago and possibly earlier. These results and the recent redating of a number of purportedly old modern human skeletal remains in Europe to younger time periods highlight the importance of fine chronological control when studying this biocultural time period and the tenuous nature of monolithic scenarios for the establishment of modern humans and earlier phases of the Upper Paleolithic in Europe. PMID:16407102

  4. Mid1p-dependent regulation of the M-G1 transcription wave in fission yeast.

    PubMed

    Agarwal, Monica; Papadopoulou, Kyriaki; Mayeux, Adeline; Vajrala, Vasanthi; Quintana, Daniela M; Paoletti, Anne; McInerny, Christopher J

    2010-12-15

    The control of gene expression at certain times during the mitotic cell division cycle is a common feature in eukaryotes. In fission yeast, at least five waves of gene expression have been described, with one transcribed at the M-G1 interval under the control of the PBF transcription factor complex. PBF consists of at least three transcription factors, two forkhead-like proteins Sep1p and Fkh2p, and a MADS box-like protein Mbx1p, and binds to PCB motifs found in the gene promoters. Mbx1p is under the direct control of the polo-like kinase Plo1p and the Cdc14p-like phosphatase Clp1p (Flp1p). Here, we show that M-G1 gene expression in fission yeast is also regulated by the anillin-like protein, Mid1p (Dmf1p). Mid1p binds in vivo to both Fkh2p and Sep1p, and to the promoter regions of M-G1 transcribed genes. Mid1p promoter binding is dependent on Fkh2p, Plo1p and Clp1p. The absence of mid1(+) in cells results in partial loss of M-G1 specific gene expression, suggesting that it has a negative role in controlling gene expression. This phenotype is exacerbated by also removing clp1(+), suggesting that Mid1p and Clp1p have overlapping functions in controlling transcription. As mid1(+) is itself expressed at M-G1, these observations offer a new mechanism whereby Mid1p contributes to controlling cell cycle-specific gene expression as part of a feedback loop.

  5. Donor cells at the G1 phase enhance homogeneous gene expression among blastomeres in bovine somatic cell nuclear transfer embryos.

    PubMed

    Iwamoto, Daisaku; Kasamatsu, Aya; Ideta, Atsushi; Urakawa, Manami; Matsumoto, Kazuya; Hosoi, Yoshihiko; Iritani, Akira; Aoyagi, Yoshito; Saeki, Kazuhiro

    2012-02-01

    The success rate of bovine somatic cell nuclear transfer (SCNT) embryos to full term has been reported to be higher with G1 cells than with G0 cells. To better understand the reason for this, we analyzed the kinetics of luminescence activity in bovine SCNT embryos from G0 and G1 cells carrying a luciferase gene under the control of the β-actin promoter during early embryonic development. At 60-h postfusion, when bovine embryonic gene activation (EGA) begins, the luminescence activity was higher in G1-SCNT embryos than G0-SCNT embryos. Moreover, half of the G1-SCNT embryos exhibited homogeneous luminescence among the blastomeres, whereas more than half of the G0-SCNT embryos exhibited mosaic luminescence. To characterize the differential luminescence pattern in SCNT embryos, the expressions of several endogenous genes and the level of DNA methylation were determined in all blastomeres of SCNT embryos with or without luminescence. The expressions of several development-related genes (H2AFZ, GJA1, and BAX) and level of DNA methylation of the SCNT embryos with luminescence were the same as those of normal embryos produced by in vitro fertilization. A higher success rate in G1-SCNT embryos is thought to contribute to homogeneous expression among all blastomeres at EGA.

  6. Histone deacetylase 9 is a negative regulator of adipogenic differentiation.

    PubMed

    Chatterjee, Tapan K; Idelman, Gila; Blanco, Victor; Blomkalns, Andra L; Piegore, Mark G; Weintraub, Daniel S; Kumar, Santosh; Rajsheker, Srinivas; Manka, David; Rudich, Steven M; Tang, Yaoliang; Hui, David Y; Bassel-Duby, Rhonda; Olson, Eric N; Lingrel, Jerry B; Ho, Shuk-Mei; Weintraub, Neal L

    2011-08-05

    Differentiation of preadipocytes into mature adipocytes capable of efficiently storing lipids is an important regulatory mechanism in obesity. Here, we examined the involvement of histone deacetylases (HDACs) and histone acetyltransferases (HATs) in the regulation of adipogenesis. We find that among the various members of the HDAC and HAT families, only HDAC9 exhibited dramatic down-regulation preceding adipogenic differentiation. Preadipocytes from HDAC9 gene knock-out mice exhibited accelerated adipogenic differentiation, whereas HDAC9 overexpression in 3T3-L1 preadipocytes suppressed adipogenic differentiation, demonstrating its direct role as a negative regulator of adipogenesis. HDAC9 expression was higher in visceral as compared with subcutaneous preadipocytes, negatively correlating with their potential to undergo adipogenic differentiation in vitro. HDAC9 localized in the nucleus, and its negative regulation of adipogenesis segregates with the N-terminal nuclear targeting domain, whereas the C-terminal deacetylase domain is dispensable for this function. HDAC9 co-precipitates with USF1 and is recruited with USF1 at the E-box region of the C/EBPα gene promoter in preadipocytes. Upon induction of adipogenic differentiation, HDAC9 is down-regulated, leading to its dissociation from the USF1 complex, whereas p300 HAT is up-regulated to allow its association with USF1 and accumulation at the E-box site of the C/EBPα promoter in differentiated adipocytes. This reciprocal regulation of HDAC9 and p300 HAT in the USF1 complex is associated with increased C/EBPα expression, a master regulator of adipogenic differentiation. These findings provide new insights into mechanisms of adipogenic differentiation and document a critical regulatory role for HDAC9 in adipogenic differentiation through a deacetylase-independent mechanism.

  7. Inhibitors of Histone Deacetylases Attenuate Noise-Induced Hearing Loss.

    PubMed

    Chen, Jun; Hill, Kayla; Sha, Su-Hua

    2016-08-01

    Loss of auditory sensory hair cells is the major pathological feature of noise-induced hearing loss (NIHL). Currently, no established clinical therapies for prevention or amelioration of NIHL are available. The absence of treatments is due to our lack of a comprehensive understanding of the molecular mechanisms underlying noise-induced damage. Our previous study indicates that epigenetic modification of histones alters hair cell survival. In this study, we investigated the effect of noise exposure on histone H3 lysine 9 acetylation (H3K9ac) in the inner ear of adult CBA/J mice and determined if inhibition of histone deacetylases by systemic administration of suberoylanilide hydroxamic acid (SAHA) could attenuate NIHL. Our results showed that H3K9ac was decreased in the nuclei of outer hair cells (OHCs) and marginal cells of the stria vascularis in the basal region after exposure to a traumatic noise paradigm known to induce permanent threshold shifts (PTS). Consistent with these results, levels of histone deacetylases 1, 2, and 3 (HDAC1, HDAC2 and HDAC3) were increased predominately in the nuclei of cochlear cells. Silencing of HDAC1, HDAC2, or HDAC3 with siRNA reduced the expression of the target HDAC in OHCs, but did not attenuate noise-induced PTS, whereas treatment with the pan-HDAC inhibitor SAHA, also named vorinostat, reduced OHC loss, and attenuated PTS. These findings suggest that histone acetylation is involved in the pathogenesis of noise-induced OHC death and hearing loss. Pharmacological targeting of histone deacetylases may afford a strategy for protection against NIHL.

  8. The potential of histone deacetylase inhibitors in lung cancer.

    PubMed

    Aparicio, Ana

    2006-03-01

    In the nucleus, DNA is wrapped around octamers of histone proteins. Histones, like other proteins, are posttranslationally modified by the addition of an array of chemical groups that affect their interactions with surrounding structures. Histone acetyltransferases and histone deacetylases (HDACs) are the enzymes involved in the addition and removal, respectively, of acetyl groups from the aminoterminal tails of histones. A number of structurally diverse compounds are capable of inhibiting HDACs and exert a variety of biologic effects on cancer cells in preclinical models. Early clinical trials with the first generation of HDAC inhibitors (HDACIs) have demonstrated promising therapeutic activity, and HDACs have become one of the hottest targets in drug development today.

  9. Design and synthesis of a potent histone deacetylase inhibitor.

    PubMed

    Liu, Tao; Kapustin, Galina; Etzkorn, Felicia A

    2007-05-03

    Histone deacetylase (HDAC) inhibitors have potential for cancer therapy. An HDAC inhibitor based on a cyclic peptide mimic of known structure, linked by an aliphatic chain to a hydroxamic acid, was designed and synthesized. The chimeric compound showed potent competitive inhibition of nuclear HDACs, with an IC50 value of 46 nM and a Ki value of 13.7 nM. The designed inhibitor showed 4-fold selectivity for HDAC1 (57 nM) over HDAC8 (231 nM).

  10. A non-isotopic assay for histone deacetylase activity.

    PubMed

    Hoffmann, K; Brosch, G; Loidl, P; Jung, M

    1999-05-01

    Inhibitors of histone deacetylase (HD) bear great potential as new drugs due to their ability to modulate transcription and to induce apoptosis or differentiation in cancer cells. To study the activity of HD and the effect of potential inhibitors in vitro so far only radio-active assays have existed. For the search of new inhibitors and for the use in HD identification and purification we established a simple, non-radioactive assay that allows screening of large numbers of compounds. The assay is based on an aminocoumarin derivative of an Omega-acetylated lysine as enzyme substrate.

  11. Histone deacetylase inhibitor activity in royal jelly might facilitate caste switching in bees.

    PubMed

    Spannhoff, Astrid; Kim, Yong Kee; Raynal, Noel J-M; Gharibyan, Vazganush; Su, Ming-Bo; Zhou, Yue-Yang; Li, Jia; Castellano, Sabrina; Sbardella, Gianluca; Issa, Jean-Pierre J; Bedford, Mark T

    2011-03-01

    Worker and queen bees are genetically indistinguishable. However, queen bees are fertile, larger and have a longer lifespan than their female worker counterparts. Differential feeding of larvae with royal jelly controls this caste switching. There is emerging evidence that the queen-bee phenotype is driven by epigenetic mechanisms. In this study, we show that royal jelly--the secretion produced by the hypopharyngeal and mandibular glands of worker bees--has histone deacetylase inhibitor (HDACi) activity. A fatty acid, (E)-10-hydroxy-2-decenoic acid (10HDA), which accounts for up to 5% of royal jelly, harbours this HDACi activity. Furthermore, 10HDA can reactivate the expression of epigenetically silenced genes in mammalian cells. Thus, the epigenetic regulation of queen-bee development is probably driven, in part, by HDACi activity in royal jelly.

  12. Acetylation of EGF Receptor Contributes to Tumor Cell Resistance to Histone Deacetylase Inhibitors

    PubMed Central

    Song, Hui; Li, Chia-Wei; Labaff, Adam M.; Lim, Seung-Oe; Li, Long-Yuan; Kan, Shu-Fen; Chen, Yue; Zhang, Kai; Lang, Jingyu; Xie, Xiaoming; Wang, Yan; Huo, Long-Fei; Hsu, Sheng-Chieh; Chen, Xiaomin; Zhao, Yingming; Hung, Mien-Chie

    2011-01-01

    Alteration of epidermal growth factor receptor (EGFR) is involved in various human cancers and has been intensively investigated. A plethora of evidence demonstrates that posttranslational modifications of EGFR play a pivotal role in controlling its function and metabolism. Here, we show that EGFR can be acetylated by CREB binding protein (CBP) acetyltransferase. Interestingly, EGFR acetylation affects its tyrosine phosphorylation, which may contribute to cancer cell resistance to histone deacetylase inhibitors (HDACIs). Since there is an increasing interest in using HDACIs to treat various cancers in the clinic, our current study provides insights and rationale for selecting effective therapeutic regimen. Consistent with the previous reports, we also show that HDACI combined with EGFR inhibitors achieves better therapeutic outcomes and provides a molecular rationale for the enhanced effect of combination therapy. Our results unveil a critical role of EGFR acetylation that regulates EGFR function, which may have an important clinical implication. PMID:21094134

  13. Dynamic phosphorylation of Histone Deacetylase 1 by Aurora kinases during mitosis regulates zebrafish embryos development

    PubMed Central

    Loponte, Sara; Segré, Chiara V.; Senese, Silvia; Miccolo, Claudia; Santaguida, Stefano; Deflorian, Gianluca; Citro, Simona; Mattoscio, Domenico; Pisati, Federica; Moser, Mirjam A.; Visintin, Rosella; Seiser, Christian; Chiocca, Susanna

    2016-01-01

    Histone deacetylases (HDACs) catalyze the removal of acetyl molecules from histone and non-histone substrates playing important roles in chromatin remodeling and control of gene expression. Class I HDAC1 is a critical regulator of cell cycle progression, cellular proliferation and differentiation during development; it is also regulated by many post-translational modifications (PTMs). Herein we characterize a new mitosis-specific phosphorylation of HDAC1 driven by Aurora kinases A and B. We show that this phosphorylation affects HDAC1 enzymatic activity and it is critical for the maintenance of a proper proliferative and developmental plan in a complex organism. Notably, we find that Aurora-dependent phosphorylation of HDAC1 regulates histone acetylation by modulating the expression of genes directly involved in the developing zebrafish central nervous system. Our data represent a step towards the comprehension of HDAC1 regulation by its PTM code, with important implications in unravelling its roles both in physiology and pathology. PMID:27458029

  14. Histone Deacetylase (HDAC) Inhibitors - emerging roles in neuronal memory, learning, synaptic plasticity and neural regeneration.

    PubMed

    Ganai, Shabir Ahmad; Ramadoss, Mahalakshmi; Mahadevan, Vijayalakshmi

    2016-01-01

    Epigenetic regulation of neuronal signalling through histone acetylation dictates transcription programs that govern neuronal memory, plasticity and learning paradigms. Histone Acetyl Transferases (HATs) and Histone Deacetylases (HDACs) are antagonistic enzymes that regulate gene expression through acetylation and deacetylation of histone proteins around which DNA is wrapped inside a eukaryotic cell nucleus. The epigenetic control of HDACs and the cellular imbalance between HATs and HDACs dictate disease states and have been implicated in muscular dystrophy, loss of memory, neurodegeneration and autistic disorders. Altering gene expression profiles through inhibition of HDACs is now emerging as a powerful technique in therapy. This review presents evolving applications of HDAC inhibitors as potential drugs in neurological research and therapy. Mechanisms that govern their expression profiles in neuronal signalling, plasticity and learning will be covered. Promising and exciting possibilities of HDAC inhibitors in memory formation, fear conditioning, ischemic stroke and neural regeneration have been detailed.

  15. Benefits of histone deacetylase inhibitors for acute brain injury: a systematic review of animal studies.

    PubMed

    Gibson, Claire L; Murphy, Sean P

    2010-11-01

    Drugs that inhibit histone deacetylase (HDAC) activities have enormous potential as neuroprotective agents. We performed a systematic review of controlled animal studies that administered known inhibitors of the zinc-dependent HDACs before and/or after acute cerebral injury and assessed anatomic/functional outcomes. Relevant studies were found by searching PubMed, Embase and Web of Science. From more than 100 identified publications, those data meeting specific criteria were analyzed using the Cochrane Review Manager software. A beneficial effect of administering HDAC inhibitors was seen in studies involving cerebral ischemia or non-ischemic models of acute cerebral injury. Specific studies assessed efficacy when drug was administered up to 14 days prior to, and 14 days following, the onset of cerebral injury. This systematic review provides objective evidence of a neuroprotective role for drugs that inhibit HDACs and highlights particular areas that require further experimental investigation. © 2010 The Authors. Journal of Neurochemistry © 2010 International Society for Neurochemistry.

  16. Histone deacetylase inhibitor activity in royal jelly might facilitate caste switching in bees

    PubMed Central

    Spannhoff, Astrid; Kim, Yong Kee; Raynal, Noel J -M; Gharibyan, Vazganush; Su, Ming-Bo; Zhou, Yue-Yang; Li, Jia; Castellano, Sabrina; Sbardella, Gianluca; Issa, Jean-Pierre J; Bedford, Mark T

    2011-01-01

    Worker and queen bees are genetically indistinguishable. However, queen bees are fertile, larger and have a longer lifespan than their female worker counterparts. Differential feeding of larvae with royal jelly controls this caste switching. There is emerging evidence that the queen-bee phenotype is driven by epigenetic mechanisms. In this study, we show that royal jelly—the secretion produced by the hypopharyngeal and mandibular glands of worker bees—has histone deacetylase inhibitor (HDACi) activity. A fatty acid, (E)-10-hydroxy-2-decenoic acid (10HDA), which accounts for up to 5% of royal jelly, harbours this HDACi activity. Furthermore, 10HDA can reactivate the expression of epigenetically silenced genes in mammalian cells. Thus, the epigenetic regulation of queen-bee development is probably driven, in part, by HDACi activity in royal jelly. PMID:21331099

  17. Histone Deacetylase (HDAC) Inhibitors - Emerging Roles in Neuronal Memory, Learning, Synaptic Plasticity and Neural Regeneration

    PubMed Central

    Ahmad Ganai, Shabir; Ramadoss, Mahalakshmi; Mahadevan, Vijayalakshmi

    2016-01-01

    Epigenetic regulation of neuronal signalling through histone acetylation dictates transcription programs that govern neuronal memory, plasticity and learning paradigms. Histone Acetyl Transferases (HATs) and Histone Deacetylases (HDACs) are antagonistic enzymes that regulate gene expression through acetylation and deacetylation of histone proteins around which DNA is wrapped inside a eukaryotic cell nucleus. The epigenetic control of HDACs and the cellular imbalance between HATs and HDACs dictate disease states and have been implicated in muscular dystrophy, loss of memory, neurodegeneration and autistic disorders. Altering gene expression profiles through inhibition of HDACs is now emerging as a powerful technique in therapy. This review presents evolving applications of HDAC inhibitors as potential drugs in neurological research and therapy. Mechanisms that govern their expression profiles in neuronal signalling, plasticity and learning will be covered. Promising and exciting possibilities of HDAC inhibitors in memory formation, fear conditioning, ischemic stroke and neural regeneration have been detailed. PMID:26487502

  18. Delaying histone deacetylase response to injury accelerates conversion into repair Schwann cells and nerve regeneration

    PubMed Central

    Brügger, Valérie; Duman, Mert; Bochud, Maëlle; Münger, Emmanuelle; Heller, Manfred; Ruff, Sophie; Jacob, Claire

    2017-01-01

    The peripheral nervous system (PNS) regenerates after injury. However, regeneration is often compromised in the case of large lesions, and the speed of axon reconnection to their target is critical for successful functional recovery. After injury, mature Schwann cells (SCs) convert into repair cells that foster axonal regrowth, and redifferentiate to rebuild myelin. These processes require the regulation of several transcription factors, but the driving mechanisms remain partially understood. Here we identify an early response to nerve injury controlled by histone deacetylase 2 (HDAC2), which coordinates the action of other chromatin-remodelling enzymes to induce the upregulation of Oct6, a key transcription factor for SC development. Inactivating this mechanism using mouse genetics allows earlier conversion into repair cells and leads to faster axonal regrowth, but impairs remyelination. Consistently, short-term HDAC1/2 inhibitor treatment early after lesion accelerates functional recovery and enhances regeneration, thereby identifying a new therapeutic strategy to improve PNS regeneration after lesion. PMID:28139683

  19. SIRT1 deacetylase in POMC neurons is required for homeostatic defenses against diet-induced obesity

    PubMed Central

    Ramadori, Giorgio; Fujikawa, Teppei; Fukuda, Makoto; Anderson, Jason; Morgan, Donald A.; Mostoslavsky, Raul; Stuart, Ronald C.; Perello, Mario; Vianna, Claudia R.; Nillni, Eduardo A.; Rahmouni, Kamal; Coppari, Roberto

    2010-01-01

    Summary Feeding on high-calorie (HC) diets induces serious metabolic imbalances, including obesity. Understanding the mechanisms against excessive body weight gain is critical for developing effective anti-obesity strategies. Here, we show that lack of nicotinamide adenosine dinucleotide (NAD+)-dependent deacetylase SIRT1 in pro-opiomelanocortin (POMC) neurons causes hypersensitivity to diet-induced obesity due to reduced energy expenditure. The ability of leptin to properly engage the phosphoinositide 3-kinase (PI3K) signaling in POMC neurons and elicit remodeling of perigonadal white adipose tissue (WAT) is severely compromised in mutant mice. Also, electrophysiological and histomorphomolecular analyses indicate a selective reduction in sympathetic nerve activity and brown-fat-like characteristics in perigonadal WAT of mutant mice; suggesting a physiologically important role for POMC neurons in controlling this visceral fat depot. In summary, our results provide direct genetic evidence that SIRT1 in POMC neurons is required for normal autonomic adaptations against diet-induced obesity. PMID:20620997

  20. Functional Interaction between Class II Histone Deacetylases and ICP0 of Herpes Simplex Virus Type 1

    PubMed Central

    Lomonte, Patrick; Thomas, Joëlle; Texier, Pascale; Caron, Cécile; Khochbin, Saadi; Epstein, Alberto L.

    2004-01-01

    This study describes the physical and functional interactions between ICP0 of herpes simplex virus type 1 and class II histone deacetylases (HDACs) 4, 5, and 7. Class II HDACs are mainly known for their participation in the control of cell differentiation through the regulation of the activity of the transcription factor MEF2 (myocyte enhancer factor 2), implicated in muscle development and neuronal survival. Immunofluorescence experiments performed on transfected cells showed that ICP0 colocalizes with and reorganizes the nuclear distribution of ectopically expressed class I and II HDACs. In addition, endogenous HDAC4 and at least one of its binding partners, the corepressor protein SMRT (for silencing mediator of retinoid and thyroid receptor), undergo changes in their nuclear distribution in ICP0-transfected cells. As a result, during infection endogenous HDAC4 colocalizes with ICP0. Coimmunoprecipitation and glutathione S-transferase pull-down assays confirmed that class II but not class I HDACs specifically interacted with ICP0 through their amino-terminal regions. This region, which is not conserved in class I HDACs but homologous to the MITR (MEF2-interacting transcription repressor) protein, is responsible for the repression, in a deacetylase-independent manner, of MEF2 by sequestering it under an inactive form in the nucleus. Consequently, we show that ICP0 is able to overcome the HDAC5 amino-terminal- and MITR-induced MEF2A repression in gene reporter assays. This is the first report of a viral protein interacting with and controlling the repressor activity of class II HDACs. We discuss the putative consequences of such an interaction for the biology of the virus both during lytic infection and reactivation from latency. PMID:15194749

  1. Crystal Structures of Lipoglycopeptide Antibiotic Deacetylases: Implications for the Biosynthesis of A40926 and Teicoplanin

    SciTech Connect

    Zou, Yaozhong; Brunzelle, Joseph S.; Nair, Satish K.

    2008-07-08

    The lipoglycopeptide antibiotics teicoplanin and A40926 have proven efficacy against Gram-positive pathogens. These drugs are distinguished from glycopeptide antibiotics by N-linked long chain acyl-D-glucosamine decorations that contribute to antibacterial efficacy. During the biosynthesis of lipoglycopeptides, tailoring glycosyltransferases attach an N-acetyl-D-glucosamine to the aglycone, and this N-acetyl-glucosaminyl pseudoaglycone is deacetylated prior to long chain hydrocarbon attachment. Here we present several high-resolution crystal structures of the pseudoaglycone deacetylases from the biosynthetic pathways of teicoplanin and A40926. The cocrystal structure of the teicoplanin pseudoaglycone deacetylase with a fatty acid product provides further insights into the roles of active-site residues, and suggests mechanistic similarities with structurally distinct zinc deacetylases, such as peptidoglycan deacetylase and LpxC. A unique, structurally mobile capping lid, located at the apex of these pseudoaglycone deacetylases, likely serves as a determinant of substrate specificity.

  2. Evidence of bovine immunoglobulin G1 (IgG1) protease activity in partially purified culture supernate of Pasteurella haemolytica A1.

    PubMed Central

    Lee, C W; Shewen, P E

    1996-01-01

    In the bovine respiratory tract, IgG1 is a major secretory immunoglobulin (Ig), and both IgG1 and IgG2 are believed to be important in defense against pneumonic pasteurellosis (shipping fever) in calves. Here we provide evidence for hydrolysis of IgG1 in the presence of partially purified culture supernate (ppCS) from the respiratory pathogen Pasteurella haemolytica A1. Bovine IgG1 was hydrolysed sequentially into three distinct bands (approximately 39, 12, and 7 kDa respectively). Furthermore, partial hydrolysis of bovine IgG2 was observed, but neither bovine IgA nor IgM were affected by incubation with ppCS. These findings suggest that the production of an IgG1-specific protease by P. haemolytica A1 may be a virulence mechanism contributing to the pathogenesis of bovine pneumonic pasteurellosis. Images Figure 1. Figure 2. Figure 3. Figure 4. Figure 5. PMID:8785718

  3. Antibacterial activity of the nitrovinylfuran G1 (Furvina) and its conversion products

    PubMed Central

    Allas, Ülar; Toom, Lauri; Selyutina, Anastasia; Mäeorg, Uno; Medina, Ricardo; Merits, Andres; Rinken, Ago; Hauryliuk, Vasili; Kaldalu, Niilo; Tenson, Tanel

    2016-01-01

    2-Bromo-5-(2-bromo-2-nitrovinyl)furan (G1 or Furvina) is an antimicrobial with a direct reactivity against thiol groups. It is active against Gram-positive and Gram-negative bacteria, yeasts and filamentous fungi. By reacting with thiol groups it causes direct damage to proteins but, as a result, is very short-living and interconverts into an array of reaction products. Our aim was to characterize thiol reactivity of G1 and its conversion products and establish how much of antimicrobial and cytotoxic effects are due to the primary activity of G1 and how much can be attributed to its reaction products. Stability of G1 in growth media as well as its conversion in the presence of thiols was characterized. The structures of G1 decomposition products were determined using NMR and mass-spectroscopy. Concentration- and time-dependent killing curves showed that G1 is bacteriostatic for Escherichia coli at the concentration of 16 μg/ml and bactericidal at 32 μg/ml. However, G1 is inefficient against non-growing E. coli. Addition of cysteine to medium reduces the antimicrobial potency of G1. Nevertheless, the reaction products of G1 and cysteine enabled prolonged antimicrobial action of the drug. Therefore, the activity of 2-bromo-5-(2-bromo-2-nitrovinyl)furan is a sum of its immediate reactivity and the antibacterial effects of the conversion products. PMID:27830730

  4. Histone deacetylases and their role in motor neuron degeneration

    PubMed Central

    Lazo-Gómez, Rafael; Ramírez-Jarquín, Uri N.; Tovar-y-Romo, Luis B.; Tapia, Ricardo

    2013-01-01

    Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease, characterized by the progressive loss of motor neurons. The cause of this selective neuronal death is unknown, but transcriptional dysregulation is recently emerging as an important factor. The physical substrate for the regulation of the transcriptional process is chromatin, a complex assembly of histones and DNA. Histones are subject to several post-translational modifications, like acetylation, that are a component of the transcriptional regulation process. Histone acetylation and deacetylation is performed by a group of enzymes (histone acetyltransferases (HATs) and deacetylases, respectively) whose modulation can alter the transcriptional state of many regions of the genome, and thus may be an important target in diseases that share this pathogenic process, as is the case for ALS. This review will discuss the present evidence of transcriptional dysregulation in ALS, the role of histone deacetylases (HDACs) in disease pathogenesis, and the novel pharmacologic strategies that are being comprehensively studied to prevent motor neuron death, with focus on sirtuins (SIRT) and their effectors. PMID:24367290

  5. Estrogen regulates histone deacetylases to prevent cardiac hypertrophy

    PubMed Central

    Pedram, Ali; Razandi, Mahnaz; Narayanan, Ramesh; Dalton, James T.; McKinsey, Timothy A.; Levin, Ellis R.

    2013-01-01

    The development and progression of cardiac hypertrophy often leads to heart failure and death, and important modulators of hypertrophy include the histone deacetylase proteins (HDACs). Estrogen inhibits cardiac hypertrophy and progression in animal models and humans. We therefore investigated the influence of 17-β-estradiol on the production, localization, and functions of prohypertrophic (class I) and antihypertrophic (class II) HDACs in cultured neonatal rat cardiomyocytes. 17-β-Estradiol or estrogen receptor β agonists dipropylnitrile and β-LGND2 comparably suppressed angiotensin II–induced HDAC2 (class I) production, HDAC-activating phosphorylation, and the resulting prohypertrophic mRNA expression. In contrast, estrogenic compounds derepressed the opposite effects of angiotensin II on the same parameters for HDAC4 and 5 (class II), resulting in retention of these deacetylases in the nucleus to inhibit hypertrophic gene expression. Key aspects were confirmed in vivo from the hearts of wild-type but not estrogen receptor β (ERβ) gene–deleted mice administered angiotensin II and estrogenic compounds. Our results identify a novel dual regulation of cardiomyocyte HDACs, shown here for the antihypertrophic sex steroid acting at ERβ. This mechanism potentially supports using ERβ agonists as HDAC modulators to treat cardiac disease. PMID:24152730

  6. Activity of chitin deacetylase from Colletotrichum gloeosporioides on chitinous substrates.

    PubMed

    Pacheco, Neith; Trombotto, Stéphane; David, Laurent; Shirai, Keiko

    2013-07-01

    Production of chitin deacetylases from the phytopathogenic fungus Colletotrichum gloeosporioides was successfully achieved by submerged fermentation. The highest specific activity of 0.018 U mg(-1) of protein was obtained after 96 h of cultivation at pH 6 and 28°C. Two bands with molecular weights of 35 kDa and 170 kDa determined with SDS-PAGE displayed deacetylase activities as detected in the zymograms. Reacetylated commercial chitosan (52% acetylation degree) was used as substrate for the extracellular crude extract in order to estimate the kinetic parameters of acetate production as undirected deacetylation measurement. The highest acetate production of 12.8 μmol mL(-1) was obtained using 7.5 mg mL(-1) of substrate. The produced enzyme from C. gloeosporioides achieved up to 25% deacetylation of a chitin substrate (hydrolyzed biological chitin) having 80% degree of acetylation, MW of 102×10(3) g mol(-1) and a crystallinity index of ca. 60%.

  7. Purification and characterization of chitin deacetylase from Colletotrichum lindemuthianum.

    PubMed

    Tsigos, I; Bouriotis, V

    1995-11-03

    Chitin deacetylase (EC 3.5.1.41), the enzyme that catalyzes the hydrolysis of acetamido groups of N-acetyl-D-glucosamine in chitin, has been purified to homogeneity from the culture filtrate of the fungus Colletotrichum lindemuthianum and further characterized. The enzyme is a glycoprotein, and its apparent molecular mass was determined to be approximately 150 kDa. The glycosylation pattern of the enzyme is consistent with a mixture of N-linked glycans including oligomannosidic hybrid and/or complex type, and its carbohydrate content is approximately 67% by weight. Chitin deacetylase is active on several chitinous substrates and chitin derivatives, is not considerably inhibited by carboxylic acids, especially acetic acid, and exhibits a remarkable thermostability. The enzyme requires at least two N-acetyl-D-glucosamine residues (chitobiose) for catalysis. When glycol chitin (a water-soluble chitin derivative) was used as substrate, the optimum temperature for enzyme activity was determined to be 50 degrees C, and the optimum pH was approximately 8.5.

  8. Distinct Contributions of Vaccine-Induced Immunoglobulin G1 (IgG1) and IgG2a Antibodies to Protective Immunity against Influenza

    PubMed Central

    Huber, Victor C.; McKeon, Raelene M.; Brackin, Martha N.; Miller, Laura A.; Keating, Rachael; Brown, Scott A.; Makarova, Natalia; Perez, Daniel R.; MacDonald, Gene H.; McCullers, Jonathan A.

    2006-01-01

    Vaccination represents the most effective form of protection against influenza infection. While neutralizing antibodies are typically measured as a correlate of vaccine-induced protective immunity against influenza, nonneutralizing antibodies may contribute to protection or amelioration of disease. The goal of this study was to dissect the individual contributions of the immunoglobulin G1 (IgG1) and IgG2a antibody isotypes to vaccine-induced immunity against influenza virus. To accomplish this, we utilized an influenza vaccine regimen that selectively enhanced IgG1 or IgG2a antibodies by using either DNA or viral replicon particle (VRP) vectors expressing influenza virus hemagglutinin (HA) (HA-DNA or HA-VRP, respectively). After HA-DNA vaccination, neutralizing antibodies were detected by both in vitro (microneutralization) and in vivo (lung viral titer) methods and were associated with increased IgG1 expression by enzyme-linked immunosorbent assay (ELISA). Vaccination with HA-VRP did not strongly stimulate either neutralizing or IgG1 antibodies but did induce IgG2a antibodies. Expression of IgG2a antibodies in this context correlated with clearance of virus and increased protection against lethal influenza challenge. Increased induction of both antibody isotypes as measured by ELISA was a better correlate for vaccine efficacy than neutralization alone. This study details separate but important roles for both IgG1 and IgG2a expression in vaccination against influenza and argues for the development of vaccine regimens that stimulate and measure expression of both antibody isotypes. PMID:16960108

  9. Mycobacterium tuberculosis arrests host cycle at the G1/S transition to establish long term infection.

    PubMed

    Cumming, Bridgette M; Rahman, Md Aejazur; Lamprecht, Dirk A; Rohde, Kyle H; Saini, Vikram; Adamson, John H; Russell, David G; Steyn, Adrie J C

    2017-05-01

    Signals modulating the production of Mycobacterium tuberculosis (Mtb) virulence factors essential for establishing long-term persistent infection are unknown. The WhiB3 redox regulator is known to regulate the production of Mtb virulence factors, however the mechanisms of this modulation are unknown. To advance our understanding of the mechanisms involved in WhiB3 regulation, we performed Mtb in vitro, intraphagosomal and infected host expression analyses. Our Mtb expression analyses in conjunction with extracellular flux analyses demonstrated that WhiB3 maintains bioenergetic homeostasis in response to available carbon sources found in vivo to establish Mtb infection. Our infected host expression analysis indicated that WhiB3 is involved in regulation of the host cell cycle. Detailed cell-cycle analysis revealed that Mtb infection inhibited the macrophage G1/S transition, and polyketides under WhiB3 control arrested the macrophages in the G0-G1 phase. Notably, infection with the Mtb whiB3 mutant or polyketide mutants had little effect on the macrophage cell cycle and emulated the uninfected cells. This suggests that polyketides regulated by Mtb WhiB3 are responsible for the cell cycle arrest observed in macrophages infected with the wild type Mtb. Thus, our findings demonstrate that Mtb WhiB3 maintains bioenergetic homeostasis to produce polyketide and lipid cyclomodulins that target the host cell cycle. This is a new mechanism whereby Mtb modulates the immune system by altering the host cell cycle to promote long-term persistence. This new knowledge could serve as the foundation for new host-directed therapeutic discovery efforts that target the host cell cycle.

  10. Molecular basis for the dissociation dynamics of protein A-immunoglobulin G1 complex.

    PubMed

    Liu, Fu-Feng; Huang, Bo; Dong, Xiao-Yan; Sun, Yan

    2013-01-01

    Staphylococcus aureus protein A (SpA) is the most popular affinity ligand for immunoglobulin G1 (IgG1). However, the molecular basis for the dissociation dynamics of SpA-IgG1 complex is unclear. Herein, coarse-grained (CG) molecular dynamics (MD) simulations with the Martini force field were used to study the dissociation dynamics of the complex. The CG-MD simulations were first verified by the agreement in the structural and interactional properties of SpA and human IgG1 (hIgG1) in the association process between the CG-MD and all-atom MD at different NaCl concentrations. Then, the CG-MD simulation studies focused on the molecular insight into the dissociation dynamics of SpA-hIgG1 complex at pH 3.0. It is found that there are four steps in the dissociation process of the complex. First, there is a slight conformational adjustment of helix II in SpA. This is followed by the phenomena that the electrostatic interactions provided by the three hot spots (Glu143, Arg146 and Lys154) of helix II of SpA break up, leading to the dissociation of helix II from the binding site of hIgG1. Subsequently, breakup of the hydrophobic interactions between helix I (Phe132, Tyr133 and His137) in SpA and hIgG1 occurs, resulting in the disengagement of helix I from its binding site of hIgG1. Finally, the non-specific interactions between SpA and hIgG1 decrease slowly till disappearance, leading to the complete dissociation of the SpA-hIgG1 complex. This work has revealed that CG-MD coupled with the Martini force field is an effective method for studying the dissociation dynamics of protein-protein complex.

  11. Underlying principles of cell fate determination during G1 phase of the mammalian cell cycle.

    PubMed

    Pfeuty, Benjamin; David-Pfeuty, Thérèse; Kaneko, Kunihiko

    2008-10-01

    Upon their exit from mitosis, mammalian cells enter a G(1) phase during which they acutely sense all sorts of environmental stimuli. On the basis of these signals that they first need to decipher and integrate, they decide whether to undergo division, differentiation, senescence or apoptosis. We questioned whether, despite the complexity of the G(1) regulatory network, simple organizing principles might be identified that could explain how specific input signals are converted into appropriate cell fates. For this purpose, we formulated a mathematical model of the G(1) regulatory network using a simplified description of activities linked to signal transduction, cell growth, cell division and cell death. Bifurcation analysis of the model revealed the existence of multistability between several attractor states corresponding to G(0)-arrest, G(1)-arrest, S-phase entry and apoptosis cell fates. We further unravelled interlinked feedback and feedforward loops within the G(1) regulatory network that drive the signal-dependent transition between G(0) arrest and the other cell fates. Initially, exit from G(0) and progression in early G(1) entail growth factor-dependent activation of an upstream positive feedback loop that activates the cell-growth machinery. Once ribosome synthesis is restored in G(1), a competition develops between a downstream positive feedback loop, which, upon activation, triggers S phase entry, and stress-activated pathways that promote G(1) arrest. If S phase entry prevails over G(1) arrest, cells are sensitized to apoptosis due to stress-induced activation of pro-apoptotic pathways or repression of pro-survival pathways. Thus, the choice between the four possible cell fates in the G(1) phase relies on the flexibly interlinked growth-activatory and division-activatory modules, certain components of which have antagonistic effects on pathways involved in driving apoptosis and G(1) arrest. The final outcome ultimately depends on the context

  12. FoxG1 and TLE2 act cooperatively to regulate ventral telencephalon formation

    PubMed Central

    Roth, Martin; Bonev, Boyan; Lindsay, Jennefer; Lea, Robert; Panagiotaki, Niki; Houart, Corinne; Papalopulu, Nancy

    2010-01-01

    FoxG1 is a conserved transcriptional repressor that plays a key role in the specification, proliferation and differentiation of the telencephalon, and is expressed from the earliest stages of telencephalic development through to the adult. How the interaction with co-factors might influence the multiplicity and diversity of FoxG1 function is not known. Here, we show that interaction of FoxG1 with TLE2, a Xenopus tropicalis co-repressor of the Groucho/TLE family, is crucial for regulating the early activity of FoxG1. We show that TLE2 is co-expressed with FoxG1 in the ventral telencephalon from the early neural plate stage and functionally cooperates with FoxG1 in an ectopic neurogenesis assay. FoxG1 has two potential TLE binding sites: an N-terminal eh1 motif and a C-terminal YWPMSPF motif. Although direct binding seems to be mediated by the N-terminal motif, both motifs appear important for functional synergism. In the neurogenesis assay, mutation of either motif abolishes functional cooperation of TLE2 with FoxG1, whereas in the forebrain deletion of both motifs renders FoxG1 unable to induce the ventral telencephalic marker Nkx2.1. Knocking down either FoxG1 or TLE2 disrupts the development of the ventral telencephalon, supporting the idea that endogenous TLE2 and FoxG1 work together to specify the ventral telencephalon. PMID:20356955

  13. Molecular Basis for the Dissociation Dynamics of Protein A-Immunoglobulin G1 Complex

    PubMed Central

    Liu, Fu-Feng; Huang, Bo; Dong, Xiao-Yan; Sun, Yan

    2013-01-01

    Staphylococcus aureus protein A (SpA) is the most popular affinity ligand for immunoglobulin G1 (IgG1). However, the molecular basis for the dissociation dynamics of SpA-IgG1 complex is unclear. Herein, coarse-grained (CG) molecular dynamics (MD) simulations with the Martini force field were used to study the dissociation dynamics of the complex. The CG-MD simulations were first verified by the agreement in the structural and interactional properties of SpA and human IgG1 (hIgG1) in the association process between the CG-MD and all-atom MD at different NaCl concentrations. Then, the CG-MD simulation studies focused on the molecular insight into the dissociation dynamics of SpA-hIgG1 complex at pH 3.0. It is found that there are four steps in the dissociation process of the complex. First, there is a slight conformational adjustment of helix II in SpA. This is followed by the phenomena that the electrostatic interactions provided by the three hot spots (Glu143, Arg146 and Lys154) of helix II of SpA break up, leading to the dissociation of helix II from the binding site of hIgG1. Subsequently, breakup of the hydrophobic interactions between helix I (Phe132, Tyr133 and His137) in SpA and hIgG1 occurs, resulting in the disengagement of helix I from its binding site of hIgG1. Finally, the non-specific interactions between SpA and hIgG1 decrease slowly till disappearance, leading to the complete dissociation of the SpA-hIgG1 complex. This work has revealed that CG-MD coupled with the Martini force field is an effective method for studying the dissociation dynamics of protein-protein complex. PMID:23776704

  14. Overexpression of Histone Deacetylase 6 Enhances Resistance to Porcine Reproductive and Respiratory Syndrome Virus in Pigs

    PubMed Central

    Li, Qiuyan; Li, Zhiguo; Wang, Meng; Liu, Lin; Tian, Kegong; Li, Ning

    2017-01-01

    Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most economically relevant viral pathogens in pigs and causes substantial losses in the pig industry worldwide each year. At present, PRRSV vaccines do not effectively prevent and control this disease. Consequently, it is necessary to develop new antiviral strategies to compensate for the inefficacy of the available vaccines. Histone deacetylase 6 (HDAC6) is an important member of the histone deacetylase family that is responsible for regulating many important biological processes. Studies have shown that HDAC6 has anti-viral activities during the viral life cycle. However, whether HDAC6 overexpression enhances resistance to PRRSV in pigs remains unknown. In this study, we used a somatic cell cloning method to produce transgenic (TG) pigs that constitutively overexpress porcine HDAC6. These TG pigs showed germ line transmission with continued overexpression of HDAC6. In vitro, virus-challenged porcine alveolar macrophages (PAMs) overexpressed HDAC6, which suppressed viral gene expression and PRRSV production. In vivo, resistance to PRRSV in TG pigs was evaluated by direct or cohabitation mediated infection with a highly pathogenic PRRSV (HP-PRRSV) strain. Compared with non-TG (NTG) siblings, TG pigs showed a significantly lower viral load in the lungs and an extended survival time after infection with HP-PRRSV via intramuscular injection. In the cohabitation study, NTG pigs housed with challenged NTG pigs exhibited significantly worse clinical symptoms than the other three in-contact groups. These results collectively suggest that HDAC6 overexpression enhances resistance to PRRSV infection both in vitro and in vivo. Our findings suggest the potential involvement of HDAC6 in the response to PRRSV, which will facilitate the development of novel therapies for PRRSV. PMID:28052127

  15. Detrimental Effect of Class-selective Histone Deacetylase Inhibitors during Tissue Regeneration following Hindlimb Ischemia

    PubMed Central

    Spallotta, Francesco; Tardivo, Silvia; Nanni, Simona; Rosati, Jessica D.; Straino, Stefania; Mai, Antonello; Vecellio, Matteo; Valente, Sergio; Capogrossi, Maurizio C.; Farsetti, Antonella; Martone, Julie; Bozzoni, Irene; Pontecorvi, Alfredo; Gaetano, Carlo; Colussi, Claudia

    2013-01-01

    Histone deacetylase inhibitors (DIs) are promising drugs for the treatment of several pathologies including ischemic and failing heart where they demonstrated efficacy. However, adverse side effects and cardiotoxicity have also been reported. Remarkably, no information is available about the effect of DIs during tissue regeneration following acute peripheral ischemia. In this study, mice made ischemic by femoral artery excision were injected with the DIs MS275 and MC1568, selective for class I and IIa histone deacetylases (HDACs), respectively. In untreated mice, soon after damage, class IIa HDAC phosphorylation and nuclear export occurred, paralleled by dystrophin and neuronal nitric-oxide synthase (nNOS) down-regulation and decreased protein phosphatase 2A activity. Between 14 and 21 days after ischemia, dystrophin and nNOS levels recovered, and class IIa HDACs relocalized to the nucleus. In this condition, the MC1568 compound increased the number of newly formed muscle fibers but delayed their terminal differentiation, whereas MS275 abolished the early onset of the regeneration process determining atrophy and fibrosis. The selective DIs had differential effects on the vascular compartment: MC1568 increased arteriogenesis whereas MS275 inhibited it. Capillarogenesis did not change. Chromatin immunoprecipitations revealed that class IIa HDAC complexes bind promoters of proliferation-associated genes and of class I HDAC1 and 2, highlighting a hierarchical control between class II and I HDACs during tissue regeneration. Our findings indicate that class-selective DIs interfere with normal mouse ischemic hindlimb regeneration and suggest that their use could be limited by alteration of the regeneration process in peripheral ischemic tissues. PMID:23836913

  16. Identification and Cloning of a Cryptococcal Deacetylase That Produces Protective Immune Responses

    PubMed Central

    Biondo, Carmelo; Beninati, Concetta; Delfino, Demetrio; Oggioni, Marco; Mancuso, Giuseppe; Midiri, Angelina; Bombaci, Mauro; Tomaselli, Giuseppe; Teti, Giuseppe

    2002-01-01

    Cell-mediated immunity plays a crucial role in host defenses against Cryptococcus (Filobasidiella) neoformans. Therefore, the identification of cryptococcal antigens capable of producing T-cell-mediated responses, such as delayed-type hypersensitivity (DTH) reactions, may be useful in the development of immune-based strategies to control cryptococcosis. In order to characterize DTH-producing antigens, culture supernatants from the unencapsulated Cap-67 strain were separated by anion-exchange chromatography. After further fractionation by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis, a purified protein with an apparent molecular mass of 25 kDa was found to produce DTH, as evidenced by increased footpad swelling in mice immunized with culture supernatants, relative to unimmunized mice. The 20-amino-acid N-terminal sequence of the 25-kDa protein was used to search data of the C. neoformans Genome Project. Based on the genomic DNA sequence, a DNA probe was used to screen a λ cDNA library prepared from strain B3501. Clones were isolated containing the full-length gene (d25), which showed homology with a number of polysaccharide deacetylases from fungi and bacteria. The recombinant d25 protein expressed in Escherichia coli was similar to the natural one in DTH-producing activity. Moreover, immunization with either the natural or the recombinant protein prolonged survival and decreased fungal burden in mice challenged with the highly virulent C. neoformans strain H99. In conclusion, we have described the first cryptococcal gene whose product, a 25-kDa extracellular polysaccharide deacetylase, has been shown to induce protective immunity responses. PMID:11953374

  17. Histone deacetylase inhibition blunts ischemia/reperfusion injury by inducing cardiomyocyte autophagy.

    PubMed

    Xie, Min; Kong, Yongli; Tan, Wei; May, Herman; Battiprolu, Pavan K; Pedrozo, Zully; Wang, Zhao V; Morales, Cyndi; Luo, Xiang; Cho, Geoffrey; Jiang, Nan; Jessen, Michael E; Warner, John J; Lavandero, Sergio; Gillette, Thomas G; Turer, Aslan T; Hill, Joseph A

    2014-03-11

    Reperfusion accounts for a substantial fraction of the myocardial injury occurring with ischemic heart disease. Yet, no standard therapies are available targeting reperfusion injury. Here, we tested the hypothesis that suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor approved for cancer treatment by the US Food and Drug Administration, will blunt reperfusion injury. Twenty-one rabbits were randomly assigned to 3 groups: (1) vehicle control, (2) SAHA pretreatment (1 day before and at surgery), and (3) SAHA treatment at the time of reperfusion only. Each arm was subjected to ischemia/reperfusion surgery (30 minutes coronary ligation, 24 hours reperfusion). In addition, cultured neonatal and adult rat ventricular cardiomyocytes were subjected to simulated ischemia/reperfusion to probe mechanism. SAHA reduced infarct size and partially rescued systolic function when administered either before surgery (pretreatment) or solely at the time of reperfusion. SAHA plasma concentrations were similar to those achieved in patients with cancer. In the infarct border zone, SAHA increased autophagic flux, assayed in both rabbit myocardium and in mice harboring an RFP-GFP-LC3 transgene. In cultured myocytes subjected to simulated ischemia/reperfusion, SAHA pretreatment reduced cell death by 40%. This reduction in cell death correlated with increased autophagic activity in SAHA-treated cells. RNAi-mediated knockdown of ATG7 and ATG5, essential autophagy proteins, abolished SAHA's cardioprotective effects. The US Food and Drug Administration-approved anticancer histone deacetylase inhibitor, SAHA, reduces myocardial infarct size in a large animal model, even when delivered in the clinically relevant context of reperfusion. The cardioprotective effects of SAHA during ischemia/reperfusion occur, at least in part, through the induction of autophagic flux.

  18. Detrimental effect of class-selective histone deacetylase inhibitors during tissue regeneration following hindlimb ischemia.

    PubMed

    Spallotta, Francesco; Tardivo, Silvia; Nanni, Simona; Rosati, Jessica D; Straino, Stefania; Mai, Antonello; Vecellio, Matteo; Valente, Sergio; Capogrossi, Maurizio C; Farsetti, Antonella; Martone, Julie; Bozzoni, Irene; Pontecorvi, Alfredo; Gaetano, Carlo; Colussi, Claudia

    2013-08-09

    Histone deacetylase inhibitors (DIs) are promising drugs for the treatment of several pathologies including ischemic and failing heart where they demonstrated efficacy. However, adverse side effects and cardiotoxicity have also been reported. Remarkably, no information is available about the effect of DIs during tissue regeneration following acute peripheral ischemia. In this study, mice made ischemic by femoral artery excision were injected with the DIs MS275 and MC1568, selective for class I and IIa histone deacetylases (HDACs), respectively. In untreated mice, soon after damage, class IIa HDAC phosphorylation and nuclear export occurred, paralleled by dystrophin and neuronal nitric-oxide synthase (nNOS) down-regulation and decreased protein phosphatase 2A activity. Between 14 and 21 days after ischemia, dystrophin and nNOS levels recovered, and class IIa HDACs relocalized to the nucleus. In this condition, the MC1568 compound increased the number of newly formed muscle fibers but delayed their terminal differentiation, whereas MS275 abolished the early onset of the regeneration process determining atrophy and fibrosis. The selective DIs had differential effects on the vascular compartment: MC1568 increased arteriogenesis whereas MS275 inhibited it. Capillarogenesis did not change. Chromatin immunoprecipitations revealed that class IIa HDAC complexes bind promoters of proliferation-associated genes and of class I HDAC1 and 2, highlighting a hierarchical control between class II and I HDACs during tissue regeneration. Our findings indicate that class-selective DIs interfere with normal mouse ischemic hindlimb regeneration and suggest that their use could be limited by alteration of the regeneration process in peripheral ischemic tissues.

  19. Deacetylase inhibitors modulate proliferation and self-renewal properties of leukemic stem and progenitor cells

    PubMed Central

    Romanski, Annette; Schwarz, Kerstin; Keller, Maren; Wietbrauk, Sarah; Vogel, Anja; Roos, Jessica; Oancea, Claudia; Brill, Boris; Krämer, Oliver H.; Serve, Hubert; Ruthardt, Martin; Bug, Gesine

    2012-01-01

    Acute myeloid leukemia (AML) is a highly malignant disease that is not curable in the majority of patients. Numerous non-random genetic abnormalities are known, among which several translocations such as PLZF/RARα or AML1/ETO are known to aberrantly recruit histone deacetylases. Deacetylase inhibitors (DACi) are promising drugs leading to growth inhibition, cell cycle arrest, premature senescence and apoptosis in malignant cells. It is believed that DACi may have clinical efficacy by eradicating the most primitive population of leukemic stem and progenitor cells, possibly by interfering with self-renewal. The aim of the study was to investigate the effects of DACi on leukemic stem and progenitor cells using murine transduction-transplantation models of hematopoietic cells harboring the leukemia-associated fusion proteins (LAFP) PLZF/RARα or a truncated AML1/ETO protein (AML1/ETO exon 9). We show that the self-renewal and short-term repopulation capacity of AML1/ETO- or PLZF/RARα-expressing Sca1+/lin- stem and progenitor cells are profoundly inhibited by clinically applicable concentrations of the DACi dacinostat and vorinostat. To further investigate the mechanisms underlying these effects, we examined the impact of DACi on the transcription factor c-MYC and the Polycomb group protein BMI1, which are induced by LAFP and involved in leukemic transformation. In AML1/ETO or PLZF/RARα-positive 32D cells, DACi-mediated antiproliferative effects were associated with downregulation of BMI1 and c-MYC protein levels. Similar effects were demonstrated in primary samples of cytogenetically defined high-risk AML patients. In conclusion, DACi may be effective as maintenance therapy by negatively interfering with signaling pathways that control survival and proliferation of leukemic stem and progenitor cells. PMID:22895185

  20. A Recombinant Fungal Chitin Deacetylase Produces Fully Defined Chitosan Oligomers with Novel Patterns of Acetylation.

    PubMed

    Naqvi, Shoa; Cord-Landwehr, Stefan; Singh, Ratna; Bernard, Frank; Kolkenbrock, Stephan; El Gueddari, Nour Eddine; Moerschbacher, Bruno M

    2016-11-15

    Partially acetylated chitosan oligosaccharides (paCOS) are potent biologics with many potential applications, and their bioactivities are believed to be dependent on their structure, i.e., their degrees of polymerization and acetylation, as well as their pattern of acetylation. However, paCOS generated via chemical N-acetylation or de-N-acetylation of GlcN or GlcNAc oligomers, respectively, typically display random patterns of acetylation, making it difficult to control and predict their bioactivities. In contrast, paCOS produced from chitin deacetylases (CDAs) acting on chitin oligomer substrates may have specific patterns of acetylation, as shown for some bacterial CDAs. However, compared to what we know about bacterial CDAs, we know little about the ability of fungal CDAs to produce defined paCOS with known patterns of acetylation. Therefore, we optimized the expression of a chitin deacetylase from the fungus Puccinia graminis f. sp. tritici in Escherichia coli The best yield of functional enzyme was obtained as a fusion protein with the maltose-binding protein (MBP) secreted into the periplasmic space of the bacterial host. We characterized the MBP fusion protein from P. graminis (PgtCDA) and tested its activity on different chitinous substrates. Mass spectrometric sequencing of the products obtained by enzymatic deacetylation of chitin oligomers, i.e., tetramers to hexamers, revealed that PgtCDA generated paCOS with specific acetylation patterns of A-A-D-D, A-A-D-D-D, and A-A-D-D-D-D, respectively (A, GlcNAc; D, GlcN), indicating that PgtCDA cannot deacetylate the two GlcNAc units closest to the oligomer's nonreducing end. This unique property of PgtCDA significantly expands the so far very limited library of well-defined paCOS available to test their bioactivities for a wide variety of potential applications.

  1. Whole exome sequencing of rare variants in EIF4G1 and VPS35 in Parkinson disease

    PubMed Central

    Nuytemans, Karen; Bademci, Guney; Inchausti, Vanessa; Dressen, Amy; Kinnamon, Daniel D.; Mehta, Arpit; Wang, Liyong; Züchner, Stephan; Beecham, Gary W.; Martin, Eden R.; Scott, William K.

    2013-01-01

    Objective: Recently, vacuolar protein sorting 35 (VPS35) and eukaryotic translation initiation factor 4 gamma 1 (EIF4G1) have been identified as 2 causal Parkinson disease (PD) genes. We used whole exome sequencing for rapid, parallel analysis of variations in these 2 genes. Methods: We performed whole exome sequencing in 213 patients with PD and 272 control individuals. Those rare variants (RVs) with <5% frequency in the exome variant server database and our own control data were considered for analysis. We performed joint gene-based tests for association using RVASSOC and SKAT (Sequence Kernel Association Test) as well as single-variant test statistics. Results: We identified 3 novel VPS35 variations that changed the coded amino acid (nonsynonymous) in 3 cases. Two variations were in multiplex families and neither segregated with PD. In EIF4G1, we identified 11 (9 nonsynonymous and 2 small indels) RVs including the reported pathogenic mutation p.R1205H, which segregated in all affected members of a large family, but also in 1 unaffected 86-year-old family member. Two additional RVs were found in isolated patients only. Whereas initial association studies suggested an association (p = 0.04) with all RVs in EIF4G1, subsequent testing in a second dataset for the driving variant (p.F1461) suggested no association between RVs in the gene and PD. Conclusions: We confirm that the specific EIF4G1 variation p.R1205H seems to be a strong PD risk factor, but is nonpenetrant in at least one 86-year-old. A few other select RVs in both genes could not be ruled out as causal. However, there was no evidence for an overall contribution of genetic variability in VPS35 or EIF4G1 to PD development in our dataset. PMID:23408866

  2. H3K9me-Independent Gene Silencing in Fission Yeast Heterochromatin by Clr5 and Histone Deacetylases

    PubMed Central

    Hansen, Klavs R.; Hazan, Idit; Shanker, Sreenath; Watt, Stephen; Verhein-Hansen, Janne; Bähler, Jürg; Martienssen, Robert A.; Partridge, Janet F.; Cohen, Amikam; Thon, Geneviève

    2011-01-01

    Nucleosomes in heterochromatic regions bear histone modifications that distinguish them from euchromatic nucleosomes. Among those, histone H3 lysine 9 methylation (H3K9me) and hypoacetylation have been evolutionarily conserved and are found in both multicellular eukaryotes and single-cell model organisms such as fission yeast. In spite of numerous studies, the relative contributions of the various heterochromatic histone marks to the properties of heterochromatin remain largely undefined. Here, we report that silencing of the fission yeast mating-type cassettes, which are located in a well-characterized heterochromatic region, is hardly affected in cells lacking the H3K9 methyltransferase Clr4. We document the existence of a pathway parallel to H3K9me ensuring gene repression in the absence of Clr4 and identify a silencing factor central to this pathway, Clr5. We find that Clr5 controls gene expression at multiple chromosomal locations in addition to affecting the mating-type region. The histone deacetylase Clr6 acts in the same pathway as Clr5, at least for its effects in the mating-type region, and on a subset of other targets, notably a region recently found to be prone to neo-centromere formation. The genomic targets of Clr5 also include Ste11, a master regulator of sexual differentiation. Hence Clr5, like the multi-functional Atf1 transcription factor which also modulates chromatin structure in the mating-type region, controls sexual differentiation and genome integrity at several levels. Globally, our results point to histone deacetylases as prominent repressors of gene expression in fission yeast heterochromatin. These deacetylases can act in concert with, or independently of, the widely studied H3K9me mark to influence gene silencing at heterochromatic loci. PMID:21253571

  3. The TCP4 transcription factor of Arabidopsis blocks cell division in yeast at G1 {yields} S transition

    SciTech Connect

    Aggarwal, Pooja; Padmanabhan, Bhavna; Bhat, Abhay; Sarvepalli, Kavitha; Sadhale, Parag P.; Nath, Utpal

    2011-07-01

    Highlights: {yields} TCP4 is a class II TCP transcription factor, that represses cell division in Arabidopsis. {yields} TCP4 expression in yeast retards cell division by blocking G1 {yields} S transition. {yields} Genome-wide expression studies and Western analysis reveals stabilization of cell cycle inhibitor Sic1, as possible mechanism. -- Abstract: The TCP transcription factors control important aspects of plant development. Members of class I TCP proteins promote cell cycle by regulating genes directly involved in cell proliferation. In contrast, members of class II TCP proteins repress cell division. While it has been postulated that class II proteins induce differentiation signal, their exact role on cell cycle has not been studied. Here, we report that TCP4, a class II TCP protein from Arabidopsis that repress cell proliferation in developing leaves, inhibits cell division by blocking G1 {yields} S transition in budding yeast. Cells expressing TCP4 protein with increased transcriptional activity fail to progress beyond G1 phase. By analyzing global transcriptional status of these cells, we show that expression of a number of cell cycle genes is altered. The possible mechanism of G1 {yields} S arrest is discussed.

  4. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 16 Commercial Practices 1 2011-01-01 2011-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305 Commercial Practices FEDERAL TRADE COMMISSION REGULATIONS UNDER SPECIFIC ACTS OF CONGRESS RULE CONCERNING... Part 305—Furnaces—Gas Manufacturer's rated heating capacities (Btu's/hr.) Range of annual...

  5. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 16 Commercial Practices 1 2010-01-01 2010-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305 Commercial Practices FEDERAL TRADE COMMISSION REGULATIONS UNDER SPECIFIC ACTS OF CONGRESS RULE CONCERNING... Part 305—Furnaces—Gas Manufacturer's rated heating capacities (Btu's/hr.) Range of annual...

  6. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 16 Commercial Practices 1 2013-01-01 2013-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305 Commercial Practices FEDERAL TRADE COMMISSION REGULATIONS UNDER SPECIFIC ACTS OF CONGRESS RULE CONCERNING... Part 305—Furnaces—Gas Manufacturer's rated heating capacities (Btu's/hr.) Range of annual...

  7. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 16 Commercial Practices 1 2012-01-01 2012-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305 Commercial Practices FEDERAL TRADE COMMISSION REGULATIONS UNDER SPECIFIC ACTS OF CONGRESS RULE CONCERNING... Part 305—Furnaces—Gas Manufacturer's rated heating capacities (Btu's/hr.) Range of annual...

  8. Acrocentric Chromosomes in Cultured Leukocytes from Mothers of Children Affected With the G1- Trisomy Syndrome

    ERIC Educational Resources Information Center

    And Others; Cotton, James E.

    1973-01-01

    Analysis of venous blood samples from 24 mothers of G1-trisomy-affected (Down's Syndrome) children and 23 mothers of chromosomally normal children indicated that mothers of G1-trisomy-affected children had a greater than expected involvement of the G-chromosomes in associations of acrocentric satellited (chromosome configuration) chromosomes.…

  9. A hyperactive transcriptional state marks genome reactivation at the mitosis-G1 transition.

    PubMed

    Hsiung, Chris C-S; Bartman, Caroline R; Huang, Peng; Ginart, Paul; Stonestrom, Aaron J; Keller, Cheryl A; Face, Carolyne; Jahn, Kristen S; Evans, Perry; Sankaranarayanan, Laavanya; Giardine, Belinda; Hardison, Ross C; Raj, Arjun; Blobel, Gerd A

    2016-06-15

    During mitosis, RNA polymerase II (Pol II) and many transcription factors dissociate from chromatin, and transcription ceases globally. Transcription is known to restart in bulk by telophase, but whether de novo transcription at the mitosis-G1 transition is in any way distinct from later in interphase remains unknown. We tracked Pol II occupancy genome-wide in mammalian cells progressing from mitosis through late G1. Unexpectedly, during the earliest rounds of transcription at the mitosis-G1 transition, ∼50% of active genes and distal enhancers exhibit a spike in transcription, exceeding levels observed later in G1 phase. Enhancer-promoter chromatin contacts are depleted during mitosis and restored rapidly upon G1 entry but do not spike. Of the chromatin-associated features examined, histone H3 Lys27 acetylation levels at individual loci in mitosis best predict the mitosis-G1 transcriptional spike. Single-molecule RNA imaging supports that the mitosis-G1 transcriptional spike can constitute the maximum transcriptional activity per DNA copy throughout the cell division cycle. The transcriptional spike occurs heterogeneously and propagates to cell-to-cell differences in mature mRNA expression. Our results raise the possibility that passage through the mitosis-G1 transition might predispose cells to diverge in gene expression states. © 2016 Hsiung et al.; Published by Cold Spring Harbor Laboratory Press.

  10. Enhanced HIV-1 neutralization by a CD4-VH3-IgG1 fusion protein

    SciTech Connect

    Meyuhas, Ronit; Noy, Hava; Fishman, Sigal; Margalit, Alon; Montefiori, David C.; Gross, Gideon

    2009-08-21

    HIV-1 gp120 is an alleged B cell superantigen, binding certain VH3+ human antibodies. We reasoned that a CD4-VH3 fusion protein could possess higher affinity for gp120 and improved HIV-1 inhibitory capacity. To test this we produced several human IgG1 immunoligands harboring VH3. Unlike VH3-IgG1 or VH3-CD4-IgG1, CD4-VH3-IgG1 bound gp120 considerably stronger than CD4-IgG1. CD4-VH3-IgG1 exhibited {approx}1.5-2.5-fold increase in neutralization of two T-cell laboratory-adapted strains when compared to CD4-IgG1. CD4-VH3-IgG1 improved neutralization of 7/10 clade B primary isolates or pseudoviruses, exceeding 20-fold for JR-FL and 13-fold for Ba-L. It enhanced neutralization of 4/8 clade C viruses, and had negligible effect on 1/4 clade A pseudoviruses. We attribute this improvement to possible pairing of VH3 with CD4 D1 and stabilization of an Ig Fv-like structure, rather than to superantigen interactions. These novel findings support the current notion that CD4 fusion proteins can act as better HIV-1 entry inhibitors with potential clinical implications.

  11. 26 CFR 1.45G-1 - Railroad track maintenance credit.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 26 Internal Revenue 1 2013-04-01 2013-04-01 false Railroad track maintenance credit. 1.45G-1... TAXES Rules for Computing Credit for Investment in Certain Depreciable Property § 1.45G-1 Railroad track maintenance credit. (a) In general. For purposes of section 38, the railroad track maintenance credit...

  12. 26 CFR 1.45G-1 - Railroad track maintenance credit.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 26 Internal Revenue 1 2012-04-01 2012-04-01 false Railroad track maintenance credit. 1.45G-1... TAXES Rules for Computing Credit for Investment in Certain Depreciable Property § 1.45G-1 Railroad track maintenance credit. (a) In general. For purposes of section 38, the railroad track maintenance credit...

  13. 26 CFR 1.45G-1 - Railroad track maintenance credit.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 26 Internal Revenue 1 2014-04-01 2013-04-01 true Railroad track maintenance credit. 1.45G-1... TAXES Rules for Computing Credit for Investment in Certain Depreciable Property § 1.45G-1 Railroad track maintenance credit. (a) In general. For purposes of section 38, the railroad track maintenance credit...

  14. 26 CFR 1.45G-1 - Railroad track maintenance credit.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 1 2010-04-01 2010-04-01 true Railroad track maintenance credit. 1.45G-1... TAXES Rules for Computing Credit for Investment in Certain Depreciable Property § 1.45G-1 Railroad track maintenance credit. (a) In general. For purposes of section 38, the railroad track maintenance credit (RTMC...

  15. A Hot, Extended Horizontal Branch in the Massive M31 Globular Cluster G1

    NASA Astrophysics Data System (ADS)

    Rich, Robert M.; Piotto, G.; Reitzel, D.; Origlia, L.; Bedin, L.

    2013-01-01

    The globular cluster G1 in M31 is among the most massive in the Local Group and may host a supermassive central black hole of ~20,000 Msun. We have used WFC3 on HST to image G1 in the F275, B, and V bands. G1 has also been proposed as likely to host a complex stellar population like that of ω Cen, which has been proposed as a possible nucleus of a dwarf galaxy. We find an extended blue horizontal branch in G1, although it appears the blue HB is <20% of the red clump population. Although we detect a wide red giant branch, neither the RGB nor HB have the complexity of those observed in Omega Cen. We have a solid detection of an extended HB in G1, but we do not strong evidence for the extreme complexity exhibited by ω Cen.

  16. Alum Directly Modulates Murine B Lymphocytes to Produce IgG1 Isotype

    PubMed Central

    Jin, Bo-Ra; Kim, Sun-Jin; Lee, Jeong-Min; Kang, Seong-Ho; Han, Hye-Ju; Jang, Young-Saeng; Seo, Goo-young

    2013-01-01

    Aluminum hydroxide (alum) is the most widely used adjuvant in human vaccines. Nevertheless, it is virtually unknown whether alum acts on B cells. In the present study, we explored the direct effect of alum on Ig expression by murine B cells in vitro. LPS-activated mouse spleen B cells were cultured with alum, and the level of isotype-specific Ig secretion, IgG1 secreting cell numbers, and Ig germ-line transcripts (GLT) were measured using ELISA, ELISPOT, and RT-PCR, respectively. Alum consistently enhanced total IgG1 production, numbers of IgG1 secreting cells, and GLTγ1 expression. These results demonstrate that alum can directly cause IgG1 isotype switching leading to IgG1 production. PMID:23559895

  17. Histone Deacetylase Inhibitors Inhibit the Proliferation of Gallbladder Carcinoma Cells by Suppressing AKT/mTOR Signaling.

    PubMed

    Zhang, Peng; Guo, Zhiyong; Wu, Ying; Hu, Ronglin; Du, Jun; He, Xiaoshun; Jiao, Xingyuan; Zhu, Xiaofeng

    2015-01-01

    Gallbladder carcinoma is an aggressive malignancy with high mortality mainly due to the limited potential for curative resection and its resistance to chemotherapeutic agents. Here, we show that the histone deacetylase inhibitors (HDACIs) trichostatin-A (TSA) and suberoylanilide hydroxamic acid (SAHA) reduce the proliferation and induce apoptosis of gallbladder carcinoma cells by suppressing the AKT/mammalian target of rapamycin (mTOR) signaling. Gallbladder carcinoma SGC-996 cells were treated with different concentrations of TSA and SAHA for different lengths of time. Cell proliferation and morphology were assessed with MTT assay and microscopy, respectively. Cell cycle distribution and cell apoptosis were analyzed with flow cytometry. Western blotting was used to detect the proteins related to apoptosis, cell cycle, and the AKT/mTOR signaling pathway. Our data showed that TSA and SAHA reduced SGC-996 cell viability and arrested cell cycle at the G1 phase in a dose- and time-dependent manner. TSA and SAHA promoted apoptosis of SGC-996 cells, down-regulated the expression of cyclin D1, c-Myc and Bmi1, and decreased the phosphorylation of AKT, mTOR p70S6K1, S6 and 4E-BP1. Additionally, the mTOR inhibitor rapamycin further reduced the cell viability of TSA- and SAHA-treated SGC-996 cells and the phosphorylation of mTOR, whereas the mTOR activator 1,2-dioctanoyl-sn-glycero-3-phosphate (C8-PA) exerted the opposite influence. Our results demonstrate that histone deacetylase inhibitors (HDACIs) suppress the proliferation of gallbladder carcinoma cell via inhibition of AKT/mTOR signaling. These findings offer a mechanistic rationale for the application of HDACIs in gallbladder carcinoma treatment.

  18. Msa1 and Msa2 Modulate G1-Specific Transcription to Promote G1 Arrest and the Transition to Quiescence in Budding Yeast

    PubMed Central

    Miles, Shawna; Croxford, Matthew W.; Abeysinghe, Amali P.; Breeden, Linda L.

    2016-01-01

    Yeast that naturally exhaust their glucose source can enter a quiescent state that is characterized by reduced cell size, and high cell density, stress tolerance and longevity. The transition to quiescence involves highly asymmetric cell divisions, dramatic reprogramming of transcription and global changes in chromatin structure and chromosome topology. Cells enter quiescence from G1 and we find that there is a positive correlation between the length of G1 and the yield of quiescent cells. The Swi4 and Swi6 transcription factors, which form the SBF transcription complex and promote the G1 to S transition in cycling cells, are also critical for the transition to quiescence. Swi6 forms a second complex with Mbp1 (MBF), which is not required for quiescence. These are the functional analogues of the E2F complexes of higher eukaryotes. Loss of the RB analogue, Whi5, and the related protein Srl3/Whi7, delays G1 arrest, but it also delays recovery from quiescence. Two MBF- and SBF-Associated proteins have been identified that have little effect on SBF or MBF activity in cycling cells. We show that these two related proteins, Msa1 and Msa2, are specifically required for the transition to quiescence. Like the E2F complexes that are quiescence-specific, Msa1 and Msa2 are required to repress the transcription of many SBF target genes, including SWI4, the CLN2 cyclin and histones, specifically after glucose is exhausted from the media. They also activate transcription of many MBF target genes. msa1msa2 cells fail to G1 arrest and rapidly lose viability upon glucose exhaustion. msa1msa2 mutants that survive this transition are very large, but they attain the same thermo-tolerance and longevity of wild type quiescent cells. This indicates that Msa1 and Msa2 are required for successful transition to quiescence, but not for the maintenance of that state. PMID:27272642

  19. Role of HLA-G1 in trophoblast cell proliferation, adhesion and invasion

    SciTech Connect

    Jiang, Feng; Zhao, Hongxi; Wang, Li; Guo, Xinyu; Wang, Xiaohong; Yin, Guowu; Hu, Yunsheng; Li, Yi; Yao, Yuanqing

    2015-02-27

    Trophoblast cells are important in embryo implantation and fetomaternal tolerance. HLA-G is specifically expressed at the maternal–fetal interface and is a regulator in pregnancy. The aim of the present study was to detect the effect of HLA-G1 on trophoblast cell proliferation, adhesion, and invasion. Human trophoblast cell lines (JAR and HTR-8/SVneo cells) were infected with HLA-G1-expressing lentivirus. After infection, HLA-G1 expression of the cells was detected by western blotting. Cell proliferation was detected by the BrdU assay. The cell cycle and apoptosis of JAR and HTR-8/SVneo cells was measured by flow cytometry (FCM). The invasion of the cells under different conditions was detected by the transwell invasion chamber assay. HLA-G1 didn't show any significant influence on the proliferation, apoptosis, adhesion, and invasion of trophocytes in normal culture conditions. However, HLA-G1 inhibited JAR and HTR-8/SVneo cells invasion induced by hepatocyte growth factor (HGF) under normal oxygen conditions. In conditions of hypoxia, HLA-G1 couldn't inhibit the induction of cell invasion by HGF. HLA-G1 is not an independent factor for regulating the trophocytes. It may play an indirect role in embryo implantation and formation of the placenta. - Highlights: • HLA-G1 could not influence trophocytes under normal conditions. • HLA-G1 inhibited cell invasion induced by HGF under normal oxygen condition. • HLA-G1 could not influence cell invasion under hypoxia conditions.

  20. Resetting the epigenetic histone code in the MRL-lpr/lpr mouse model of lupus by histone deacetylase inhibition.

    PubMed

    Garcia, Benjamin A; Busby, Scott A; Shabanowitz, Jeffrey; Hunt, Donald F; Mishra, Nilamadhab

    2005-01-01

    The baseline level of gene expression varies between healthy controls and systemic lupus erythematosus (SLE) patients, and among SLE patients themselves. These variations may explain the different clinical manifestations and severity of disease observed in SLE. Epigenetic mechanisms, which involve DNA and histone modifications, are predictably associated with distinct transcriptional states. To understand the interplay between various histone modifications, including acetylation and methylation, and lupus disease, we performed differential expression histone modification analysis in splenocytes from the MRL-lpr/lpr mouse model of lupus. Using stable isotope labeling in combination with mass spectrometry, we found global site-specific hypermethylation (except H3 K4 methylation) and hypoacetylation in histone H3 and H4 MRL-lpr/lpr mice compared to control MRL/MPJ mice. Moreover, we have identified novel histone modifications such as H3 K18 methylation, H4 K31 methylation, and H4 K31 acetylation that are differentially expressed in MRL-lpr/lpr mice compared to controls. Finally, in vivo administration of the histone deacetylase inhibitor trichostatin A (TSA) corrected the site-specific hypoacetylation states on H3 and H4 in MRL-lpr/lpr mice with improvement of disease phenotype. Thus, this study is the first to establish the association between aberrant histone codes and pathogenesis of autoimmune disease SLE. These aberrant post-translational histone modifications can therefore be reset with histone deacetylase inhibition in vivo.

  1. Micropatterning of Single Endothelial Cell Shape Reveals a Tight Coupling between Nuclear Volume in G1 and Proliferation

    PubMed Central

    Roca-Cusachs, Pere; Alcaraz, Jordi; Sunyer, Raimon; Samitier, Josep; Farré, Ramon; Navajas, Daniel

    2008-01-01

    Shape-dependent local differentials in cell proliferation are considered to be a major driving mechanism of structuring processes in vivo, such as embryogenesis, wound healing, and angiogenesis. However, the specific biophysical signaling by which changes in cell shape contribute to cell cycle regulation remains poorly understood. Here, we describe our study of the roles of nuclear volume and cytoskeletal mechanics in mediating shape control of proliferation in single endothelial cells. Micropatterned adhesive islands were used to independently control cell spreading and elongation. We show that, irrespective of elongation, nuclear volume and apparent chromatin decondensation of cells in G1 systematically increased with cell spreading and highly correlated with DNA synthesis (percent of cells in the S phase). In contrast, cell elongation dramatically affected the organization of the actin cytoskeleton, markedly reduced both cytoskeletal stiffness (measured dorsally with atomic force microscopy) and contractility (measured ventrally with traction microscopy), and increased mechanical anisotropy, without affecting either DNA synthesis or nuclear volume. Our results reveal that the nuclear volume in G1 is predictive of the proliferative status of single endothelial cells within a population, whereas cell stiffness and contractility are not. These findings show that the effects of cell mechanics in shape control of proliferation are far more complex than a linear or straightforward relationship. Our data are consistent with a mechanism by which spreading of cells in G1 partially enhances proliferation by inducing nuclear swelling and decreasing chromatin condensation, thereby rendering DNA more accessible to the replication machinery. PMID:18326659

  2. Cytoplasmic-Nuclear Trafficking of G1/S Cell Cycle Molecules and Adult Human β-Cell Replication

    PubMed Central

    Fiaschi-Taesch, Nathalie M.; Kleinberger, Jeffrey W.; Salim, Fatimah G.; Troxell, Ronnie; Wills, Rachel; Tanwir, Mansoor; Casinelli, Gabriella; Cox, Amy E.; Takane, Karen K.; Srinivas, Harish; Scott, Donald K.; Stewart, Andrew F.

    2013-01-01

    Harnessing control of human β-cell proliferation has proven frustratingly difficult. Most G1/S control molecules, generally presumed to be nuclear proteins in the human β-cell, are in fact constrained to the cytoplasm. Here, we asked whether G1/S molecules might traffic into and out of the cytoplasmic compartment in association with activation of cell cycle progression. Cdk6 and cyclin D3 were used to drive human β-cell proliferation and promptly translocated into the nucleus in association with proliferation. In contrast, the cell cycle inhibitors p15, p18, and p19 did not alter their location, remaining cytoplasmic. Conversely, p16, p21, and p27 increased their nuclear frequency. In contrast once again, p57 decreased its nuclear frequency. Whereas proliferating β-cells contained nuclear cyclin D3 and cdk6, proliferation generally did not occur in β-cells that contained nuclear cell cycle inhibitors, except p21. Dynamic cytoplasmic-nuclear trafficking of cdk6 was confirmed using green fluorescent protein–tagged cdk6 and live cell imaging. Thus, we provide novel working models describing the control of cell cycle progression in the human β-cell. In addition to known obstacles to β-cell proliferation, cytoplasmic-to-nuclear trafficking of G1/S molecules may represent an obstacle as well as a therapeutic opportunity for human β-cell expansion. PMID:23493571

  3. Transformation abrogates an early G1-phase arrest point required for specification of the Chinese hamster DHFR replication origin.

    PubMed

    Wu, J R; Keezer, S M; Gilbert, D M

    1998-03-16

    The origin decision point (ODP) was originally identified as a distinct point during G1-phase when Chinese hamster ovary (CHO) cell nuclei experience a transition that is required for specific recognition of the dihydrofolate reductase (DHFR) origin locus by Xenopus egg extracts. Passage of cells through the ODP requires a mitogen-independent protein kinase that is activated prior to restriction point control. Here we show that inhibition of an early G1-phase protein kinase pathway by the addition of 2-aminopurine (2-AP) prior to the ODP arrests CHO cells in G1-phase. Transformation with simian virus 40 (SV40) abrogated this arrest point, resulting in the entry of cultured cells into S-phase in the presence of 2-AP and a disruption of the normal pattern of initiation sites at the DHFR locus. Cells treated with 2-AP after the ODP initiated replication specifically within the DHFR origin locus. Transient exposure of transformed cells to 2-AP during the ODP transition also disrupted origin choice, whereas non-transformed cells arrested in G1-phase and then passed through a delayed ODP after removal of 2-AP from the medium. We conclude that mammalian cells have many potential sites at which they can initiate replication. Normally, events occurring during the early G1-phase ODP transition determine which of these sites will be the preferred initiation site. However, if chromatin is exposed to S-phase-promoting factors prior to this transition, mammalian cells, like Xenopus and Drosophila embryos, can initiate replication without origin specification.

  4. Transformation abrogates an early G1-phase arrest point required for specification of the Chinese hamster DHFR replication origin.

    PubMed Central

    Wu, J R; Keezer, S M; Gilbert, D M

    1998-01-01

    The origin decision point (ODP) was originally identified as a distinct point during G1-phase when Chinese hamster ovary (CHO) cell nuclei experience a transition that is required for specific recognition of the dihydrofolate reductase (DHFR) origin locus by Xenopus egg extracts. Passage of cells through the ODP requires a mitogen-independent protein kinase that is activated prior to restriction point control. Here we show that inhibition of an early G1-phase protein kinase pathway by the addition of 2-aminopurine (2-AP) prior to the ODP arrests CHO cells in G1-phase. Transformation with simian virus 40 (SV40) abrogated this arrest point, resulting in the entry of cultured cells into S-phase in the presence of 2-AP and a disruption of the normal pattern of initiation sites at the DHFR locus. Cells treated with 2-AP after the ODP initiated replication specifically within the DHFR origin locus. Transient exposure of transformed cells to 2-AP during the ODP transition also disrupted origin choice, whereas non-transformed cells arrested in G1-phase and then passed through a delayed ODP after removal of 2-AP from the medium. We conclude that mammalian cells have many potential sites at which they can initiate replication. Normally, events occurring during the early G1-phase ODP transition determine which of these sites will be the preferred initiation site. However, if chromatin is exposed to S-phase-promoting factors prior to this transition, mammalian cells, like Xenopus and Drosophila embryos, can initiate replication without origin specification. PMID:9501102

  5. [Histone deacetylase inhibitors: new synergistic third-line option in multiple myeloma].

    PubMed

    Stegmann, Danielle A

    2016-04-01

    Despite advances in drug therapy of the orphan disease multiple myeloma, patients relapse or become refractory to first-line therapy, and the disease remains incurable. Therefore, histone deacetylase inhibitors have emerged as a new class of anti-myeloma drugs, with synergistic results on progression free survival when given in combination to current first-line therapy. Histone deacetylase inhibitors influence gene expression of target genes. Based on results of an extensive multicenter phase III trial, panobinostat was approved by the FDA in February 2015 as the first histone deacetylase inhibitor for the treatment of multiple myeloma. In Europe, panobinostat received marketing authorization by August 2015.

  6. Novel Antiproliferative Chimeric Compounds with Marked Histone Deacetylase Inhibitory Activity

    PubMed Central

    2014-01-01

    Given our interest in finding potential antitumor agents and in view of the multifactorial mechanistic nature of cancer, in the present work, taking advantage of the multifunctional ligands approach, new chimeric molecules were designed and synthesized by combining in single chemical entities structural features of SAHA, targeting histone deacetylases (HDACs), with substituted stilbene or terphenyl derivatives previously obtained by us and endowed with antiproliferative and pro-apoptotic activity. The new chimeric derivatives were characterized with respect to their cytotoxic activity and their effects on cell cycle progression on different tumor cell lines, as well as their HDACs inhibition. Among the other, trans-6 showed the most interesting biological profile, as it exhibited a strong pro-apoptotic activity in tumor cell lines in comparison with both of its parent compounds and a marked HDAC inhibition. PMID:25221651

  7. Histone deacetylase inhibitors reverse gene silencing in Friedreich's ataxia.

    PubMed

    Herman, David; Jenssen, Kai; Burnett, Ryan; Soragni, Elisabetta; Perlman, Susan L; Gottesfeld, Joel M

    2006-10-01

    Expansion of GAA x TTC triplets within an intron in FXN (the gene encoding frataxin) leads to transcription silencing, forming the molecular basis for the neurodegenerative disease Friedreich's ataxia. Gene silencing at expanded FXN alleles is accompanied by hypoacetylation of histones H3 and H4 and trimethylation of histone H3 at Lys9, observations that are consistent with a heterochromatin-mediated repression mechanism. We describe the synthesis and characterization of a class of histone deacetylase (HDAC) inhibitors that reverse FXN silencing in primary lymphocytes from individuals with Friedreich's ataxia. We show that these molecules directly affect the histones associated with FXN, increasing acetylation at particular lysine residues on histones H3 and H4 (H3K14, H4K5 and H4K12). This class of HDAC inhibitors may yield therapeutics for Friedreich's ataxia.

  8. Histone deacetylases as regulators of inflammation and immunity.

    PubMed

    Shakespear, Melanie R; Halili, Maria A; Irvine, Katharine M; Fairlie, David P; Sweet, Matthew J

    2011-07-01

    Histone deacetylases (HDACs) remove an acetyl group from lysine residues of target proteins to regulate cellular processes. Small-molecule inhibitors of HDACs cause cellular growth arrest, differentiation and/or apoptosis, and some are used clinically as anticancer drugs. In animal models, HDAC inhibitors are therapeutic for several inflammatory diseases, but exacerbate atherosclerosis and compromise host defence. Loss of HDAC function has also been linked to chronic lung diseases in humans. These contrasting effects might reflect distinct roles for individual HDACs in immune responses. Here, we review the current understanding of innate and adaptive immune pathways that are regulated by classical HDAC enzymes. The objective is to provide a rationale for targeting (or not targeting) individual HDAC enzymes with inhibitors for future immune-related applications. Copyright © 2011 Elsevier Ltd. All rights reserved.

  9. Histone deacetylases 9 and 10 are required for homologous recombination.

    PubMed

    Kotian, Shweta; Liyanarachchi, Sandhya; Zelent, Arthur; Parvin, Jeffrey D

    2011-03-11

    We tested the role of histone deacetylases (HDACs) in the homologous recombination process. A tissue-culture based homology-directed repair assay was used in which repair of a double-stranded break by homologous recombination results in gene conversion of an inactive GFP allele to an active GFP gene. Our rationale was that hyperacetylation caused by HDAC inhibitor treatment would increase chromatin accessibility to repair factors, thereby increasing homologous recombination. Contrary to expectation, treatment of cells with the inhibitors significantly reduced homologous recombination activity. Using RNA interference to deplete each HDAC, we found that depletion of either HDAC9 or HDAC10 specifically inhibited homologous recombination. By assaying for sensitivity of cells to the interstrand cross-linker mitomycin C, we found that treatment of cells with HDAC inhibitors or depletion of HDAC9 or HDAC10 resulted in increased sensitivity to mitomycin C. Our data reveal an unanticipated function of HDAC9 and HDAC10 in the homologous recombination process.

  10. Lysine Deacetylase Inhibitors in Parasites: Past, Present, and Future Perspectives.

    PubMed

    Hailu, Gebremedhin S; Robaa, Dina; Forgione, Mariantonietta; Sippl, Wolfgang; Rotili, Dante; Mai, Antonello

    2017-06-22

    Current therapies for human parasite infections rely on a few drugs, most of which have severe side effects, and their helpfulness is being seriously compromised by the drug resistance problem. Globally, this is pushing discovery research of antiparasitic drugs toward new agents endowed with new mechanisms of action. By using a "drug repurposing" strategy, histone deacetylase inhibitors (HDACi), which are presently clinically approved for cancer use, are now under investigation for various parasite infections. Because parasitic Zn(2+)- and NAD(+)-dependent HDACs play crucial roles in the modulation of parasite gene expression and many of them are pro-survival for several parasites under various conditions, they are now emerging as novel potential antiparasitic targets. This Perspective summarizes the state of knowledge of HDACi (both class I/II HDACi and sirtuin inhibitors) targeted to the main human parasitic diseases (schistosomiasis, malaria, trypanosomiasis, leishmaniasis, and toxoplasmosis) and provides visions into the main issues that challenge their development as antiparasitic agents.

  11. Histone Deacetylase Inhibitors Preserve Function in Aging Axons

    PubMed Central

    Baltan, Selva

    2012-01-01

    Aging increases the vulnerability of aging white matter to ischemic injury. Histone deacetylase (HDAC) inhibitors preserve young adult white matter structure and function during ischemia by conserving ATP and reducing excitotoxicity. In isolated optic nerve from 12 month old mice, deprived of oxygen and glucose, we show that pan- and Class I specific HDAC inhibitors promote functional recovery of axons. This protection correlates with preservation of axonal mitochondria. The cellular expression of HDAC 3, in the central nervous system (CNS) and HDAC 2 in optic nerve considerably changed with age expanding to more cytoplasmic domains from nuclear compartments suggesting that changes in glial cell protein acetylation may confer protection to aging axons. Our results indicate manipulation of HDAC activities in glial cells may have a universal potential for stroke therapy across age groups. PMID:23050648

  12. Treatment of chronic kidney diseases with histone deacetylase inhibitors

    PubMed Central

    Liu, Na; Zhuang, Shougang

    2015-01-01

    Histone deacetylases (HDACs) induce deacetylation of both histone and non-histone proteins and play a critical role in the modulation of physiological and pathological gene expression. Pharmacological inhibition of HDAC has been reported to attenuate progression of renal fibrogenesis in obstructed kidney and reduce cyst formation in polycystic kidney disease. HDAC inhibitors (HDACis) are also able to ameliorate renal lesions in diabetes nephropathy, lupus nephritis, aristolochic acid nephropathy, and transplant nephropathy. The beneficial effects of HDACis are associated with their anti-fibrosis, anti-inflammation, and immunosuppressant effects. In this review, we summarize recent advances on the treatment of various chronic kidney diseases with HDACis in pre-clinical models. PMID:25972812

  13. Quarter variation and correlations of colostrum albumin, immunoglobulin G1 and G2 in dairy cows.

    PubMed

    Samarütel, Jaak; Baumrucker, Craig R; Gross, Josef J; Dechow, Chad D; Bruckmaier, Rupert M

    2016-05-01

    A high variation in immunoglobulin G1 (IgG1) concentration in first milked quarter colostrum has been reported, but BSA quarter colostrum variation is not known. The occurrence of serum albumin in milk has been attributed to increased blood-milk barrier penetration. Reports of serum albumin binding to the Fc Receptor of the neonate, the receptor thought to be responsible for IgG1 transcytosis, suggested that a correlation with the appearance of IgG1 in colostrum of dairy cows was likely. The objective of the study was to establish the quarter colostrum concentration and mass of immunoglobulins and serum albumin. First colostrum was quarter collected within 4 h of parturition from healthy udders of 31 multiparous dairy cows. Individual quarter colostrum weight was determined and a sample of each was frozen for subsequent analysis. Concentrations of immunoglobulin G1, G2, and BSA were measured by ELISA and total mass of components was calculated. In addition, colostrum was also analysed for L-lactate dehydrogenase activity. Analysis of concentration and mass of BSA, immunoglobulin G1, G2 established that the quarter variations were different by cow, quarter and quarter within cow. Partial correlations corrected for colostrum weight indicated that BSA and IgG2 concentration and mass are closely correlated while that of BSA and IgG1 concentration and mass exhibited no correlation suggesting that BSA and IgG1 may have different transport mechanisms. Interestingly, immunoglobulin G1 and G2 concentration and mass exhibited strong correlations suggesting that also some unknown mechanism of immunoglobulin G2 appearance in colostrum is occurring. Finally, no measured protein exhibited any correlation with the activity of lactate dehydrogenase in colostrum.

  14. Neuroblastoma stem cells - mechanisms of chemoresistance and histone deacetylase inhibitors.

    PubMed

    Khalil, M A; Hrabeta, J; Cipro, S; Stiborova, M; Vicha, A; Eckschlager, T

    2012-01-01

    Cancer stem cells (CSCs) form a small proportion of tumor cells that have stem cell properties: self-renewal capacity, the ability to develop into different lineages and proliferative potential. The interest in CSCs emerged from their expected role in initiation, progression and recurrence of many tumors. They are generally resistant to conventional chemotherapy and radiotherapy. There are two hypotheses about their origin: The first assumes that CSCs may arise from normal stem cells, and the second supposes that differentiated cells acquire the properties of CSCs. Both hypotheses are not mutually exclusive, as it is possible that CSCs have a diverse origin in different tumors. CD133+ cells (CD133 is marker of CSC in some tumors) isolated from NBL, osteosarcoma and Ewing sarcoma cell lines are resistant to cisplatin, carboplatin, etoposide and doxorubicin than the CD133- ones. Being resistant to chemotherapy, there were many attempts to target CSCs epigenetically including the use of histone deacetylase inhibitors. The diverse influence of valproic acid (histone deacetylase inhibitor) on normal and cancer stem cells was proved in different experiments. We have found an increase percentage of CD133+ NBL cells after their incubation with VPA in a dose that does not induce apoptosis. Further researches on CSCs and clinical application for their detection are necessary: (i) to define the CSC function in carcinogenesis, cancer development and their role in metastasis; (ii) to find a specific marker for CSCs in different tumors; (iii) to explain the role of different pathways that determine their behavior and (iv) to explain mechanisms of chemoresistance of CSCs.

  15. Histone deacetylase inhibitor (HDACI) mechanisms of action: emerging insights

    PubMed Central

    Bose, Prithviraj; Dai, Yun; Grant, Steven

    2014-01-01

    Initially regarded as “epigenetic modifiers” acting predominantly through chromatin remodeling via histone acetylation, HDACIs, alternatively referred to as lysine deacetylase or simply deacetylase inhibitors, have since been recognized to exert multiple cytotoxic actions in cancer cells, often through acetylation of non-histone proteins. Some well-recognized mechanisms of HDACI lethality include, in addition to relaxation of DNA and de-repression of gene transcription, interference with chaperone protein function, free radical generation, induction of DNA damage, up-regulation of endogenous inhibitors of cell cycle progression, e.g., p21, and promotion of apoptosis. Intriguingly, this class of agents is relatively selective for transformed cells, at least in pre-clinical studies. In recent years, additional mechanisms of action of these agents have been uncovered. For example, HDACIs interfere with multiple DNA repair processes, as well as disrupt cell cycle checkpoints, critical to the maintenance of genomic integrity in the face of diverse genotoxic insults. Despite their pre-clinical potential, the clinical use of HDACIs remains restricted to certain subsets of T-cell lymphoma. Currently, it appears likely that the ultimate role of these agents will lie in rational combinations, only a few of which have been pursued in the clinic to date. This review focuses on relatively recently identified mechanisms of action of HDACIs, with particular emphasis on those that relate to the DNA damage response (DDR), and discuss synergistic strategies combining HDACIs with several novel targeted agents that disrupt the DDR or antagonize anti-apoptotic proteins that could have implications for the future use of HDACIs in patients with cancer. PMID:24769080

  16. Histone Deacetylases with Antagonistic Roles in Saccharomyces cerevisiae Heterochromatin Formation.

    PubMed

    Thurtle-Schmidt, Deborah M; Dodson, Anne E; Rine, Jasper

    2016-09-01

    As the only catalytic member of the Sir-protein gene-silencing complex, Sir2's catalytic activity is necessary for silencing. The only known role for Sir2's catalytic activity in Saccharomyces cerevisiae silencing is to deacetylate N-terminal tails of histones H3 and H4, creating high-affinity binding sites for the Sir-protein complex, resulting in association of Sir proteins across the silenced domain. This histone deacetylation model makes the simple prediction that preemptively removing Sir2's H3 and H4 acetyl substrates, by mutating these lysines to unacetylatable arginines, or removing the acetyl transferase responsible for their acetylation, should restore silencing in the Sir2 catalytic mutant. However, this was not the case. We conducted a genetic screen to explore what aspect of Sir2's catalytic activity has not been accounted for in silencing. Mutation of a nonsirtuin histone deacetylase, Rpd3, restored Sir-protein-based silencing in the absence of Sir2's catalytic activity. Moreover, this antagonism could be mediated by either the large or the small Rpd3-containing complex. Interestingly, this restoration of silencing appeared independent of any known histone H3 or H4 substrates of Rpd3 Investigation of Sir-protein association in the Rpd3 mutant revealed that the restoration of silencing was correlated with an increased association of Sir proteins at the silencers, suggesting that Rpd3 was an antagonist of Sir2's function in nucleation of Sir proteins to the silencer. Additionally, restoration of silencing by Rpd3 was dependent on another sirtuin family member, Hst3, indicating multiple antagonistic roles for deacetylases in S. cerevisiae silencing. Copyright © 2016 by the Genetics Society of America.

  17. Shrimp chitin as substrate for fungal chitin deacetylase.

    PubMed

    Win, N N; Stevens, W F

    2001-10-01

    The fungal chitin deacetylases (CDA) studied so far are able to perform heterogeneous enzymatic deacetylation on their solid substrate, but only to a limited extent. Kinetic data show that about 5-10% of the N-acetyl glucosamine residues are deacetylated rapidly. Thereafter enzymatic deacetylation is slow. In this study, chitin was exposed to various physical and chemical conditions such as heating, sonicating, grinding, derivatization and interaction with saccharides and presented as a substrate to the CDA of the fungus Absidia coerulea. None of these treatments of the substrate resulted in a more efficient enzymatic deacetylation. Dissolution of chitin in specific solvents followed by fast precipitation by changing the composition of the solvent was not successful either in making microparticles that would be more accessible to the enzyme. However, by treating chitin in this way, a decrystallized chitin with a very small particle size called superfine (SF) chitin could be obtained. This SF chitin, pretreated with 18% formic acid, appeared to be a good substrate for fungal deacetylase. This was confirmed both by enzyme-dependent deacetylation measured by acetate production as well as by isolation and assay for the degree of deacetylation (DD). In this way chitin (10% DD) was deacetylated by the enzyme into chitosan with DD of 90%. The formic acid treatment reduced the molecular weight of the polymeric chain from 2x10(5) in chitin to 1.2 x 10(4) in the chitosan product. It is concluded that nearly complete enzymatic deacetylation has been demonstrated for low-molecular chitin.

  18. A distinct G1 step required to specify the Chinese hamster DHFR replication origin.

    PubMed

    Wu, J R; Gilbert, D M

    1996-03-01

    Nuclei isolated from Chinese hamster ovary (CHO) cells at various times during the G1 phase of the cell cycle were stimulated to enter S phase by incubation in Xenopus egg cytosol. Replication of DNA initiated within the dihydrofolate reductase (DHFR) origin locus in nuclei isolated late in G1, but at random sites in nuclei isolated early in G1. A discrete transition point occurred 3 to 4 hours after metaphase. Neither replication licensing nor nuclear assembly was sufficient for origin recognition. Thus, a distinct cell cycle-regulated event in the nucleus restricts the initiation of replication to specific sites downstream of the DHFR gene.

  19. Arabidopsis thaliana histone deacetylase 1 (AtHD1) is localized in euchromatic regions and demonstrates histone deacetylase activity in vitro

    PubMed Central

    Fong, Paulus M; Tian, Lu; Chen, Z Jeffrey

    2007-01-01

    Arabidopsis thaliana histone deacetylase 1 (AtHD1 or AtHDA19), a homolog of yeast RPD3, is a global regulator of many physiological and developmental processes in plants. In spite of the genetic evidence for a role of AtHD1 in plant gene regulation and development, the biochemical and cellular properties of AtHD1 are poorly understood. Here we report cellular localization patterns of AtHD1 in vivo and histone deacetylase activity in vitro. The transient and stable expression of a green fluorescent (GFP)-tagged AtHD1 in onion cells and in roots, seeds and leaves of the transgenic Arabidopsis, respectively, revealed that AtHD1 is localized in the nucleus presumably in the euchromatic regions and excluded from the nucleolus. The localization patterns of AtHD1 are different from those of AtHD2 and AtHDA6 that are involved in nucleolus formation and silencing of transgenes and repeated DNA elements, respectively. In addition, a histone deacetylase activity assay showed that the recombinant AtHD1 produced in bacteria demonstrated a specific histone deacetylase activity in vitro. The data suggest that AtHD1 is a nuclear protein and possesses histone deacetylase activities responsible for global transcriptional regulation important to plant growth and development. PMID:16699543

  20. Moments of the Spin Structure Functions g1p and g1d for 0.05 < Q2 < 3.0 GeV2

    SciTech Connect

    Prok, Yelena; Bosted, Peter; Burkert, Volker; Deur, Alexandre; Dharmawardane, Kahanawita; Dodge, Gail; Griffioen, Keith; Kuhn, Sebastian; Minehart, Ralph; Adams, Gary; Amaryan, Moscov; Amaryan, Moskov; Anghinolfi, Marco; Asryan, G.; Audit, Gerard; Avagyan, Harutyun; Baghdasaryan, Hovhannes; Baillie, Nathan; Ball, J.P.; Ball, Jacques; Baltzell, Nathan; Barrow, Steve; Battaglieri, Marco; Beard, Kevin; Bedlinskiy, Ivan; Bektasoglu, Mehmet; Bellis, Matthew; Benmouna, Nawal; Berman, Barry; Biselli, Angela; Blaszczyk, Lukasz; Boyarinov, Sergey; Bonner, Billy; Bouchigny, Sylvain; Bradford, Robert; Branford, Derek; Briscoe, William; Brooks, William; Bultmann, S.; Bueltmann, Stephen; Butuceanu, Cornel; Calarco, John; Careccia, Sharon; Carman, Daniel; Casey, Liam; Cazes, Antoine; Chen, Shifeng; Cheng, Lu; Cole, Philip; Collins, Patrick; Coltharp, Philip; Cords, Dieter; Corvisiero, Pietro; Crabb, Donald; Crede, Volker; Cummings, John; Dale, Daniel; Dashyan, Natalya; De Masi, Rita; De Vita, Raffaella; De Sanctis, Enzo; Degtiarenko, Pavel; Denizli, Haluk; Dennis, Lawrence; Dhuga, Kalvir; Dickson, Richard; Djalali, Chaden; Doughty, David; Dugger, Michael; Dytman, Steven; Dzyubak, Oleksandr; Egiyan, Hovanes; Egiyan, Kim; Elfassi, Lamiaa; Elouadrhiri, Latifa; Eugenio, Paul; Fatemi, Renee; Fedotov, Gleb; Feldman, Gerald; Fersch, Robert; Feuerbach, Robert; Forest, Tony; Fradi, Ahmed; Funsten, Herbert; Garcon, Michel; Gavalian, Gagik; Gevorgyan, Nerses; Gilfoyle, Gerard; Giovanetti, Kevin; Girod, Francois-Xavier; Goetz, John; Golovach, Evgeny; Gothe, Ralf; Guidal, Michel; Guillo, Matthieu; Guler, Nevzat; Guo, Lei; Gyurjyan, Vardan; Hadjidakis, Cynthia; Hafidi, Kawtar; Hakobyan, Hayk; Hanretty, Charles; Hardie, John; Hassall, Neil; Heddle, David; Hersman, F.; Hicks, Kenneth; Hleiqawi, Ishaq; Holtrop, Maurik; Huertas, Marco; Hyde, Charles; Ilieva, Yordanka; Ireland, David; Ishkhanov, Boris; Isupov, Evgeny; Ito, Mark; Jenkins, David; Jo, Hyon-Suk; Johnstone, John; Joo, Kyungseon; Juengst, Henry; Kalantarians, Narbe; Keith, Christopher; Kellie, James; Khandaker, Mahbubul; Kim, Kui; Kim, Kyungmo; Kim, Wooyoung; Klein, Andreas; Klein, Franz; Klusman, Mike; Kossov, Mikhail; Krahn, Zebulun; Kramer, Laird; Kubarovsky, Valery; Kuhn, Joachim; Kuleshov, Sergey; Kuznetsov, Viacheslav; Lachniet, Jeff; Laget, Jean; Langheinrich, Jorn; Lawrence, Dave; Lima, Ana; Livingston, Kenneth; Lu, Haiyun; Lukashin, K.; MacCormick, Marion; Marchand, Claude; Markov, Nikolai; Mattione, Paul; McAleer, Simeon; McKinnon, Bryan; McNabb, John; Mecking, Bernhard; Mestayer, Mac; Meyer, Curtis; Mibe, Tsutomu; Mikhaylov, Konstantin; Mirazita, Marco; Miskimen, Rory; Mokeev, Viktor; Morand, Ludyvine; Moreno, Brahim; Moriya, Kei; Morrow, Steven; Moteabbed, Maryam; Mueller, James; Munevar Espitia, Edwin; Mutchler, Gordon; Nadel-Turonski, Pawel; Nasseripour, Rakhsha; Niccolai, Silvia; Niculescu, Gabriel; Niculescu, Maria-Ioana; Niczyporuk, Bogdan; Niroula, Megh; Niyazov, Rustam; Nozar, Mina; O'Rielly, Grant; Osipenko, Mikhail; Ostrovidov, Alexander; Park, Kijun; Pasyuk, Evgueni; Paterson, Craig; Anefalos Pereira, S.; Philips, Sasha; Pierce, J.; Pivnyuk, Nikolay; Pocanic, Dinko; Pogorelko, Oleg; Popa, Iulian; Pozdnyakov, Sergey; Preedom, Barry; Price, John; Procureur, Sebastien; Protopopescu, Dan; Qin, Liming; Raue, Brian; Riccardi, Gregory; Ricco, Giovanni; Ripani, Marco; Ritchie, Barry; Rosner, Guenther; Rossi, Patrizia; Rowntree, David; Rubin, Philip; Sabatie, Franck; Salamanca, Julian; Salgado, Carlos; Santoro, Joseph; Sapunenko, Vladimir; Schumacher, Reinhard; Seely, Mikell; Serov, Vladimir; Sharabian, Youri; Sharov, Dmitri; Shaw, Jeffrey; Shvedunov, Nikolay; Skabelin, Alexander; Smith, Elton; Smith, Lee; Sober, Daniel; Sokhan, Daria; Stavinskiy, Aleksey; Stepanyan, Samuel; Stepanyan, Stepan; Stokes, Burnham; Stoler, Paul; Strakovski, Igor; Strauch, Steffen; Suleiman, Riad; Taiuti, Mauro; Tedeschi, David; Tkabladze, Avtandil; Tkachenko, Svyatoslav; Todor, Luminita; Ungaro, Maurizio; V

    2009-02-01

    The spin structure functions $g_1$ for the proton and the deuteron have been measured over a wide kinematic range in $x$ and \\Q2 using 1.6 and 5.7 GeV longitudinally polarized electrons incident upon polarized NH$_3$ and ND$_3$ targets at Jefferson Lab. Scattered electrons were detected in the CEBAF Large Acceptance Spectrometer, for $0.05 < Q^2 < 5 $\\ GeV$^2$ and $W < 3$ GeV. The first moments of $g_1$ for the proton and deuteron are presented -- both have a negative slope at low \\Q2, as predicted by the extended Gerasimov-Drell-Hearn sum rule. The first result for the generalized forward spin polarizability of the proton $\\gamma_0^p$ is also reported, and shows evidence of scaling above $Q^2$ = 1.5 GeV$^2$. Although the first moments of $g_1$ are consistent with Chiral Perturbation Theory (\\ChPT) calculations up to approximately $Q^2 = 0.06$ GeV$^2$, a significant discrepancy is observed between the $\\gamma_0^p$ data and \\ChPT\\ for $\\gamma_0^p$,even at the lowest \\Q2.

  1. Cation exchange surface-mediated denaturation of an aglycosylated immunoglobulin (IgG1).

    PubMed

    Gillespie, Ron; Nguyen, Thao; Macneil, Sean; Jones, Laurie; Crampton, Shon; Vunnum, Suresh

    2012-08-17

    Cation exchange chromatography of an aglycosylated IgG1 resulted in two distinct peaks during gradient elution. The early eluting peak contained <1% high molecular weight (HMW) species, while the later peak contained 23% HMW species. Analysis by hydrogen-deuterium exchange and Fourier transform infrared spectroscopy (FTIR) indicated that aggregate formation and generation of the second peak were caused by antibody denaturation on the resin surface. Denaturation and HMW generation was increased by the use of strong cation exchange media, by increasing antibody residence time on the exchanger, or increasing temperature. Denaturation and HMW generation was reduced by increasing pH or ionic strength, by the use of preferentially excluded solutes such as citrate or glycine and controlled entirely by addition of 125 mM arginine to the process buffers. This leads to the hypothesis that denaturation and HMW generation of this antibody can be managed by reducing the strength of binding, by increasing its conformational stability, or by suppressing non-native protein-protein interactions. The glycosylated version of this antibody exhibited less than 2% denatured form, suggesting that glycosylation contributes significantly to the stability of this antibody. These findings may be helpful in managing aggregation in other antibodies, and particularly useful in developing purification processes for aglycosylated antibodies.

  2. Liquid high concentration IgG1 antibody formulations by precipitation.

    PubMed

    Matheus, Susanne; Friess, Wolfgang; Schwartz, Daniel; Mahler, Hanns-Christian

    2009-09-01

    A manufacturing approach for liquid high concentration antibody formulations based on precipitation and subsequent re-dissolution was investigated. IgG1 antibody solutions were concentrated from 20 to 100 mg/mL by intermediate precipitation, with a recovery exceeding 95%, retention of the native secondary structure and binding activity as well as adequate stability. Quantitative, reproducible precipitation was performed using 1.45 M ammonium sulphate (pH 5.5 and 8.0), 0.67 M sodium citrate (pH 8.0) and 9% (w/v) PEG 4000 (pH 5.5 and 8.0). Scalability was confirmed from 1 to 100 mL. The concentrations achievable in the re-dissolution step were less affected by the re-dissolution medium, but limited by the residual precipitant. Both, improved removal of remaining precipitant liquid and larger precipitation scales were successful in increasing the final protein concentration. SEC and turbidity analysis directly after re-dissolution indicated that similar protein qualities were obtained, independent from the precipitant used. However, increased aggregate formation was observed after short term storage of the precipitated protein particles at either 2-8 degrees C or ambient temperature. An accelerated mechanical and thermal stability program verified comparable stability of the re-dissolved liquid 100 mg/mL formulations produced by intermediate precipitation to a control formulation obtained by standard ultrafiltration.

  3. Bacillus amyloliquefaciens G1: A Potential Antagonistic Bacterium against Eel-Pathogenic Aeromonas hydrophila

    PubMed Central

    Cao, Haipeng; He, Shan; Wei, Ruopeng; Diong, Marek; Lu, Liqun

    2011-01-01

    Recent studies have revealed that the use of probiotics is an alternative to control marine aeromonas. However, few probiotics are available against Aeromonas hydrophila infections in eels. In the present study, a potential antagonistic strain G1 against the eel-pathogenic A. hydrophila was isolated from sediment underlying brackish water. Its extracellular products with antibacterial activities were shown to be stable under wide range of pH, temperature, and proteinase K. It was initially identified as Bacillus amyloliquefaciens using API identification kits and confirmed to be B. amyloliquefaciens strain (GenBank accession number DQ422953) by phylogenetic analysis. In addition, it was shown to be safe for mammalians, had a wide anti-A. hydrophila spectrum, and exhibited significant effects on inhibiting the growth of the eel-pathogenic A. hydrophila both in vitro and in vivo. To the best of our knowledge, this is the first report on a promising antagonistic Bacillus amyloliquefaciens strain from brackish water sediment against eel-pathogenic A. hydrophila. PMID:21754944

  4. Histone hypoacetylation and increased histone deacetylases in peripheral blood mononuclear cells from patients with Graves' disease.

    PubMed

    Yan, Ni; Zhou, Jiao-zhen; Zhang, Jin-an; Cai, Tiantian; Zhang, Wen; Wang, Yuan; Muhali, Fatuma-Said; Guan, Lijuan; Song, Rong-hua

    2015-10-15

    The objective of this study was to investigate histone modification patterns in peripheral blood mononuclear cells (PBMCs) of patients with Graves' disease (GD). Thirty GD patients and 20 healthy controls were enrolled in this study. Global histone H3/H4 acetylation levels of PBMCs in all subjects were detected by enzyme-linked immunosorbent assay. mRNA levels of histone-related chromatin modifier genes were measured by real-time quantitative reverse transcription-polymerase chain reaction. Global histone H4 acetylation level in PBMCs of GD patients was significantly decreased compared with controls (p=0.005). The mRNA expression of histone deacetylases HDAC1 and HDAC2 were significantly increased in PBMCs of GD patients compared with controls (p=0.004 and 0.018; respectively). No significant difference was observed either in SIRT1 or in HATs mRNA including p300, CREBBP between GD patients and controls (p>0.05). Our findings firstly suggested that histone acetylation modifications are aberrant in PBMCs of GD patients, possibly due to the deregulation of epigenetic modifier genes. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  5. miR-150 inhibits proliferation and tumorigenicity via retarding G1/S phase transition in nasopharyngeal carcinoma

    PubMed Central

    Li, Xiangyong; Liu, Fumei; Lin, Bihua; Luo, Haiqing; Liu, Meilian; Wu, Jinhua; Li, Caihong; Li, Ronggang; Zhang, Xin; Zhou, Keyuan; Ren, Dong

    2017-01-01

    Cancer cells are characterized by a pathological manifestation of uncontrolled proliferation, which results in tumor formation. Therefore, it is necessary to improve understanding of the underlying mechanism of cell cycle control. Here, we report that miR-150 is downregulated in nasopharyngeal carcinoma tissues and cells. Upregulation of miR-150 suppresses nasopharyngeal carcinoma (NPC) cell proliferation and induces G1/S arrest in vitro, and inhibits tumorigenesis in vivo. Conversely, silencing miR-150 yields the opposite effect. Our results further demonstrate that miR-150 retards nasopharyngeal carcinoma cell proliferation and G1/S transition via targeting multiple cell cycle-related genes, including CCND1, CCND2, CDK2 and CCNE2. Therefore, our results uncover a novel mechanistic understanding of miR-150-mediated tumor suppression in NPC, which will facilitate the development of effective cancer therapies against nasopharyngeal carcinoma. PMID:28350089

  6. Hcp and VgrG1 are secreted components of the Helicobacter hepaticus type VI secretion system and VgrG1 increases the bacterial colitogenic potential.

    PubMed

    Bartonickova, Lucie; Sterzenbach, Torsten; Nell, Sandra; Kops, Friederike; Schulze, Jessika; Venzke, Annika; Brenneke, Birgit; Bader, Sophie; Gruber, Achim D; Suerbaum, Sebastian; Josenhans, Christine

    2013-06-01

    The enterohepatic Epsilonproteobacterium Helicobacter hepaticus persistently colonizes the intestine of mice and causes chronic inflammatory symptoms in susceptible mouse strains. The bacterial factors causing intestinal inflammation are poorly characterized. A large genomic pathogenicity island, HHGI1, which encodes components of a type VI secretion system (T6SS), was previously shown to contribute to the colitogenic potential of H. hepaticus. We have now characterized the T6SS components Hcp, VgrG1, VgrG2 and VgrG3, encoded on HHGI1, including the potential impact of the T6SS on intestinal inflammation in a mouse T-cell transfer model. The H. hepaticus T6SS components were expressed during the infection and secreted in a T6SS-dependent manner, when the bacteria were cultured either in the presence or in the absence of mouse intestinal epithelial cells. Mutants deficient in VgrG1 displayed a significantly lower colitogenic potential in T-cell-transferred C57BL/6 Rag2(-/-) mice, despite an unaltered ability to colonize mice persistently. Intestinal microbiota analyses demonstrated only minor changes in mice infected with wild-typeH. hepaticus as compared with mice infected with VgrG1-deficient isogenic bacteria. In addition, competitive assays between both wild-type and T6SS-deficient H. hepaticus, and between wild-type H. hepaticus and Campylobacter jejuni or Enterobacteriaceae species did not show an effect of the T6SS on interbacterial competitiveness. Therefore, we suggest that microbiota alterations did not play a major role in the changes of pro-inflammatory potential mediated by the T6SS. Cellular innate pro-inflammatory responses were increased by the secreted T6SS proteins VgrG1 and VgrG2. We therefore concluded that the type VI secretion component VgrG1 can modulate and specifically exacerbate the innate pro-inflammatory effect of the chronic H. hepaticus infection.

  7. Redox-mediated bypass of restriction point via skipping of G1pm

    PubMed Central

    Hoffman, Arnold; Greene, James J; Spetner, Lee M; Burke, Michael

    2006-01-01

    Background It is well known that cancer cells bypass the restriction point, R, and undergo uncontrolled cell proliferation. Hypothesis and evidence We suggest here that fibrosarcoma cells enter G1ps directly from M, skipping G1pm, hence bypassing R, in response to redox modulation. Evidence is presented from the published literature that demonstrate a shortening of the cycle period of transformed fibroblasts (SV-3T3) compared to the nontransformed 3T3 fibroblasts, corresponding to the duration of G1pm in the 3T3 fibroblasts. Evidence is also presented that demonstrate that redox modulation can induce the CUA-4 fibroblasts to bypass R, resulting in a cycle period closely corresponding to the cycle period of fibrosarcoma cells (HT1080). Conclusion The evidence supports our hypothesis that a low internal redox potential can cause fibrosarcoma cells to skip the G1pm phase of the cell cycle. PMID:16867189

  8. AMBIENT PARTICULATE MATTER INDUCES ALVEOLAR EPITHELIAL CELL CYCLE ARREST: ROLE OF G1 CYCLINS

    PubMed Central

    Zhang, Jingmei; Ghio, Andrew J.; Gao, Mingxing; Wei, Ke; Rosen, Glenn D.; Upadhyay, Daya

    2007-01-01

    We hypothesized that the ambient air pollution particles (PM) induce cell cycle arrest in alveolar epithelial cells (AEC). Exposure of PM (25μg/cm2) to AEC induced cells cycle arrest in G1 phase, inhibited DNA synthesis, blocked cell proliferation and caused decrease in cyclin E, A, D1 and Cyclin E- cyclin-dependent kinase(CDK)-2 kinase activity after 4h. PM induced upregulation of CDK inhibitor, p21 protein and p21 activity in AEC. SiRNAp21 blocked PM–induced downregulation of cyclins and AEC G1 arrest. Accordingly, we provide the evidence that PM induces AEC G1 arrest by altered regulation of G1 cyclins and CDKs. PMID:17977533

  9. Ambient particulate matter induces alveolar epithelial cell cycle arrest: role of G1 cyclins.

    PubMed

    Zhang, Jingmei; Ghio, Andrew J; Gao, Mingxing; Wei, Ke; Rosen, Glenn D; Upadhyay, Daya

    2007-11-13

    We hypothesized that the ambient air pollution particles (particulate matter; PM) induce cell cycle arrest in alveolar epithelial cells (AEC). Exposure of PM (25microg/cm(2)) to AEC induced cells cycle arrest in G1 phase, inhibited DNA synthesis, blocked cell proliferation and caused decrease in cyclin E, A, D1 and Cyclin E- cyclin-dependent kinase (CDK)-2 kinase activity after 4h. PM induced upregulation of CDK inhibitor, p21 protein and p21 activity in AEC. SiRNAp21 blocked PM-induced downregulation of cyclins and AEC G1 arrest. Accordingly, we provide the evidence that PM induces AEC G1 arrest by altered regulation of G1 cyclins and CDKs.

  10. IgG1 variations in the colostrum of Holstein dairy cows.

    PubMed

    Le Cozler, Y; Guatteo, R; Le Dréan, E; Turban, H; Leboeuf, F; Pecceu, K; Guinard-Flament, J

    2016-02-01

    High-immune quality colostrum (IgG1 concentration ⩾50 g/l) is crucial for the health and development of the young calf. Studies on colostrum quality tend to focus on external factors such as breed, parity or dry period length, but few have focused on within-cow variations. Here we ran experiments to gain a deeper insight into within-cow variation in IgG1 concentrations in dairy cow colostrum. Trials were performed in an experimental farm, located in the Western part of France. Colostrum from each quarter and a composite sample (mix of four quarters) were concomitantly collected on 77 Holstein dairy cows just after calving to assess the influence of sample type on IgG1 concentrations. Variation in IgG1 concentrations during the first milking was studied on samples from nine cows collected every minute from the start of milking. Repeatability of colostral IgG1 concentration was estimated from 2009 and 2010 data on 16 healthy cows. IgG1 concentrations were tested using a radial immunodiffusion method. Sensitivity and specificity were similar regardless of sample type tested (individual quarter or composite milk). Mean average IgG1 concentration was 54.1 g/l in composite colostrum, and was significantly higher in hind quarter teats (56.2 g/l) than front quarter teats (53.1 g/l). Average IgG1 concentration did not change significantly during colostrum milking, and the variations observed (15% or less) were likely due to the laboratory method (CV 15%). IgG1 concentrations in dam colostrum increased slightly from 2009 to 2010 due to BW and parity effects. In 56% of cases, colostrum quality could have been assessed on either individual or composite colostrum samples collected at any time during the first milking without affecting the reliability of the measurement. However, in other cases, differences were significant enough to mean that estimates of average IgG1 concentration in colostrum from any one quarter would not be reliable. It is concluded that colostrum quality

  11. APOL1-G1 in Nephrocytes Induces Hypertrophy and Accelerates Cell Death.

    PubMed

    Fu, Yulong; Zhu, Jun-Yi; Richman, Adam; Zhang, Yi; Xie, Xuefang; Das, Jharna R; Li, Jinliang; Ray, Patricio E; Han, Zhe

    2017-04-01

    People of African ancestry carrying certain APOL1 mutant alleles are at elevated risk of developing renal diseases. However, the mechanisms underlying APOL1-associated renal diseases are unknown. Because the APOL1 gene is unique to humans and some primates, new animal models are needed to understand the function of APOL1 in vivo We generated transgenic Drosophila fly lines expressing the human APOL1 wild type allele (G0) or the predominant APOL1 risk allele (G1) in different tissues. Ubiquitous expression of APOL1 G0 or G1 in Drosophila induced lethal phenotypes, and G1 was more toxic than was G0. Selective expression of the APOL1 G0 or G1 transgene in nephrocytes, fly cells homologous to mammalian podocytes, induced increased endocytic activity and accumulation of hemolymph proteins, dextran particles, and silver nitrate. As transgenic flies with either allele aged, nephrocyte function declined, cell size increased, and nephrocytes died prematurely. Compared with G0-expressing cells, however, G1-expressing cells showed more dramatic phenotypes, resembling those observed in cultured mammalian podocytes overexpressing APOL1-G1. Expressing the G0 or G1 APOL1 transgene in nephrocytes also impaired the acidification of organelles. We conclude that expression of an APOL1 transgene initially enhances nephrocyte function, causing hypertrophy and subsequent cell death. This new Drosophila model uncovers a novel mechanism by which upregulated expression of APOL1-G1 could precipitate renal disease in humans. Furthermore, this model may facilitate the identification of APOL1-interacting molecules that could serve as new drug targets to treat APOL1-associated renal diseases.

  12. Dillapiol and Apiol as specific inhibitors of the biosynthesis of aflatoxin G1 in Aspergillus parasiticus.

    PubMed

    Razzaghi-Abyaneh, Mehdi; Yoshinari, Tomoya; Shams-Ghahfarokhi, Masoomeh; Rezaee, Mohammad-Bagher; Nagasawa, Hiromichi; Sakuda, Shohei

    2007-09-01

    Dillapiol was isolated from the essential oil of dill as a specific inhibitor of aflatoxin G1 production. It inhibited aflatoxin G1 production by Aspergillus parasiticus with an IC50 value of 0.15 microM without inhibiting aflatoxin B1 production or fungal growth. Apiol and myristicin, congeners of dillapiol, showed similar activity with IC50 values of 0.24 and 3.5 microM, respectively.

  13. Comprehensive phenotypic analysis of knockout mice deficient in cyclin G1 and cyclin G2

    PubMed Central

    Ohno, Shouichi; Ikeda, Jun-ichiro; Naito, Yoko; Okuzaki, Daisuke; Sasakura, Towa; Fukushima, Kohshiro; Nishikawa, Yukihiro; Ota, Kaori; Kato, Yorika; Wang, Mian; Torigata, Kosuke; Kasama, Takashi; Uchihashi, Toshihiro; Miura, Daisaku; Yabuta, Norikazu; Morii, Eiichi; Nojima, Hiroshi

    2016-01-01

    Cyclin G1 (CycG1) and Cyclin G2 (CycG2) play similar roles during the DNA damage response (DDR), but their detailed roles remain elusive. To investigate their distinct roles, we generated knockout mice deficient in CycG1 (G1KO) or CycG2 (G2KO), as well as double knockout mice (DKO) deficient in both proteins. All knockouts developed normally and were fertile. Generation of mouse embryonic fibroblasts (MEFs) from these mice revealed that G2KO MEFs, but not G1KO or DKO MEFs, were resistant to DNA damage insults caused by camptothecin and ionizing radiation (IR) and underwent cell cycle arrest. CycG2, but not CycG1, co-localized with γH2AX foci in the nucleus after γ-IR, and γH2AX-mediated DNA repair and dephosphorylation of CHK2 were delayed in G2KO MEFs. H2AX associated with CycG1, CycG2, and protein phosphatase 2A (PP2A), suggesting that γH2AX affects the function of PP2A via direct interaction with its B’γ subunit. Furthermore, expression of CycG2, but not CycG1, was abnormal in various cancer cell lines. Kaplan–Meier curves based on TCGA data disclosed that head and neck cancer patients with reduced CycG2 expression have poorer clinical prognoses. Taken together, our data suggest that reduced CycG2 expression could be useful as a novel prognostic marker of cancer. PMID:27982046

  14. Commercial geophysical well logs from the USW G-1 drill hole, Nevada Test Site, Nevada

    USGS Publications Warehouse

    Muller, D.C.; Kibler, J.E.

    1983-01-01

    Drill hole USW G-1 was drilled at Yucca Mountain, Nevada Test Site, Nevada, as part of the ongoing exploration program for the Nevada Nuclear Waste Storage Investigations. Contract geophysical well logs run at USW G-1 show only limited stratigraphic correlations, but correlate reasonably well with the welding of the ash-flow and ash-fall tuffs. Rocks in the upper part of the section have highly variable physical properties, but are more uniform and predictably lower in the section.

  15. The multifaceted influence of histone deacetylases on DNA damage signalling and DNA repair

    PubMed Central

    Roos, Wynand Paul; Krumm, Andrea

    2016-01-01

    Histone/protein deacetylases play multiple roles in regulating gene expression and protein activation and stability. Their deregulation during cancer initiation and progression cause resistance to therapy. Here, we review the role of histone deacetylases (HDACs) and the NAD+ dependent sirtuins (SIRTs) in the DNA damage response (DDR). These lysine deacetylases contribute to DNA repair by base excision repair (BER), nucleotide excision repair (NER), mismatch repair (MMR), non-homologous end joining (NHEJ), homologous recombination (HR) and interstrand crosslink (ICL) repair. Furthermore, we discuss possible mechanisms whereby these histone/protein deacetylases facilitate the switch between DNA double-strand break (DSB) repair pathways, how SIRTs play a central role in the crosstalk between DNA repair and cell death pathways due to their dependence on NAD+, and the influence of small molecule HDAC inhibitors (HDACi) on cancer cell resistance to genotoxin based therapies. Throughout the review, we endeavor to identify the specific HDAC targeted by HDACi leading to therapy sensitization. PMID:27738139

  16. Deacetylase inhibitors as a novel modality in the treatment of multiple myeloma.

    PubMed

    Richardson, Paul G; Moreau, Philippe; Laubach, Jacob P; Maglio, Michelle E; Lonial, Sagar; San-Miguel, Jesus

    2017-03-01

    Deacetylase enzymes remove acetyl groups from histone and nonhistone proteins. Dysregulation of deacetylase activity is a hallmark of malignancy, including multiple myeloma (MM). Deacetylase inhibitors (DACi) cause epigenetic modification and inhibition of the aggresome pathway, resulting in death of MM cells. Panobinostat, a pan-DACi, has shown significant clinical benefit and is the first DACi approved for the treatment of MM. It is approved for use in combination with bortezomib and dexamethasone for the treatment of patients with relapsed or relapsed and refractory MM who have received ≥2 prior regimens including bortezomib and an immunomodulatory drug. Ricolinostat and ACY-241, which selectively inhibit HDAC6 and the aggresome pathway, are currently being studied in combination with dexamethasone and bortezomib or an immunomodulatory drug for the treatment of relapsed and refractory MM. In this review, we discuss the data from key clinical trials investigating deacetylase inhibitors as novel treatment options for MM.

  17. Evaluation of the hydrometer for testing immunoglobulin G1 concentrations in Holstein colostrum.

    PubMed

    Pritchett, L C; Gay, C C; Hancock, D D; Besser, T E

    1994-06-01

    Hydrometer measurement in globulin and IgG1 concentration measured by the radial immunodiffusion technique were compared for 915 samples of first milking colostrum from Holstein cows. Least squares analysis of the relationship between hydrometer measurement and IgG1 concentration was improved by log transformation of IgG1 concentration and resulted in a significant linear relationship between hydrometer measurement and log10 IgG1 concentration; r2 = .469. At 50 mg of globulin/ml of colostrum, the recommended hydrometer cutoff point for colostrum selection, the sensitivity of the hydrometer as a test of IgG1 concentration in Holstein colostrum was 26%, and the negative predictive value was 67%. The negative predictive value and sensitivity of the hydrometer as a test of IgG1 in Holstein colostrum was improved, and the cost of misclassification of colostrum was minimized, when the cutoff point for colostrum selection was increased above the recommended 50 mg/ml.

  18. Guiding bispecific monovalent antibody formation through proteolysis of IgG1 single-chain.

    PubMed

    Dimasi, Nazzareno; Fleming, Ryan; Sachsenmeier, Kris F; Bezabeh, Binyam; Hay, Carl; Wu, Jincheng; Sult, Erin; Rajan, Saravanan; Zhuang, Li; Cariuk, Peter; Buchanan, Andrew; Bowen, Michael A; Wu, Herren; Gao, Changshou

    2017-04-01

    We developed an IgG1 domain-tethering approach to guide the correct assembly of 2 light and 2 heavy chains, derived from 2 different antibodies, to form bispecific monovalent antibodies in IgG1 format. We show here that assembling 2 different light and heavy chains by sequentially connecting them with protease-cleavable polypeptide linkers results in the generation of monovalent bispecific antibodies that have IgG1 sequence, structure and functional properties. This approach was used to generate a bispecific monovalent antibody targeting the epidermal growth factor receptor and the type I insulin-like growth factor receptor that: 1) can be produced and purified using standard IgG1 techniques; 2) exhibits stability and structural features comparable to IgG1; 3) binds both targets simultaneously; and 4) has potent anti-tumor activity. Our strategy provides new engineering opportunities for bispecific antibody applications, and, most importantly, overcomes some of the limitations (e.g., half-antibody and homodimer formation, light chains mispairing, multi-step purification), inherent with some of the previously described IgG1-based bispecific monovalent antibodies.

  19. GPER agonist G-1 decreases adrenocortical carcinoma (ACC) cell growth in vitro and in vivo

    PubMed Central

    Zolea, Fabiana; Rizza, Pietro; Avena, Paola; Malivindi, Rocco; De Luca, Arianna; Campana, Carmela; Martire, Emilia; Domanico, Francesco; Fallo, Francesco; Carpinelli, Giulia; Cerquetti, Lidia; Amendola, Donatella; Stigliano, Antonio; Pezzi, Vincenzo

    2015-01-01

    We have previously demonstrated that estrogen receptor (ER) alpha (ESR1) increases proliferation of adrenocortical carcinoma (ACC) through both an estrogen-dependent and -independent (induced by IGF-II/IGF1R pathways) manner. Then, the use of tamoxifen, a selective estrogen receptor modulator (SERM), appears effective in reducing ACC growth in vitro and in vivo. However, tamoxifen not only exerts antiestrogenic activity, but also acts as full agonist on the G protein-coupled estrogen receptor (GPER). Aim of this study was to investigate the effect of a non-steroidal GPER agonist G-1 in modulating ACC cell growth. We found that G-1 is able to exert a growth inhibitory effect on H295R cells both in vitro and, as xenograft model, in vivo. Treatment of H295R cells with G-1 induced cell cycle arrest, DNA damage and cell death by the activation of the intrinsic apoptotic mechanism. These events required sustained extracellular regulated kinase (ERK) 1/2 activation. Silencing of GPER by a specific shRNA partially reversed G-1-mediated cell growth inhibition without affecting ERK activation. These data suggest the existence of G-1 activated but GPER-independent effects that remain to be clarified. In conclusion, this study provides a rational to further study G-1 mechanism of action in order to include this drug as a treatment option to the limited therapy of ACC. PMID:26131713

  20. IgG1 Fc N-glycan galactosylation as a biomarker for immune activation

    PubMed Central

    de Jong, Sanne E.; Selman, Maurice H. J.; Adegnika, Ayola A.; Amoah, Abena S.; van Riet, Elly; Kruize, Yvonne C. M.; Raynes, John G.; Rodriguez, Alejandro; Boakye, Daniel; von Mutius, Erika; Knulst, André C.; Genuneit, Jon; Cooper, Philip J.; Hokke, Cornelis H.; Wuhrer, Manfred; Yazdanbakhsh, Maria

    2016-01-01

    Immunoglobulin G (IgG) Fc N-glycosylation affects antibody-mediated effector functions and varies with inflammation rooted in both communicable and non-communicable diseases. Worldwide, communicable and non-communicable diseases tend to segregate geographically. Therefore, we studied whether IgG Fc N-glycosylation varies in populations with different environmental exposures in different parts of the world. IgG Fc N-glycosylation was analysed in serum/plasma of 700 school-age children from different communities of Gabon, Ghana, Ecuador, the Netherlands and Germany. IgG1 galactosylation levels were generally higher in more affluent countries and in more urban communities. High IgG1 galactosylation levels correlated with low total IgE levels, low C-reactive protein levels and low prevalence of parasitic infections. Linear mixed modelling showed that only positivity for parasitic infections was a significant predictor of reduced IgG1 galactosylation levels. That IgG1 galactosylation is a predictor of immune activation is supported by the observation that asthmatic children seemed to have reduced IgG1 galactosylation levels as well. This indicates that IgG1 galactosylation levels could be used as a biomarker for immune activation of populations, providing a valuable tool for studies examining the epidemiological transition from communicable to non-communicable diseases. PMID:27306703

  1. Increased expression of histone deacetylases 2 in temporal lobe epilepsy: a study of epileptic patients and rat models.

    PubMed

    Huang, Yuanyuan; Zhao, Fenghua; Wang, Liang; Yin, Huan; Zhou, Chunlei; Wang, Xuefeng

    2012-02-01

    Histone deacetylases 2 (HDAC2) is expressed in the central nervous system; it has multiple functions in neural plasticity. However, we do not know if HDAC2 is also involved in the pathology of epilepsy. Here we report that HDAC2 was expressed in the brain tissues of both control and temporal lobe epilepsy (TLE) patients. Results from immunofluorescence and immunohistochemistry showed that HDAC2 was primarily located in the nucleus and that TLE patients exhibit significantly more HDAC2 positive cells than control. Western blotting showed that HDAC2 protein levels were significantly higher in TLE than in control brain. Moreover, in the rat model of TLE, there was a sustained enhancement of HDAC2 expression in rat models of TLE. HDAC2 was significantly increased in both the acute (1 day) and chronic (60 days) animals compared with control group. These results suggest that HDAC2 play an important role in the pathogenesis of human TLE.

  2. Histone deacetylase inhibitor induces cell apoptosis and cycle arrest in lung cancer cells via mitochondrial injury and p53 up-acetylation.

    PubMed

    Bao, Lianmin; Diao, Hua; Dong, Nian; Su, Xiaoqiong; Wang, Bingbin; Mo, Qiongya; Yu, Heguo; Wang, Xiangdong; Chen, Chengshui

    2016-12-01

    The reversibility of non-genotoxic phenotypic changes has been explored in order to develop novel preventive and therapeutic approaches for cancer. Quisinostat (JNJ-26481585), a novel second-generation histone deacetylase inhibitor (HDACi), has efficient therapeutic actions on non-small cell lung cancer (NSCLC) cell. The present study aims at investigating underlying molecular mechanisms involved in the therapeutic activity of quisinostat on NSCLC cells. We found that quisinostat significantly inhibited A549 cell proliferation in dose- and time-dependent manners. Up-acetylation of histones H3 and H4 and non-histone protein α-tubulin was induced by quisinostat treatment in a nanomolar concentration. We also demonstrated that quisinostat increased reactive oxygen species (ROS) production and destroyed mitochondrial membrane potential (ΔΨm), inducing mitochondria-mediated cell apoptosis. Furthermore, exposure of A549 cells to quisinostat significantly suppressed cell migration by inhibiting epithelial-mesenchymal transition (EMT) process. Bioinformatics analysis indicated that effects of quisinostat on NSCLC cells were associated with activated p53 signaling pathway. We found that quisinostat increased p53 acetylation at K382/K373 sites, upregulated the expression of p21((Waf1/Cip1)), and resulted in G1 phase arrest. Thus, our results suggest that the histone deacetylase can be a therapeutic target of NSCLC to discover and develop a new category of therapy for lung cancer.

  3. Cystathionine γ-lyase protects vascular endothelium: a role for inhibition of histone deacetylase 6.

    PubMed

    Leucker, Thorsten M; Nomura, Yohei; Kim, Jae Hyung; Bhatta, Anil; Wang, Victor; Wecker, Andrea; Jandu, Sandeep; Santhanam, Lakshmi; Berkowitz, Dan; Romer, Lewis; Pandey, Deepesh

    2017-04-01

    Endothelial cystathionine γ-lyase (CSEγ) contributes to cardiovascular homeostasis, mainly through production of H2S. However, the molecular mechanisms that control CSEγ gene expression in the endothelium during cardiovascular diseases are unclear. The aim of the current study is to determine the role of specific histone deacetylases (HDACs) in the regulation of endothelial CSEγ. Reduced CSEγ mRNA expression and protein abundance were observed in human aortic endothelial cells (HAEC) exposed to oxidized LDL (OxLDL) and in aortas from atherogenic apolipoprotein E knockout (ApoE(-/-)) mice fed a high-fat diet compared with controls. Intact murine aortic rings exposed to OxLDL (50 μg/ml) for 24 h exhibited impaired endothelium-dependent vasorelaxation that was blocked by CSEγ overexpression or the H2S donor NaHS. CSEγ expression was upregulated by pan-HDAC inhibitors and by class II-specific HDAC inhibitors, but not by other class-specific inhibitors. The HDAC6 selective inhibitor tubacin and HDAC6-specific siRNA increased CSEγ expression and blocked OxLDL-mediated reductions in endothelial CSEγ expression and CSEγ promoter activity, indicating that HDAC6 is a specific regulator of CSEγ expression. Consistent with this finding, HDAC6 mRNA, protein expression, and activity were upregulated in OxLDL-exposed HAEC, but not in human aortic smooth muscle cells. HDAC6 protein levels in aortas from high-fat diet-fed ApoE(-/-) mice were comparable to those in controls, whereas HDAC6 activity was robustly upregulated. Together, our findings indicate that HDAC6 is upregulated by atherogenic stimuli via posttranslational modifications and is a critical regulator of CSEγ expression in vascular endothelium. Inhibition of HDAC6 activity may improve endothelial function and prevent or reverse the development of atherosclerosis.NEW & NOTEWORTHY Oxidative injury to endothelial cells by oxidized LDL reduced cystathionine γ-lyase (CSEγ) expression and H2S production

  4. Dominant recognition of a cross-reactive B-cell epitope in Mycobacterium leprae 10 K antigen by immunoglobulin G1 antibodies across the disease spectrum in leprosy

    PubMed Central

    Hussain, R; Dockrell, H M; Chiang, T J

    1999-01-01

    Mycobacterium leprae-specific immunoglobulin G1 (IgG1) antibodies in patients with leprosy show a direct correlation with bacterial load (ρ=0·748; P < 0002) suggesting that IgG1 B-cell responses may be surrogate markers of disease progression. To investigate if this upregulation was a general feature of IgG1 responses to all M. leprae (ML) antigens, we analysed responses to several recombinant purified ML heat-shock proteins (HSP). Three recombinant HSPs (ML10 K, ML 18 K and ML 65 K) were tested for their ability to induce various IgG subclasses in patients with either the lepromatous (LL/BL, n = 26) or tuberculoid form (BT/TT, n = 39) of the disease as well as in healthy households (HC, n = 14) and endemic controls (EC = 19). Our major findings were: (1) selective augmentation of IgG1 antibody responses to ML10 K; (2) recognition of a restricted number of epitopes across the disease spectrum and healthy controls by IgG1 antibodies; (3) dominant recognition of cross-reactive epitopes which were common to both ML and MT 10 K. This response was not related to contamination with endotoxin. Epitope mapping using 15-mer overlapping peptides spanning the ML 10 000 MW revealed an immunodominant IgG1 binding peptide (aa41–55) in patients as well as healthy controls. This peptide is a shared epitope with M. tuberculosis 10 K suggesting that postswitched IgG1 B cells recognizing this epitope rather than naive B cells are being expanded. PMID:10233750

  5. IgG1 deficiency exacerbates experimental autoimmune myasthenia gravis in BALB/c mice.

    PubMed

    Huda, Ruksana; Strait, Richard T; Tüzün, Erdem; Finkelman, Fred D; Christadoss, Premkumar

    2015-04-15

    Myasthenia gravis is an autoimmune disease characterized by muscle weakness due to neuromuscular junction (NMJ) damage by anti-acetylcholine receptor (AChR) auto-antibodies and complement. In experimental autoimmune myasthenia gravis (EAMG), which is induced by immunization with Torpedo AChR in CFA, anti-AChR IgG2b and IgG1 are the predominant isotypes in the circulation. Complement activation by isotypes such as IgG2b plays a crucial role in EAMG pathogenesis; this suggested the possibility that IgG1, which does not activate complement through the classical pathway, may suppress EAMG. In this study, we show that AChR-immunized BALB/c mice genetically deficient for IgG1 produce higher levels of complement-activating isotypes of anti-AChR, especially IgG3 and IgG2a, and develop increased IgG3/IgG2a deposits at the NMJ, as compared to wild type (WT) BALB/c mice. Consistent with this, AChR-immunized IgG1(-/-) BALB/c mice lose muscle strength and muscle AChR to a greater extent than AChR-immunized WT mice. These observations demonstrate that IgG1 deficiency leads to increased severity of EAMG associated with an increase in complement activating IgG isotypes. Further studies are needed to dissect the specific role or mechanism of IgG1 in limiting EAMG and that of EAMG exacerbating role of complement activating IgG3 and IgG2a in IgG1 deficiency.

  6. IL-27 induces the production of IgG1 by human B cells.

    PubMed

    Boumendjel, Amel; Tawk, Lina; Malefijt, René de Waal; Boulay, Vera; Yssel, Hans; Pène, Jérôme

    2006-12-01

    It has been reported that IL-27 specifically induces the production of IgG2a by mouse B cells and inhibits IL-4-induced IgG1 synthesis. Here, we show that human naïve cord blood expresses a functional IL-27 receptor, consisting of the TCCR and gp130 subunits, although at lower levels as compared to naïve and memory splenic B cells. IL-27 does not induce proliferative responses and does not increase IgG1 production by CD19(+)CD27(+) memory B cells. However, it induces a low, but significant production of IgG1 by naïve CD19(+)CD27(-)IgD(+)IgG(-) spleen and cord blood B cells, activated via CD40, whereas it has no effect on the production of the other IgG subclasses. In addition, IL-27 induces the differentiation of a population of B cells that express high levels of CD38, in association with a down-regulation of surface IgD expression, and that are surface IgG(+/int), CD20(low), CD27(high), indicating that IL-27 promotes isotype switching and plasma cell differentiation of naive B cells. However, as compared to the effects of IL-21 and IL-10, both switch factors for human IgG1 and IgG3, those of IL-27 are modest and regulate exclusively the production of IgG1. Finally, although IL-27 has no effect on IL-4 and anti-CD40-induced Cepsilon germline promoter activity, it up-regulates IL-4-induced IgE production by naive B cells. These results point to a partial redundancy of switch factors regulating the production of IgG1 in humans, and furthermore indicate the existence of a common regulation of the human IgG1and murine IgG2a isotypes by IL-27.

  7. Detection of G1 proteins in Chinese hamster cells synchronized by isoleucine deprivation or mitotic selection.

    PubMed

    Ley, K D

    1975-07-01

    Examination of labeling patterns of proteins in Chinese hamster cells(line CHO) revealed the presence of a class of protein(s) that is synthesized during G1 phase of the cell cycle. Cells arrested in G1 by isoleucine (Ile) deprivation were prelabeded with [14-C]Ile, induced to traverse G1 by addition of unlabeled Ile, and labeled with [3-H]Ile at hourly intervals. Cells were fractionated into neclear and cytoplasmic portions, and proteins were separated by sodium dodecyl sulfate-polyacrylamide get electrophoresis. Gel profiles of proteins in the 45,000-160,000 mol wt range from the cytoplasm of cells in G1 were similar to those from cells arrested in G1 except for the presence of a mojor peak of [1-H]Ile incorporated into a protein(s) of approximately 80,000 mol wt. Peaks of net [3-H]Ile incorporation were not detected in neclear preparations. Cellular fractionation by differential centrifugation showed the peak I protein was located in the soluble supernatant fraction of the cytoplasm. Time-course studies showed that synthesis of this protein began 1-2 h after initiation of G1 traverse; the protein reached maximum levels in 4-6 h and was reduced to undetectable levels by 9 h. A cytoplasmic protein with similar electrophoretic mobility was found in G1 phase of cells synchronized by mitotic selection. This class of proteins is synthesized by cells before entry into S phase and may be involved in initiation of DNA synthesis.

  8. Nuclear levels and patterns of histone H3 modification and HP1 proteins after inhibition of histone deacetylases.

    PubMed

    Bártová, Eva; Pacherník, Jirí; Harnicarová, Andrea; Kovarík, Ales; Kovaríková, Martina; Hofmanová, Jirina; Skalníková, Magdalena; Kozubek, Michal; Kozubek, Stanislav

    2005-11-01

    The effects of the histone deacetylase inhibitors (HDACi) trichostatin A (TSA) and sodium butyrate (NaBt) were studied in A549, HT29 and FHC human cell lines. Global histone hyperacetylation, leading to decondensation of interphase chromatin, was characterized by an increase in H3(K9) and H3(K4) dimethylation and H3(K9) acetylation. The levels of all isoforms of heterochromatin protein, HP1, were reduced after HDAC inhibition. The observed changes in the protein levels were accompanied by changes in their interphase patterns. In control cells, H3(K9) acetylation and H3(K4) dimethylation were substantially reduced to a thin layer at the nuclear periphery, whereas TSA and NaBt caused the peripheral regions to become intensely acetylated at H3(K9) and dimethylated at H3(K4). The dispersed pattern of H3(K9) dimethylation was stable even at the nuclear periphery of HDACi-treated cells. After TSA and NaBt treatment, the HP1 proteins were repositioned more internally in the nucleus, being closely associated with interchromatin compartments, while centromeric heterochromatin was relocated closer to the nuclear periphery. These findings strongly suggest dissociation of HP1 proteins from peripherally located centromeres in a hyperacetylated and H3(K4) dimethylated environment. We conclude that inhibition of histone deacetylases caused dynamic reorganization of chromatin in parallel with changes in its epigenetic modifications.

  9. Impact of the GeoMIP G1 sunshade geoengineering experiment on the Atlantic meridional overturning circulation

    NASA Astrophysics Data System (ADS)

    Hong, Yu; Moore, John C.; Jevrejeva, Svetlana; Ji, Duoying; Phipps, Steven J.; Lenton, Andrew; Tilmes, Simone; Watanabe, Shingo; Zhao, Liyun

    2017-03-01

    We analyze the multi-earth system model responses of ocean temperatures and the Atlantic Meridional Overturning Circulation (AMOC) under an idealized solar radiation management scenario (G1) from the Geoengineering Model Intercomparison Project. All models simulate warming of the northern North Atlantic relative to no geoengineering, despite geoengineering substantially offsetting the increases in mean global ocean temperatures. Increases in the temperature of the North Atlantic Ocean at the surface (˜0.25 K) and at a depth of 500 m (˜0.10 K) are mainly due to a 10 Wm-2 reduction of total heat flux from ocean to atmosphere. Although the AMOC is slightly reduced under the solar dimming scenario, G1, relative to piControl, it is about 37% stronger than under abrupt4 × CO2 . The reduction of the AMOC under G1 is mainly a response to the heat flux change at the northern North Atlantic rather than to changes in the water flux and the wind stress. The AMOC transfers heat from tropics to high latitudes, helping to warm the high latitudes, and its strength is maintained under solar dimming rather than weakened by greenhouse gas forcing acting alone. Hence the relative reduction in high latitude ocean temperatures provided by solar radiation geoengineering, would tend to be counteracted by the correspondingly active AMOC circulation which furthermore transports warm surface waters towards the Greenland ice sheet, warming Arctic sea ice and permafrost.

  10. Effect of high mobility group nonhistone proteins HMG-20 (ubiquitin) and HMG-17 on histone deacetylase activity assayed in vitro.

    PubMed Central

    Mezquita, J; Chiva, M; Vidal, S; Mezquita, C

    1982-01-01

    We have used a method previously described by Reeves and Candido (1) to partially release histone deacetylase from cell nuclei together with putative regulators of the enzyme. Histone deacetylase released from testis cell nuclei and its putative regulators were separated by gel filtration in Sepharose 6B. A peak of low molecular weight contains a heat-stable factor that stimulate histone deacetylase in vitro. Many of the properties of the activator coincide with those of the protein HMG-20 (ubiquitin). Ubiquitin isolated from testis cell nuclei stimulated histone deacetylase in vitro. It has been suggested that HMG-17 partially inhibits histone deacetylase in Fried cell nuclei (2). In our system, HMG-17 shows no inhibitory effect on histone deacetylase activity Images PMID:6280157

  11. Echinococcus granulosus genotypes in livestock of Iran indicating high frequency of G1 genotype in camels.

    PubMed

    Sharbatkhori, Mitra; Mirhendi, Hossein; Harandi, Majid Fasihi; Rezaeian, Mostafa; Mohebali, Mehdi; Eshraghian, Mohammadreza; Rahimi, Hamidreza; Kia, Eshrat Beigom

    2010-04-01

    In this study, 112 Echinococcus granulosus isolates from different livestock of Iran were genotyped by PCR amplification of ribosomal DNA-internal transcribed spacer 1 (rDNA-ITS1) region followed by restriction fragment length polymorphism (RFLP) with the enzyme RsaI. The possibility of intra-genotype variation was also investigated using randomly amplified polymorphic DNA (RAPD) analysis. Isolates from sheep, goats, cattle and the majority of camels (12 of 18; 66.7%) were identified as the G1 genotype and a few camel isolates (6 of 18; 33.3%) belonged to the G6 genotype. Overall G1 and G6 genotypes were identified in 94.6% (106 of 112) and 5.3% (6 of 112) of all isolates, respectively. RAPD analysis based on 15 separate primers showed 7-14 bands of 200-3000bp for strain G1. Considering each individual primer, no differences observed among isolates from different hosts and between livers and lungs. This study confirmed the existence of G1 and G6 genotypes in Iran. Moreover, G1 is much more prevalent even in camels, indicating the importance of sheep-dog cycle in public health. Studying intra-genotypic variation of E. granulosus warrants more research using other primers and methods.

  12. Selection of G1 Phase Yeast Cells for Synchronous Meiosis and Sporulation.

    PubMed

    Stuart, David T

    2017-01-01

    Centrifugal elutriation is a procedure that allows the fractionation of cell populations based upon their size and shape. This allows cells in distinct cell cycle stages can be captured from an asynchronous population. The technique is particularly helpful when performing an experiment to monitor the progression of cells through the cell cycle or meiosis. Yeast sporulation like gametogenesis in other eukaryotes initiates from the G1 phase of the cell cycle. Conveniently, S. cerevisiae arrest in G1 phase when starved for nutrients and so withdrawal of nitrogen and glucose allows cells to abandon vegetative growth in G1 phase before initiating the sporulation program. This simple starvation protocol yields a partial synchronization that has been used extensively in studies of progression through meiosis and sporulation. By using centrifugal elutriation it is possible to isolate a homogeneous population of G1 phase cells and induce them to sporulate synchronously, which is beneficial for investigating progression through meiosis and sporulation. An additionally benefit of this protocol is that cell populations can be isolated based upon size and both large and small cell populations can be tested for progression through meiosis and sporulation. Here we present a protocol for purification of G1 phase diploid cells for examining synchronous progression through meiosis and sporulation.

  13. A Dynamical Framework for the All-or-None G1/S Transition

    PubMed Central

    Barr, Alexis R.; Heldt, Frank S.; Zhang, Tongli; Bakal, Chris; Novák, Béla

    2016-01-01

    Summary The transition from G1 into DNA replication (S phase) is an emergent behavior resulting from dynamic and complex interactions between cyclin-dependent kinases (Cdks), Cdk inhibitors (CKIs), and the anaphase-promoting complex/cyclosome (APC/C). Understanding the cellular decision to commit to S phase requires a quantitative description of these interactions. We apply quantitative imaging of single human cells to track the expression of G1/S regulators and use these data to parametrize a stochastic mathematical model of the G1/S transition. We show that a rapid, proteolytic, double-negative feedback loop between Cdk2:Cyclin and the Cdk inhibitor p27Kip1 drives a switch-like entry into S phase. Furthermore, our model predicts that increasing Emi1 levels throughout S phase are critical in maintaining irreversibility of the G1/S transition, which we validate using Emi1 knockdown and live imaging of G1/S reporters. This work provides insight into the general design principles of the signaling networks governing the temporally abrupt transitions between cell-cycle phases. PMID:27136687

  14. A comparative genomic analysis of the alkalitolerant soil bacterium Bacillus lehensis G1.

    PubMed

    Noor, Yusuf Muhammad; Samsulrizal, Nurul Hidayah; Jema'on, Noor Azah; Low, Kheng Oon; Ramli, Aizi Nor Mazila; Alias, Noor Izawati; Damis, Siti Intan Rosdianah; Fuzi, Siti Fatimah Zaharah Mohd; Isa, Mohd Noor Mat; Murad, Abdul Munir Abdul; Raih, Mohd Firdaus Mohd; Bakar, Farah Diba Abu; Najimudin, Nazalan; Mahadi, Nor Muhammad; Illias, Rosli Md

    2014-07-25

    Bacillus lehensis G1 is a Gram-positive, moderately alkalitolerant bacterium isolated from soil samples. B. lehensis produces cyclodextrin glucanotransferase (CGTase), an enzyme that has enabled the extensive use of cyclodextrin in foodstuffs, chemicals, and pharmaceuticals. The genome sequence of B. lehensis G1 consists of a single circular 3.99 Mb chromosome containing 4017 protein-coding sequences (CDSs), of which 2818 (70.15%) have assigned biological roles, 936 (23.30%) have conserved domains with unknown functions, and 263 (6.55%) have no match with any protein database. Bacillus clausii KSM-K16 was established as the closest relative to B. lehensis G1 based on gene content similarity and 16S rRNA phylogenetic analysis. A total of 2820 proteins from B. lehensis G1 were found to have orthologues in B. clausii, including sodium-proton antiporters, transport proteins, and proteins involved in ATP synthesis. A comparative analysis of these proteins and those in B. clausii and other alkaliphilic Bacillus species was carried out to investigate their contributions towards the alkalitolerance of the microorganism. The similarities and differences in alkalitolerance-related genes among alkalitolerant/alkaliphilic Bacillus species highlight the complex mechanism of pH homeostasis. The B. lehensis G1 genome was also mined for proteins and enzymes with potential viability for industrial and commercial purposes.

  15. RESEARCH PAPER: Old stellar population synthesis: new age and mass estimates for Mayall II = G1

    NASA Astrophysics Data System (ADS)

    Ma, Jun; de Grijs, Richard; Fan, Zhou; Rey, Soo-Chang; Wu, Zhen-Yu; Zhou, Xu; Wu, Jiang-Hua; Jiang, Zhao-Ji; Chen, Jian-Sheng; Lee, Kyungsook; Sohn, Sangmo Tony

    2009-06-01

    Mayall II = G1 is one of the most luminous globular clusters (GCs) in M31. Here, we determine its age and mass by comparing multicolor photometry with theoretical stellar population synthesis models. Based on far- and near-ultraviolet GALEX photometry, broad-band UBVRI, and infrared JHKS 2MASS data, we construct the most extensive spectral energy distribution of G1 to date, spanning the wavelength range from 1538 to 20 000 Å. A quantitative comparison with a variety of simple stellar population (SSP) models yields a mean age which is consistent with G1 being among the oldest building blocks of M31 and having formed within ~1.7 Gyr after the Big Bang. Irrespective of the SSP model or stellar initial mass function adopted, the resulting mass estimates (of order 107 Modot) indicate that G1 is one of the most massive GCs in the Local Group. However, we speculate that the cluster's exceptionally high mass suggests that it may not be a genuine GC. Our results also suggest that G1 may contain, on average, (1.65±0.63) × 102 Lodot far-ultraviolet-bright, hot, extreme horizontal-branch stars, depending on the adopted SSP model. In addition, we demonstrate that extensive multi-passband photometry coupled with SSP analysis enables one to obtain age estimates for old SSPs that have similar accuracies as those from integrated spectroscopy or resolved stellar photometry, provided that some of the free parameters can be constrained independently.

  16. The role of RBF in the introduction of G1 regulation during Drosophila embryogenesis.

    PubMed Central

    Du, W; Dyson, N

    1999-01-01

    The first appearance of G1 during Drosophila embryogenesis, at cell cycle 17, is accompanied by the down-regulation of E2F-dependent transcription. Mutant alleles of rbf were generated and analyzed to determine the role of RBF in this process. Embryos lacking both maternal and zygotic RBF products show constitutive expression of PCNA and RNR2, two E2F-regulated genes, indicating that RBF is required for their transcriptional repression. Despite the ubiquitous expression of E2F target genes, most epidermal cells enter G1 normally. Rather than pausing in G1 until the appropriate time for cell cycle progression, many of these cells enter an ectopic S-phase. These results indicate that the repression of E2F target genes by RBF is necessary for the maintenance but not the initiation of a G1 phase. The phenotype of RBF-deficient embryos suggests that rbf has a function that is complementary to the roles of dacapo and fizzy-related in the introduction of G1 during Drosophila embryogenesis. PMID:10022834

  17. A Plasmodium falciparum histone deacetylase regulates antigenic variation and gametocyte conversion.

    PubMed

    Coleman, Bradley I; Skillman, Kristen M; Jiang, Rays H Y; Childs, Lauren M; Altenhofen, Lindsey M; Ganter, Markus; Leung, Yvette; Goldowitz, Ilana; Kafsack, Björn F C; Marti, Matthias; Llinás, Manuel; Buckee, Caroline O; Duraisingh, Manoj T

    2014-08-13

    The asexual forms of the malaria parasite Plasmodium falciparum are adapted for chronic persistence in human red blood cells, continuously evading host immunity using epigenetically regulated antigenic variation of virulence-associated genes. Parasite survival on a population level also requires differentiation into sexual forms, an obligatory step for further human transmission. We reveal that the essential nuclear gene, P. falciparum histone deacetylase 2 (PfHda2), is a global silencer of virulence gene expression and controls the frequency of switching from the asexual cycle to sexual development. PfHda2 depletion leads to dysregulated expression of both virulence-associated var genes and PfAP2-g, a transcription factor controlling sexual conversion, and is accompanied by increases in gametocytogenesis. Mathematical modeling further indicates that PfHda2 has likely evolved to optimize the parasite's infectious period by achieving low frequencies of virulence gene expression switching and sexual conversion. This common regulation of cellular transcriptional programs mechanistically links parasite transmissibility and virulence.

  18. Histone deacetylases 1 and 2 regulate autophagy flux and skeletal muscle homeostasis in mice

    PubMed Central

    Moresi, Viviana; Carrer, Michele; Grueter, Chad E.; Rifki, Oktay F.; Shelton, John M.; Richardson, James A.; Bassel-Duby, Rhonda; Olson, Eric N.

    2012-01-01

    Maintenance of skeletal muscle structure and function requires efficient and precise metabolic control. Autophagy plays a key role in metabolic homeostasis of diverse tissues by recycling cellular constituents, particularly under conditions of caloric restriction, thereby normalizing cellular metabolism. Here we show that histone deacetylases (HDACs) 1 and 2 control skeletal muscle homeostasis and autophagy flux in mice. Skeletal muscle-specific deletion of both HDAC1 and HDAC2 results in perinatal lethality of a subset of mice, accompanied by mitochondrial abnormalities and sarcomere degeneration. Mutant mice that survive the first day of life develop a progressive myopathy characterized by muscle degeneration and regeneration, and abnormal metabolism resulting from a blockade to autophagy. HDAC1 and HDAC2 regulate skeletal muscle autophagy by mediating the induction of autophagic gene expression and the formation of autophagosomes, such that myofibers of mice lacking these HDACs accumulate toxic autophagic intermediates. Strikingly, feeding HDAC1/2 mutant mice a high-fat diet from the weaning age releases the block in autophagy and prevents myopathy in adult mice. These findings reveal an unprecedented and essential role for HDAC1 and HDAC2 in maintenance of skeletal muscle structure and function and show that, at least in some pathological conditions, myopathy may be mitigated by dietary modifications. PMID:22307625

  19. Heterochromatin assembly and transcriptome repression by Set1 in coordination with a class II histone deacetylase.

    PubMed

    Lorenz, David R; Meyer, Lauren F; Grady, Patrick J R; Meyer, Michelle M; Cam, Hugh P

    2014-12-15

    Histone modifiers play essential roles in controlling transcription and organizing eukaryotic genomes into functional domains. Here, we show that Set1, the catalytic subunit of the highly conserved Set1C/COMPASS complex responsible for histone H3K4 methylation (H3K4me), behaves as a repressor of the transcriptome largely independent of Set1C and H3K4me in the fission yeast Schizosaccharomyces pombe. Intriguingly, while Set1 is enriched at highly expressed and repressed loci, Set1 binding levels do not generally correlate with the levels of transcription. We show that Set1 is recruited by the ATF/CREB homolog Atf1 to heterochromatic loci and promoters of stress-response genes. Moreover, we demonstrate that Set1 coordinates with the class II histone deacetylase Clr3 in heterochromatin assembly at prominent chromosomal landmarks and repression of the transcriptome that includes Tf2 retrotransposons, noncoding RNAs, and regulators of development and stress-responses. Our study delineates a molecular framework for elucidating the functional links between transcriptome control and chromatin organization.

  20. Design, synthesis, and antitumor evaluation of histone deacetylase inhibitors with l-phenylglycine scaffold

    PubMed Central

    Zhang, Yingjie; Li, Xiaoguang; Hou, Jinning; Huang, Yongxue; Xu, Wenfang

    2015-01-01

    In our previous research, a novel series of histone deacetylase (HDAC) inhibitors with l-phenylglycine scaffold were designed and synthesized, among which amides D3 and D7 and ureido D18 were far superior to the positive control (suberoylanilide hydroxamic acid [SAHA]) in HDAC inhibition, but were only comparable to SAHA in antiproliferation on tumor cell lines. Herein, further structural derivation of lead compounds D3, D7, and D18 was carried out to improve their cellular activities. Most of our newly synthesized compounds exhibited more potent HDAC inhibitory activities than the positive control SAHA, and several derivatives were even better than their parent compounds. However, compared with SAHA and our lead compounds, only secondary amine series compounds exhibited improved antiproliferative activities, likely due to their appropriate topological polar surface area values and cell permeabilities. In a human histiocytic lymphoma (U937) xenograft model, the most potent secondary amine 9d exhibited similar in vivo antitumor activity to that of SAHA. PMID:26504374

  1. Polycomb- and REST-associated histone deacetylases are independent pathways toward a mature neuronal phenotype

    PubMed Central

    McGann, James C; Oyer, Jon A; Garg, Saurabh; Yao, Huilan; Liu, Jun; Feng, Xin; Liao, Lujian; Yates, John R; Mandel, Gail

    2014-01-01

    The bivalent hypothesis posits that genes encoding developmental regulators required for early lineage decisions are poised in stem/progenitor cells by the balance between a repressor histone modification (H3K27me3), mediated by the Polycomb Repressor Complex 2 (PRC2), and an activator modification (H3K4me3). In this study, we test whether this mechanism applies equally to genes that are not required until terminal differentiation. We focus on the RE1 Silencing Transcription Factor (REST) because it is expressed highly in stem cells and is an established global repressor of terminal neuronal genes. Elucidation of the REST complex, and comparison of chromatin marks and gene expression levels in control and REST-deficient stem cells, shows that REST target genes are poised by a mechanism independent of Polycomb, even at promoters which bear the H3K27me3 mark. Specifically, genes under REST control are actively repressed in stem cells by a balance of the H3K4me3 mark and a repressor complex that relies on histone deacetylase activity. Thus, chromatin distinctions between pro-neural and terminal neuronal genes are established at the embryonic stem cell stage by two parallel, but distinct, repressor pathways. DOI: http://dx.doi.org/10.7554/eLife.04235.001 PMID:25250711

  2. The histone deacetylase inhibitor, romidepsin, as a potential treatment for pulmonary fibrosis

    PubMed Central

    Calderwood, Claire J.; Thatcher, Thomas H.; Jones, Mark G.; Smart, David E.; Mahajan, Sumeet; Alzetani, Aiman; Havelock, Tom; Maher, Toby M.; Molyneaux, Philip L.; Thorley, Andrew J.; Tetley, Teresa D.; Warner, Jane A.; Packham, Graham; Ganesan, A.; Skipp, Paul J.; Marshall, Benjamin J.; Richeldi, Luca; Sime, Patricia J.; O'Reilly, Katherine M.A.; Davies, Donna E.

    2017-01-01

    Idiopathic pulmonary fibrosis (IPF) is a progressive disease that usually affects elderly people. It has a poor prognosis and there are limited therapies. Since epigenetic alterations are associated with IPF, histone deacetylase (HDAC) inhibitors offer a novel therapeutic strategy to address the unmet medical need. This study investigated the potential of romidepsin, an FDA-approved HDAC inhibitor, as an anti-fibrotic treatment and evaluated biomarkers of target engagement that may have utility in future clinical trials. The anti-fibrotic effects of romidepsin were evaluated both in vitro and in vivo together with any harmful effect on alveolar type II cells (ATII). Bronchoalveolar lavage fluid (BALF) from IPF or control donors was analyzed for the presence of lysyl oxidase (LOX). In parallel with an increase in histone acetylation, romidepsin potently inhibited fibroblast proliferation, myofibroblast differentiation and LOX expression. ATII cell numbers and their lamellar bodies were unaffected. In vivo, romidepsin inhibited bleomycin-induced pulmonary fibrosis in association with suppression of LOX expression. LOX was significantly elevated in BALF of IPF patients compared to controls. These data show the anti-fibrotic effects of romidepsin, supporting its potential use as novel treatment for IPF with LOX as a companion biomarker for evaluation of early on-target effects. PMID:28467787

  3. Histone deacetylases: revealing the molecular base of dimorphism in pathogenic fungi

    PubMed Central

    Elías-Villalobos, Alberto; Helmlinger, Dominique; Ibeas, José I.

    2015-01-01

    Fungi, as every living organism, interact with the external world and have to adapt to its fluctuations. For pathogenic fungi, such interaction involves adapting to the hostile environment of their host. Survival depends on the capacity of fungi to detect and respond to external stimuli, which is achieved through a tight and efficient genetic control. Chromatin modifications represent a well-known layer of regulation that controls gene expression in response to environmental signals. However, less is known about the chromatin modifications that are involved in fungal virulence and the specific cues and signalling pathways that target chromatin modifications to specific genes. In a recently published study, our research group identified one such regulatory pathway. We demonstrated that the histone deacetylase (HDAC) Hos2 is involved in yeast-to-hyphal transition (dimorphism) and it is associated with the virulence of the maize pathogen Ustilago maydis, the causative agent of smut disease in corn. Hos2 activates mating-type genes by directly binding to their gene bodies. Furthermore, Hos2 acts downstream of the nutrient-sensing cyclic AMP-Protein Kinase A pathway. We also found that another HDAC, Clr3, contributes to this regulation, possibly in cooperation with Hos2. As a whole, our data suggest that there is a direct link between changes in the environment and acetylation of nucleosomes within certain genes. We propose that histone acetylation is critical to the proper timing and induction of transcription of the genes encoding factors that coordinate changes in morphology with pathogenesis.

  4. Heterochromatin assembly and transcriptome repression by Set1 in coordination with a class II histone deacetylase

    PubMed Central

    Lorenz, David R; Meyer, Lauren F; Grady, Patrick J R; Meyer, Michelle M; Cam, Hugh P

    2014-01-01

    Histone modifiers play essential roles in controlling transcription and organizing eukaryotic genomes into functional domains. Here, we show that Set1, the catalytic subunit of the highly conserved Set1C/COMPASS complex responsible for histone H3K4 methylation (H3K4me), behaves as a repressor of the transcriptome largely independent of Set1C and H3K4me in the fission yeast Schizosaccharomyces pombe. Intriguingly, while Set1 is enriched at highly expressed and repressed loci, Set1 binding levels do not generally correlate with the levels of transcription. We show that Set1 is recruited by the ATF/CREB homolog Atf1 to heterochromatic loci and promoters of stress-response genes. Moreover, we demonstrate that Set1 coordinates with the class II histone deacetylase Clr3 in heterochromatin assembly at prominent chromosomal landmarks and repression of the transcriptome that includes Tf2 retrotransposons, noncoding RNAs, and regulators of development and stress-responses. Our study delineates a molecular framework for elucidating the functional links between transcriptome control and chromatin organization. DOI: http://dx.doi.org/10.7554/eLife.04506.001 PMID:25497836

  5. Histone deacetylase inhibitor significantly improved the cloning efficiency of porcine somatic cell nuclear transfer embryos.

    PubMed

    Huang, Yongye; Tang, Xiaochun; Xie, Wanhua; Zhou, Yan; Li, Dong; Yao, Chaogang; Zhou, Yang; Zhu, Jianguo; Lai, Liangxue; Ouyang, Hongsheng; Pang, Daxin

    2011-12-01

    Valproic acid (VPA), a histone deacetylase inbibitor, has been shown to generate inducible pluripotent stem (iPS) cells from mouse and human fibroblasts with a significant higher efficiency. Because successful cloning by somatic cell nuclear transfer (SCNT) undergoes a full reprogramming process in which the epigenetic state of a differentiated donor nuclear is converted into an embryonic totipotent state, we speculated that VPA would be useful in promoting cloning efficiency. Therefore, in the present study, we examined whether VPA can promote the developmental competence of SCNT embryos by improving the reprogramming state of donor nucleus. Here we report that 1 mM VPA for 14 to 16 h following activation significantly increased the rate of blastocyst formation of porcine SCNT embryos constructed from Landrace fetal fibroblast cells compared to the control (31.8 vs. 11.4%). However, we found that the acetylation level of Histone H3 lysine 14 and Histone H4 lysine 5 and expression level of Oct4, Sox2, and Klf4 was not significantly changed between VPA-treated and -untreated groups at the blastocyst stage. The SCNT embryos were transferred to 38 surrogates, and the cloning efficiency in the treated group was significantly improved compared with the control group. Taken together, we have demonstrated that VPA can improve both in vitro and in vivo development competence of porcine SCNT embryos.

  6. The effect of limited proteolysis by trypsin and chymotrypsin on bovine colostral IgG1.

    PubMed Central

    Brock, J H; Arzabe, F R; Ortega, F; Piñeiro, A

    1977-01-01

    Limited proteolysis of bovine colostral IgG1 by trypsin caused loss of specific antibody activity but column chromatography showed that relatively little cleavage into fragements had occurred. Polyacrlamide-agarose SDS electrophoresis of the 2-mercaptoethanol-treated digest revealed, however, that extensive cleavage of light chains had occurred even though most of the material before reduction had a mol. wt close to that of undigested IgG1. Although a Fab-type fragment was detected in the digest by immunoelectrophoresis it appeared to be only a minor component. Chymotrypsin had little effect upon either the structure or antibody activity of IgG1. These findings may explain the effect of trypsin and chymotrypsin on the bactericidal activity of colostral antibodies. Images Figure 2 Figure 3 PMID:321343

  7. Asymmetric High-Velocity Ejecta in the Youngest Galactic SNR G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Borkowski, Kazimierz

    2014-11-01

    Chandra has revealed highly asymmetric supernova ejecta in G1.9+0.3. Iron dominates thermal emission in the radio-bright northern rim, while only intermediate-mass elements are found along the SE-NW axis. The measured X-ray expansion rates decrease radially by about 60% along this axis from 0.84% yr^{-1) to 0.52% yr^{-1}. This corresponds to undecelerated ages of 120 - 190 yr, confirming the young age of G1.9+0.3, and implying that the blast wave is much more decelerated than the reverse shock. Only the outermost ejecta with very high (>18,000 km s^{-1}) free-expansion velocities have been shocked so far. We discuss G1.9+0.3 in the framework of recent asymmetric 3D delayed-detonation Type Ia explosions from Seitenzahl et al. (2013). Their N3 model provides the best match.

  8. Twisted K-theory in /g>1 from D-branes

    NASA Astrophysics Data System (ADS)

    Isidro, José M.

    2002-07-01

    We study the wrapping of N-type IIB D p-branes on a compact Riemann surface Σ in genus g>1 by means of the Sen-Witten construction, as a superposition of N'-type IIB D p'-brane/antibrane pairs, with p'> p. A background Neveu-Schwarz field B deforms the commutative C -algebra of functions on Σ to a non-commutative C -algebra. Our construction provides an explicit example of the N'→∞ limit advocated by Bouwknegt-Mathai and Witten in order to deal with twisted K-theory. We provide the necessary elements to formulate M(atrix) theory on this new C -algebra, by explicitly constructing a family of projective C -modules admitting constant-curvature connections. This allows us to define the g>1 analogue of the BPS spectrum of states in g=1, by means of Donaldson's formulation of the Narasimhan-Seshadri theorem.

  9. Acute appendicitis with a neuroendocrine tumor G1 (carcinoid): pitfalls of conservative treatment.

    PubMed

    Watanabe, Hiroyuki A; Fujimoto, Taketoshi; Kato, Yo; Sasaki, Mayumi; Ikusue, Toshikazu

    2016-08-01

    A man in his early thirties presented to our clinic with right lower abdominal pain. Computed tomography (CT) and ultrasonography (US) revealed a swollen appendix and an appendicolith. Abscess formation was not observed but ongoing appendiceal rupture was not ruled out. Three months after successful conservative therapy, the lumen of the apical portion was kept dilated and laparoscopic interval appendectomy was performed. No tumorous findings were observed macroscopically. However, histology revealed many tiny nests infiltrating the submucosa, muscular layer, and subserosa at the root of the appendix. An appendiceal neuroendocrine tumor G1 (NET G1; carcinoid) was diagnosed immunohistologically. Neither CT nor US visualized the tumor because of its non-tumor-forming but infiltrative growth. In conclusion, after successful conservative treatment, interval appendectomy should be considered to uncover a possible appendiceal NET G1 (carcinoid), particularly when dilatation of the distal lumen is kept under observation.

  10. ORC function in late G1: maintaining the license for DNA replication.

    PubMed

    Da-Silva, Lance F; Duncker, Bernard P

    2007-01-15

    The origin recognition complex (ORC) is essential as a scaffold for the assembly of prereplicative complexes (pre-RCs) in G(1) phase of the cell cycle. Some models have proposed that once origins have been licensed for DNA replication, ORC is dispensable for MCM protein association, and ensuing DNA replication. Although budding yeast Orc6 is not needed for origin recognition or binding in vitro, we have recently shown that this ORC subunit is required in late G(1) phase for maintenance of MCMs, and subsequent DNA replication. Further investigation shows that depletion of Orc6 results in displacement of MCM proteins from both early- and late-firing origins, and eventually results in the activation of the Rad53 checkpoint kinase, consistent with incomplete DNA replication. Loss of MCM association at origins may be mediated by the displacement of Mcm10 and/or Orc2 as a consequence of late G(1) Orc6 depletion.

  11. Nonuniform expansion and brightening of the youngest Galactic SNR G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Borkowski, Kazimierz

    2014-09-01

    We propose a 400-ks observation of the youngest Galactic supernova remnant (SNR) G1.9+0.3, to study its nonuniform expansion and monitor increase in brightness. Expansion along the major axis of G1.9+0.3 has been found to decrease with radius. The longer time baseline should help in understanding these surprising variations in expansion. No other Galactic SNR is brightening. The X-rays are mainly produced as synchrotron radiation from 10 -- 100 TeV electrons, so the magnitude and spatial dependence of the brightening rate have important implications for the physics of particle acceleration and magnetic-field amplification in fast shock waves. G1.9+0.3 is a unique SNR whose continued monitoring should greatly advance our understanding of Type Ia supernovae and nonthermal shock physics.

  12. G 1 Bézier Surface Generation from Given Boundary Curve Network with T-Junction

    NASA Astrophysics Data System (ADS)

    Oh, Min-Jae; Park, Sung Ha; Kim, Tae-Wan

    T-junctions usually appear in surface modeling processes that start with a given curve network. However, since T-shaped patches are not available in current CAD system so existing G 1 surface generation methods are restricted to n-sided patches. Therefore a designer must design a curve network without T-junctions, or subdivide it into n-sided patches, to avoid T-shaped topologies. We generate G 1 Bézier surfaces at a T-junction by combining the coplanar G 1 continuity condition with the de Casteljau algorithm to avoid the collision of twist points. Both T-junctions in the middle of boundary curves and at 3-valent vertices are considered. Our method requires no subdivision or triangulation of the surface, and the curve network is not changed.

  13. Histone deacetylase inhibitor BML-210 induces growth inhibition and apoptosis and regulates HDAC and DAPC complex expression levels in cervical cancer cells.

    PubMed

    Borutinskaite, Veronika V; Magnusson, Karl-Eric; Navakauskiene, Ruta

    2012-12-01

    Histone deacetylase inhibitors (HDACIs) represent a new class of targeted anti-cancer agents and different other diseases, like muscular disorders. A number of studies have shown that extracellular signal-activated kinases can target chromatin-modifying complexes directly and regulate their function. The molecular connection between the dystrophin-associated protein complex (DAPC) and chromatin has been described, by showing that NO signaling regulates histone deacetylase (HDAC) activity and influences gene expression in different cell types. In present study, we investigated HDACs changes in HeLa cells undergoing growth inhibition and apoptosis, caused by HDACI BML-210 and retinoic acid (ATRA). Cell cycle analysis indicated that HeLa cell treatment with 20 and 30 μM concentration of BML-210 increased the proportion of cells in G0/G1 phase, and caused accumulation in subG1, indicating that the cells are undergoing apoptosis. We determined down-regulation of HDAC 1-5 and 7 after treatment with BML-210. Also, we demonstrated expression of different isoforms of alpha-dystrobrevin (α-DB) and other components of DAPC such as syntrophin, dystrophin, beta-dystrobrevin (β-DB) and NOS in HeLa cells after treatments. We determined changes in protein expression level of dystrophin, NOS1, α- and β-DB and in subcellular localization of α-DB after treatments with BML-210 and ATRA. In conclusion, these results suggest that HDACI BML-210 can inhibit cell growth and induce apoptosis in cervical cancer cells, what correlates with down-regulation of HDAC class I and II and changes in the DAPC expression levels. This can be important for identifying target proteins in DAPC signaling to HDACs, as a target of pharmacological intervention for treatment of muscular dystrophies and other diseases.

  14. Extraction of monoclonal antibodies (IgG1) using anionic and anionic/nonionic reverse micelles.

    PubMed

    George, Daliya A; Stuckey, David C

    2010-01-01

    Purification schemes for antibody production based on affinity chromatography are trying to keep pace with increases in cell culture expression levels and many current research initiatives are focused on finding alternatives to chromatography for the purification of Monoclonal antibodies (MAbs). In this article, we have investigated an alternative separation technique based on liquid-liquid extraction called the reverse micellar extraction. We extracted MAb (IgG1) using reverse micelles of an anionic surfactant, sodium bis 2-ethyl-hexyl sulfosuccinate (AOT) and a combination of anionic (AOT) and nonionic surfactants (Brij-30, Tween-85, Span-85) using isooctane as the solvent system. The extraction efficiency of IgG1 was studied by varying parameters, such as pH of the aqueous phase, cation concentration, and type and surfactant concentration. Using the AOT/Isooctane reverse micellar system, we could achieve good overall extraction of IgG1 (between 80 and 90%), but only 30% of the bioactivity of IgG1 could be recovered at the end of the extraction by using its binding to affinity chromatography columns as a surrogate measure of activity. As anionic surfactants were suspected as being one of the reasons for the reduced activity, we decided to combine a nonionic surfactant with an anionic surfactant and then study its effect on the extraction efficiency and bioactivity. The best results were obtained using an AOT/Brij-30/Isooctane reverse micellar system, which gave an overall extraction above 90 and 59% overall activity recovery. An AOT/Tween-85/Isooctane reverse micellar system gave an overall extraction of between 75 and 80% and overall activity recovery of around 40-45%. The results showed that the activity recovery of IgG1 can be significantly enhanced using different surfactant combination systems, and if the recovery of IgG1 can be further enhanced, the technique shows considerable promise for the downstream purification of MAbs.

  15. Replication licensing promotes cyclin D1 expression and G1 progression in untransformed human cells

    PubMed Central

    Liu, Peijun; Slater, Damien M.; Lenburg, Marc; Nevis, Kathleen; Cook, Jeanette Gowen; Vaziri, Cyrus

    2011-01-01

    Defects in DNA replication are implicated as early and causal events in malignancy. However, the immediate effects of impaired DNA replication licensing on cell cycle progression of non-malignant human cells are unknown. Therefore, we have investigated the acute effects of Mcm7 ablation using synchronized cultures of untransformed Human Dermal Fibroblasts (HDF). Mcm7 ablation elicited a G1 delay associated with impaired activation of CDK4 and CDK2 and reduced Rb phosphorylation. The cell cycle delay of Mcm7-ablated cells was not associated with a DNA damage response. However, levels of cyclin D1 mRNA were specifically reduced and binding of RNA Polymerase II to the CYCD1 promoter was decreased in Mcm7-depleted cells. Similar to Mcm7-deficiency, Mcm2- or Cdc6-depletion led to impaired cyclin D expression. Ectopic overexpression of Cdc6 in quiescent cells promoted cyclin D1 expression, CDK4 activation and G1 progression. Therefore timely and efficient expression of cyclin D1 during G1 phase requires replication licensing. Reconstitution of cyclin D1 expression was insufficient to correct the G1 delay of Mcm7-depleted cells, indicating that additional cell cycle events during G1 are dependent on replication licensing. However, ectopic expression of the HPV-E7 oncoprotein, and the resulting bypass of the requirement for cyclin D1-Rb signaling enabled Mcm7-depleted cells to enter S-phase. HPV-E7-induced S-phase entry of Mcm7-depleted cells led to a DNA damage response, a hallmark of pre-malignancy. Taken together, our results suggest the existence of a ‘replication licensing restriction point’ that couples pre-RC assembly with G1 progression in normal cells to minimize replication stress, DNA damage and tumorigenesis. PMID:19106611

  16. Cytosolic phospholipase A2α regulates G1 progression through modulating FOXO1 activity

    PubMed Central

    Naini, Said Movahedi; Choukroun, Gabriel J.; Ryan, James R.; Hentschel, Dirk M.; Shah, Jagesh V.; Bonventre, Joseph V.

    2016-01-01

    Group IVA phospholipase A2 [cytosolic phospholipase A2α (cPLA2α)] is a key mediator of inflammation and tumorigenesis. In this study, by using a combination of chemical inhibition and genetic approaches in zebrafish and murine cells, we identify a mechanism by which cPLA2α promotes cell proliferation. We identified 2 cpla2α genes in zebrafish, cpla2αa and cpla2αb, with conserved phospholipase activity. In zebrafish, loss of cpla2α expression or inhibition of cpla2α activity diminished G1 progression through the cell cycle. This phenotype was also seen in both mouse embryonic fibroblasts and mesangial cells. G1 progression was rescued by the addition of arachidonic acid or prostaglandin E2 (PGE2), indicating a phospholipase-dependent mechanism. We further show that PGE2, through PI3K/AKT activation, promoted Forkhead box protein O1 (FOXO1) phosphorylation and FOXO1 nuclear export. This led to up-regulation of cyclin D1 and down-regulation of p27Kip1, thus promoting G1 progression. Finally, using pharmacologic inhibitors, we show that cPLA2α, rapidly accelerated fibrosarcoma (RAF)/MEK/ERK, and PI3K/AKT signaling pathways cooperatively regulate G1 progression in response to platelet-derived growth factor stimulation. In summary, these data indicate that cPLA2α, through its phospholipase activity, is a critical effector of G1 phase progression through the cell cycle and suggest that pharmacological targeting of this enzyme may have important therapeutic benefits in disease mechanisms that involve excessive cell proliferation, in particular, cancer and proliferative glomerulopathies.—Naini, S. M., Choukroun, G. J., Ryan, J. R., Hentschel, D. M., Shah, J. V., Bonventre, J. V. Cytosolic phospholipase A2α regulates G1 progression through modulating FOXO1 activity. PMID:26644349

  17. Molecular Analysis of VP7 Gene of Rotavirus G1 Strains Isolated from North India.

    PubMed

    Jain, Swapnil; Vashistt, Jitendraa; Gupta, Kanika; Kumar, Ashok; Changotra, Harish

    2016-12-01

    Rotavirus G1 strains are the predominant cause of diarrhoea in children. Universally common rotavirus vaccines (Rotarix and RotaTeq) include G1 as the immunological component. India has recently introduced rotavirus vaccine in Universal Immunization Programme. Therefore, in the present study, VP7 gene of rotavirus G1 strains circulating in Himachal Pradesh, India is analysed to study their phylogenetic characteristics, and further comparative analysis was performed for assessment of their divergence from the vaccine strains. The rotavirus strains (JU-SOL-5, JU-SOL-58, JU-SOL-77, JU-SOL-173 and JU-SHI-14) analysed in the study were isolated from the faeces of diarrhoeic children during active surveillance for rotaviruses. The Himachal strains clustered together in G1-Lineage 1 in the phylogenetic analysis. All five isolates showed 96.4-98.8 % similarity with the other G1-Lineage 1 strains at amino acid level. However, none of them clustered in the pre-defined sublineages within lineage 1. Interestingly, all the strains were distantly related to the vaccine strains having 93.9-94.5 and 91.9-92.6 % similarities at amino acid level with Rotarix and RotaTeq strains, respectively. The comparative sequence and structural analysis of the Himachal strains with vaccine strains revealed differences in amino acids in epitope region of the protein especially at the antibody neutralization sites. The study highlights variations between the G1 strains from Himachal Pradesh, India and Rotarix and RotaTeq vaccine strains. These differences might have an impact on the neutralization efficiency of vaccine and subsequently on vaccine efficacy. This underscores further investigation to study intragenotype antigenic variability and also impact of viral evolution on vaccine effectiveness.

  18. Histone Deacetylase Inhibitors Equipped with Estrogen Receptor Modulation Activity

    PubMed Central

    Gryder, Berkley E.; Rood, Michael K.; Johnson, Kenyetta A.; Patil, Vishal; Raftery, Eric D.; Yao, Li-Pan D.; Rice, Marcie; Azizi, Bahareh; Doyle, Donald F.; Oyelere, Adegboyega K.

    2013-01-01

    We described a set of novel histone deacetylase inhibitors (HDACi) equipped with either an antagonist or an agonist of the estrogen receptor (ER) to confer selective activity against breast cancers. These bifunctional compounds potently inhibit HDAC at nanomolar concentrations, and either agonize or antagonize ERα and ERβ. The ER antagonist activities of tamoxifen-HDACi conjugates (Tam-HDACi) are nearly identical to those of tamoxifen. Conversely, ethynyl-estradiol HDACi conjugates (EED-HDACi) have attenuated ER agonist activities relative to the parent ethynyl-estradiol. In silico docking analysis provides structural basis for the trends of ER agonism/antagonism and ER subtype selectivity. Excitingly, lead Tam-HDACi conjugates show anticancer activity that is selectively more potent against MCF-7 (ERα positive breast) compared to MDA-MB-231 (triple negative breast cancer), DU145 (prostate cancer) or Vero (non-cancerous cell line). This dual-targeting approach illustrates the utility of designing small molecules with an emphasis on cell-type selectivity, not merely improved potency, working towards a higher therapeutic index at the earliest stages of drug development. PMID:23786452

  19. The functional interactome landscape of the human histone deacetylase family

    PubMed Central

    Joshi, Preeti; Greco, Todd M; Guise, Amanda J; Luo, Yang; Yu, Fang; Nesvizhskii, Alexey I; Cristea, Ileana M

    2013-01-01

    Histone deacetylases (HDACs) are a diverse family of essential transcriptional regulatory enzymes, that function through the spatial and temporal recruitment of protein complexes. As the composition and regulation of HDAC complexes are only partially characterized, we built the first global protein interaction network for all 11 human HDACs in T cells. Integrating fluorescence microscopy, immunoaffinity purifications, quantitative mass spectrometry, and bioinformatics, we identified over 200 unreported interactions for both well-characterized and lesser-studied HDACs, a subset of which were validated by orthogonal approaches. We establish HDAC11 as a member of the survival of motor neuron complex and pinpoint a functional role in mRNA splicing. We designed a complementary label-free and metabolic-labeling mass spectrometry-based proteomics strategy for profiling interaction stability among different HDAC classes, revealing that HDAC1 interactions within chromatin-remodeling complexes are largely stable, while transcription factors preferentially exist in rapid equilibrium. Overall, this study represents a valuable resource for investigating HDAC functions in health and disease, encompassing emerging themes of HDAC regulation in cell cycle and RNA processing and a deeper functional understanding of HDAC complex stability. PMID:23752268

  20. Class IIa Histone Deacetylases Are Conserved Regulators of Circadian Function*

    PubMed Central

    Fogg, Paul C. M.; O'Neill, John S.; Dobrzycki, Tomasz; Calvert, Shaun; Lord, Emma C.; McIntosh, Rebecca L. L.; Elliott, Christopher J. H.; Sweeney, Sean T.; Hastings, Michael H.; Chawla, Sangeeta

    2014-01-01

    Class IIa histone deacetylases (HDACs) regulate the activity of many transcription factors to influence liver gluconeogenesis and the development of specialized cells, including muscle, neurons, and lymphocytes. Here, we describe a conserved role for class IIa HDACs in sustaining robust circadian behavioral rhythms in Drosophila and cellular rhythms in mammalian cells. In mouse fibroblasts, overexpression of HDAC5 severely disrupts transcriptional rhythms of core clock genes. HDAC5 overexpression decreases BMAL1 acetylation on Lys-537 and pharmacological inhibition of class IIa HDACs increases BMAL1 acetylation. Furthermore, we observe cyclical nucleocytoplasmic shuttling of HDAC5 in mouse fibroblasts that is characteristically circadian. Mutation of the Drosophila homolog HDAC4 impairs locomotor activity rhythms of flies and decreases period mRNA levels. RNAi-mediated knockdown of HDAC4 in Drosophila clock cells also dampens circadian function. Given that the localization of class IIa HDACs is signal-regulated and influenced by Ca2+ and cAMP signals, our findings offer a mechanism by which extracellular stimuli that generate these signals can feed into the molecular clock machinery. PMID:25271152

  1. Current evidence for histone deacetylase inhibitors in pancreatic cancer

    PubMed Central

    Koutsounas, Ioannis; Giaginis, Constantinos; Patsouris, Efstratios; Theocharis, Stamatios

    2013-01-01

    Pancreatic cancer is one of the most aggressive human cancers, with more than 200 000 deaths worldwide every year. Despite recent efforts, conventional treatment approaches, such as surgery and classic chemotherapy, have only slightly improved patient outcomes. More effective and well-tolerated therapies are required to reverse the current poor prognosis of this type of neoplasm. Among new agents, histone deacetylase inhibitors (HDACIs) are now being tested. HDACIs have multiple biological effects related to acetylation of histones and many non-histone proteins that are involved in regulation of gene expression, apoptosis, cell cycle progression and angiogenesis. HDACIs induce cell cycle arrest and can activate the extrinsic and intrinsic pathways of apoptosis in different cancer cell lines. In the present review, the main mechanisms by which HDACIs act in pancreatic cancer cells in vitro, as well as their antiproliferative effects in animal models are presented. HDACIs constitute a promising treatment for pancreatic cancer with encouraging anti-tumor effects, at well-tolerated doses. PMID:23430136

  2. Glucose metabolism as a target of histone deacetylase inhibitors.

    PubMed

    Wardell, Suzanne E; Ilkayeva, Olga R; Wieman, Heather L; Frigo, Daniel E; Rathmell, Jeffrey C; Newgard, Christopher B; McDonnell, Donald P

    2009-03-01

    The therapeutic efficacy of histone deacetylase inhibitors (HDACI) is generally attributed to their ability to alter gene expression secondary to their effects on the acetylation status of transcription factors and histones. However, because HDACIs exhibit similar transcriptional effects in most cells, the molecular basis for their therapeutic selectivity toward malignant cells is largely unknown. In this study, we report that HDACI, of distinct chemotypes, quantitatively inhibit glucose transporter 1 (GLUT1)-mediated glucose transport into multiple myeloma cells through both down-regulation of GLUT1 and inhibition of hexokinase 1 (HXK1) enzymatic activity. Unexpectedly, however, this inhibition of glucose utilization is accompanied by an increase in amino acid catabolism with no increase in fatty acid oxidation. Our findings suggest that an HDACI-induced change in carbon source preference could contribute to the therapeutic efficacy of these drugs by creating a pattern of fuel utilization that is incompatible with rapid tumor growth and survival. Furthermore, these results, which implicate glucose metabolism as a target of HDACI, suggest that caution should be exercised in attributing effects of this class of drug to primary alterations in gene transcription.

  3. Comparative and pharmacophore model for deacetylase SIRT1

    NASA Astrophysics Data System (ADS)

    Huhtiniemi, Tero; Wittekindt, Carsten; Laitinen, Tuomo; Leppänen, Jukka; Salminen, Antero; Poso, Antti; Lahtela-Kakkonen, Maija

    2006-09-01

    Sirtuins are NAD-dependent histone deacetylases, which cleave the acetyl-group from acetylated proteins, such as histones but also the acetyl groups from several transcription factors, and in this way can change their activities. Of all seven mammalian SirTs, the human sirtuin SirT1 has been the most extensively studied. However, there is no crystal structure or comparative model reported for SirT1. We have therefore built up a three-dimensional comparison model of the SirT1 protein catalytic core (domain area from residues 244 to 498 of the full length SirT1) in order to assist in the investigation of active site-ligand interactions and in the design of novel SirT1 inhibitors. In this study we also propose the binding-mode of recently reported set of indole-based inhibitors in SirT1. The site of interaction and the ligand conformation were predicted by the use of molecular docking techniques. To distinguish between active and inactive compounds, a post-docking filter based on H-bond network was constructed. Docking results were used to investigate the pharmacophore and to identify a filter for database mining.

  4. Inhibition of Histone Deacetylases in Inflammatory Bowel Diseases

    PubMed Central

    Glauben, Rainer; Siegmund, Britta

    2011-01-01

    This review, comprised of our own data and that of others, provides a summary overview of histone deacetylase (HDAC) inhibition on intestinal inflammation as well as inflammation-mediated carcinogenesis. Experimental colitis in mice represents an excellent in vivo model to define the specific cell populations and target tissues modulated by inhibitors of HDAC. Oral administration of either suberyolanilide hydroxamic acid (SAHA) or ITF2357 results in an amelioration in these models, as indicated by a significantly reduced colitis disease score and histological score. This effect was paralleled by suppression of proinflammatory cytokines at the site of inflammation as well as specific changes in the composition of cells within the lamina propria. In addition, tumor number and size was significantly reduced in two models of inflammation-driven tumorigenesis, namely interleukin (IL)-10–deficient mice and the azoxymethane–dextran sulfate sodium (DSS) model, respectively. The mechanisms affected by HDAC inhibition, contributing to this antiinflammatory and antiproliferative potency will be discussed in detail. Furthermore, with regard to the relevance in human inflammatory bowel disease, the doses of ITF2357 considered safe in humans and the corresponding serum concentrations are consistent with the efficacious dosing used in our in vivo as well as in vitro experiments. Thus, the data strongly suggest that HDAC inhibitors could serve as a therapeutic option in inflammatory bowel disease. PMID:21365125

  5. Metabolism as a key to histone deacetylase inhibition

    PubMed Central

    Rajendran, Praveen; Williams, David E.; Ho, Emily; Dashwood, Roderick H.

    2012-01-01

    There is growing interest in the epigenetic mechanisms that are dysregulated in cancer and other human pathologies. Under this broad umbrella, modulators of histone deacetylase (HDAC) activity have gained interest as both cancer chemopreventive and therapeutic agents. Of the first generation, FDA-approved HDAC inhibitors to have progressed to clinical trials, vorinostat represents a “direct acting” compound with structural features suitable for docking into the HDAC pocket, whereas romidepsin can be considered a prodrug that undergoes reductive metabolism to generate the active intermediate (a zinc-binding thiol). It is now evident that other agents, including those in the human diet, can be converted by metabolism to intermediates that affect HDAC activity. Examples are cited of short-chain fatty acids, seleno-α-keto acids, small molecule thiols, mercapturic acid metabolites, indoles, and polyphenols. The findings are discussed in the context of putative endogenous HDAC inhibitors generated by intermediary metabolism (e.g. pyruvate), the yin–yang of HDAC inhibition versus HDAC activation, and the screening assays that might be most appropriate for discovery of novel HDAC inhibitors in the future. PMID:21599534

  6. Therapeutic applications of histone deacetylase inhibitors in sarcoma.

    PubMed

    Tang, Fan; Choy, Edwin; Tu, Chongqi; Hornicek, Francis; Duan, Zhenfeng

    2017-09-01

    Sarcomas are a rare group of malignant tumors originating from mesenchymal stem cells. Surgery, radiation and chemotherapy are currently the only standard treatments for sarcoma. However, their response rates to chemotherapy are quite low. Toxic side effects and multi-drug chemoresistance make treatment even more challenging. Therefore, better drugs to treat sarcomas are needed. Histone deacetylase inhibitors (HDAC inhibitors, HDACi, HDIs) are epigenetic modifying agents that can inhibit sarcoma growth in vitro and in vivo through a variety of pathways, including inducing tumor cell apoptosis, causing cell cycle arrest, impairing tumor invasion and preventing metastasis. Importantly, preclinical studies have revealed that HDIs can not only sensitize sarcomas to chemotherapy and radiotherapy, but also increase treatment responses when combined with other chemotherapeutic drugs. Several phase I and II clinical trials have been conducted to assess the efficacy of HDIs either as monotherapy or in combination with standard chemotherapeutic agents or targeted therapeutic drugs for sarcomas. Combination regimen for sarcomas appear to be more promising than monotherapy when using HDIs. This review summarizes our current understanding and therapeutic applications of HDIs in sarcomas. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Purification and characterization of extracellular chin deacetylase from Colletotrichum lindemuthianum.

    PubMed

    Tokuyasu, K; Ohnishi-Kameyama, M; Hayashi, K

    1996-10-01

    Chitin deacetylase, active in the presence of acetate (96% of the enzymatic activity was retained in the presence of 100 mM sodium acetate), was purified to electrophoretic homogeneity from a culture filtrate of Colletotrichum lindemuthianum (944-fold with a recovery of 4.05%). The enzyme was induced in the medium after the eighth day of incubation simultaneously with the blackening of the medium. The molecular mass of the enzyme was 31.5 kDa and 33 kDa as judged by SDS-PAGE and gel filtration, respectively, suggesting that the enzyme is a single polypeptide. The optimum temperature was 60 degrees C and the optimum pH was 11.5-12.0 when glycol chitin was used as substrate. The enzyme was active toward glycol chitin, partially N-deacetylated water soluble chitin, and chitin oligomers the degrees of polymerization of which were more than four, but was less active with chitin trimer and dimer, and inactive with N-acetylglucosamine. The Km and kcat for glycol chitin were 2.55 mM and 27.1 s-1, respectively, and those for chitin pentamer were 414 microM and 83.2 s-1, respectively. The reaction rates of the enzyme toward glycol chitin and chitin oligomers seemed to follow the Michaelis-Menten kinetics.

  8. Histone deacetylase inhibitors induce autophagy through FOXO1-dependent pathways.

    PubMed

    Zhang, Jianbin; Ng, Shukie; Wang, Jigang; Zhou, Jing; Tan, Shi-Hao; Yang, Naidi; Lin, Qingsong; Xia, Dajing; Shen, Han-Ming

    2015-04-03

    Autophagy is a catabolic process in response to starvation or other stress conditions to sustain cellular homeostasis. At present, histone deacetylase inhibitors (HDACIs) are known to induce autophagy in cells through inhibition of mechanistic target of rapamycin (MTOR) pathway. FOXO1, an important transcription factor regulated by AKT, is also known to play a role in autophagy induction. At present, the role of FOXO1 in the HDACIs-induced autophagy has not been reported. In this study, we first observed that HDACIs increased the expression of FOXO1 at the mRNA and protein level. Second, we found that FOXO1 transcriptional activity was enhanced by HDACIs, as evidenced by increased FOXO1 nuclear accumulation and transcriptional activity. Third, suppression of FOXO1 function by siRNA knockdown or by a chemical inhibitor markedly blocked HDACIs-induced autophagy. Moreover, we found that FOXO1-mediated autophagy is achieved via its transcriptional activation, leading to a dual effect on autophagy induction: (i) enhanced expression of autophagy-related (ATG) genes, and (ii) suppression of MTOR via transcription of the SESN3 (sestrin 3) gene. Finally, we found that inhibition of autophagy markedly enhanced HDACIs-mediated cell death, indicating that autophagy serves as an important cell survival mechanism. Taken together, our studies reveal a novel function of FOXO1 in HDACIs-mediated autophagy in human cancer cells and thus support the development of a novel therapeutic strategy by combining HDACIs and autophagy inhibitors in cancer therapy.

  9. HISTONE DEACETYLASE 6 Represses Pathogen Defense Responses in Arabidopsis thaliana.

    PubMed

    Wang, Yizhong; Hu, Qin; Wu, Zhenjiang; Wang, Hui; Han, Shiming; Jin, Ye; Zhou, Jin; Zhang, Zhengfeng; Jiang, Jiafu; Shen, Yun; Shi, Huazhong; Yang, Wannian; Shi, Huazhong

    2017-08-02

    Plant defense mechanisms are suppressed in the absence of pathogen attack to prevent wasted energy and growth inhibition. However, how defense responses are repressed is not well understood. Histone Deacetylase 6 (HDA6) is a negative regulator of gene expression, and its role in pathogen defense response in plants is not known. In this study, a novel allele of hda6 (designated as shi5) with spontaneous defense response was isolated from a forward genetics screening in Arabidopsis. The shi5 mutant exhibited increased resistance to hemi-biotrophic bacterial pathogen Pst DC3000, constitutively activated expression of pathogen responsive genes including PR1, PR2, etc, and increased histone acetylation levels at the promoters of most tested genes that were up-regulated in shi5. In both wild type and shi5 plants, the expression and histone acetylation of these genes were upregulated by pathogen infection. HDA6 was found to bind to the promoters of these genes under both normal growth conditions and pathogen infection. Our research suggests that HDA6 is a general repressor of pathogen defense response and plays important roles in inhibiting and modulating the expression of pathogen responsive genes in Arabidopsis. This article is protected by copyright. All rights reserved.

  10. Histone deacetylase inhibitors promote the tumoricidal effect of HAMLET.

    PubMed

    Brest, Patrick; Gustafsson, Mattias; Mossberg, Ann-Kristin; Gustafsson, Lotta; Duringer, Caroline; Hamiche, Ali; Svanborg, Catharina

    2007-12-01

    Histone deacetylase inhibitors (HDIs) and HAMLET (human alpha-lactalbumin made lethal to tumor cells) interact with histones, modify the structure of chromatin, and trigger tumor cell death. This study investigated how the combination of HDIs and HAMLET influences cell viability, histone acetylation, and DNA integrity. The pretreatment of tumor cells with HDIs was shown to enhance the lethal effect of HAMLET and the histone hyperacetylation response to HDIs increased even further after HAMLET treatment. HDIs and HAMLET were shown to target different histone domains as HAMLET bound tailless core histones, whereas HDIs modify the acetylation of the histone tail. DNA damage in response to HAMLET was increased by HDIs. The DNA repair response (p21WAFI expression) was induced by both agonists but abolished when the two agonists were combined. The results suggest that the synergy of HDIs and HAMLET is based on different but converging death pathways, both involving chromatin alterations. We speculate that HAMLET and HDIs might be combined to promote tumor cell death in vivo.

  11. Role of histone deacetylases in gene regulation at nuclear lamina.

    PubMed

    Milon, Beatrice C; Cheng, Haibo; Tselebrovsky, Mikhail V; Lavrov, Sergei A; Nenasheva, Valentina V; Mikhaleva, Elena A; Shevelyov, Yuri Y; Nurminsky, Dmitry I

    2012-01-01

    Theoretical models suggest that gene silencing at the nuclear periphery may involve "closing" of chromatin by transcriptional repressors, such as histone deacetylases (HDACs). Here we provide experimental evidence confirming these predictions. Histone acetylation, chromatin compactness, and gene repression in lamina-interacting multigenic chromatin domains were analyzed in Drosophila S2 cells in which B-type lamin, diverse HDACs, and lamina-associated proteins were downregulated by dsRNA. Lamin depletion resulted in decreased compactness of the repressed multigenic domain associated with its detachment from the lamina and enhanced histone acetylation. Our data reveal the major role for HDAC1 in mediating deacetylation, chromatin compaction, and gene silencing in the multigenic domain, and an auxiliary role for HDAC3 that is required for retention of the domain at the lamina. These findings demonstrate the manifold and central involvement of class I HDACs in regulation of lamina-associated genes, illuminating a mechanism by which these enzymes can orchestrate normal and pathological development.

  12. Role of Histone Deacetylases in Gene Regulation at Nuclear Lamina

    PubMed Central

    Milon, Beatrice C.; Cheng, Haibo; Tselebrovsky, Mikhail V.; Lavrov, Sergei A.; Nenasheva, Valentina V.; Mikhaleva, Elena A.; Shevelyov, Yuri Y.; Nurminsky, Dmitry I.

    2012-01-01

    Theoretical models suggest that gene silencing at the nuclear periphery may involve “closing” of chromatin by transcriptional repressors, such as histone deacetylases (HDACs). Here we provide experimental evidence confirming these predictions. Histone acetylation, chromatin compactness, and gene repression in lamina-interacting multigenic chromatin domains were analyzed in Drosophila S2 cells in which B-type lamin, diverse HDACs, and lamina-associated proteins were downregulated by dsRNA. Lamin depletion resulted in decreased compactness of the repressed multigenic domain associated with its detachment from the lamina and enhanced histone acetylation. Our data reveal the major role for HDAC1 in mediating deacetylation, chromatin compaction, and gene silencing in the multigenic domain, and an auxiliary role for HDAC3 that is required for retention of the domain at the lamina. These findings demonstrate the manifold and central involvement of class I HDACs in regulation of lamina-associated genes, illuminating a mechanism by which these enzymes can orchestrate normal and pathological development. PMID:23226217

  13. Histone deacetylase regulation of ATM-mediated DNA damage signaling.

    PubMed

    Thurn, K Ted; Thomas, Scott; Raha, Paromita; Qureshi, Ian; Munster, Pamela N

    2013-10-01

    Ataxia-telangiectasia mutated (ATM) is a major regulator of the DNA damage response. ATM promotes the activation of BRCA1, CHK2, and p53 leading to the induction of response genes such as CDKN1A (p21), GADD45A, and RRM2B that promote cell-cycle arrest and DNA repair. The upregulation of these response genes may contribute to resistance of cancer cells to genotoxic therapies. Here, we show that histone deacetylases (HDAC) play a major role in mitigating the response of the ATM pathway to DNA damage. HDAC inhibition decreased ATM activation and expression, and attenuated the activation of p53 in vitro and in vivo. Select depletion of HDAC1 and HDAC2 was sufficient to modulate ATM activation, reduce GADD45A and RRM2B induction, and increase sensitivity to DNA strand breaks. The regulation of ATM by HDAC enzymes therefore suggests a vital role for HDAC1 and HDAC2 in the DNA damage response, and the potential use of the ATM pathway as a pharmacodynamic marker for combination therapies involving HDAC inhibitors. ©2013 AACR.

  14. Histone Deacetylase Inhibition Restores Retinal Pigment Epithelium Function in Hyperglycemia

    PubMed Central

    Desjardins, Danielle; Liu, Yueying; Crosson, Craig E.; Ablonczy, Zsolt

    2016-01-01

    In diabetic individuals, macular edema is a major cause of vision loss. This condition is refractory to insulin therapy and has been attributed to metabolic memory. The retinal pigment epithelium (RPE) is central to maintaining fluid balance in the retina, and this function is compromised by the activation of advanced glycation end-product receptors (RAGE). Here we provide evidence that acute administration of the RAGE agonist, glycated-albumin (gAlb) or vascular endothelial growth factor (VEGF), increased histone deacetylase (HDAC) activity in RPE cells. The administration of the class I/II HDAC inhibitor, trichostatin-A (TSA), suppressed gAlb-induced reductions in RPE transepithelial resistance (in vitro) and fluid transport (in vivo). Systemic TSA also restored normal RPE fluid transport in rats with subchronic hyperglycemia. Both gAlb and VEGF increased HDAC activity and reduced acetyl-α-tubulin levels. Tubastatin-A, a relatively specific antagonist of HDAC6, inhibited gAlb-induced changes in RPE cell resistance. These data are consistent with the idea that RPE dysfunction following exposure to gAlb, VEGF, or hyperglycemia is associated with increased HDAC6 activity and decreased acetyl-α-tubulin. Therefore, we propose inhibiting HDAC6 in the RPE as a potential therapy for preserving normal fluid homeostasis in the hyperglycemic retina. PMID:27617745

  15. Targeting macrophage Histone deacetylase 3 stabilizes atherosclerotic lesions

    PubMed Central

    Hoeksema, Marten A; Gijbels, Marion JJ; Van den Bossche, Jan; van der Velden, Saskia; Sijm, Ayestha; Neele, Annette E; Seijkens, Tom; Stöger, J Lauran; Meiler, Svenja; Boshuizen, Marieke CS; Dallinga-Thie, Geesje M; Levels, Johannes HM; Boon, Louis; Mullican, Shannon E; Spann, Nathanael J; Cleutjens, Jack P; Glass, Chris K; Lazar, Mitchell A; de Vries, Carlie JM; Biessen, Erik AL; Daemen, Mat JAP; Lutgens, Esther; de Winther, Menno PJ

    2014-01-01

    Macrophages are key immune cells found in atherosclerotic plaques and critically shape atherosclerotic disease development. Targeting the functional repertoire of macrophages may hold novel approaches for future atherosclerosis management. Here, we describe a previously unrecognized role of the epigenomic enzyme Histone deacetylase 3 (Hdac3) in regulating the atherosclerotic phenotype of macrophages. Using conditional knockout mice, we found that myeloid Hdac3 deficiency promotes collagen deposition in atherosclerotic lesions and thus induces a stable plaque phenotype. Also, macrophages presented a switch to anti-inflammatory wound healing characteristics and showed improved lipid handling. The pro-fibrotic phenotype was directly linked to epigenetic regulation of the Tgfb1 locus upon Hdac3 deletion, driving smooth muscle cells to increased collagen production. Moreover, in humans, HDAC3 was the sole Hdac upregulated in ruptured atherosclerotic lesions, Hdac3 associated with inflammatory macrophages, and HDAC3 expression inversely correlated with pro-fibrotic TGFB1 expression. Collectively, we show that targeting the macrophage epigenome can improve atherosclerosis outcome and we identify Hdac3 as a potential novel therapeutic target in cardiovascular disease. PMID:25007801

  16. Attenuation of Choroidal Neovascularization by Histone Deacetylase Inhibitor

    PubMed Central

    Chan, Nymph; He, Shikun; Spee, Christine K.; Ishikawa, Keijiro; Hinton, David R.

    2015-01-01

    Choroidal neovascularization (CNV) is a blinding complication of age-related macular degeneration that manifests as the growth of immature choroidal blood vessels through Bruch’s membrane, where they can leak fluid or hemorrhage under the retina. Here, we demonstrate that the histone deacetylase inhibitor (HDACi) trichostatin A (TSA) can down-regulate the pro-angiogenic hypoxia-inducible factor-1α and vascular endothelial growth factor (VEGF), and up-regulate the anti-angiogenic and neuro-protective pigment epithelium derived factor in human retinal pigment epithelial (RPE) cells. Most strikingly, TSA markedly down-regulates the expression of VEGF receptor-2 in human vascular endothelial cells and, thus, can knock down pro-angiogenic cell signaling. Additionally, TSA suppresses CNV-associated wound healing response and RPE epithelial-mesenchymal transdifferentiation. In the laser-induced model of CNV using C57Bl/6 mice, systemic administration of TSA significantly reduces fluorescein leakage and the size of CNV lesions at post—laser days 7 and 14 as well as the immunohistochemical expression of VEGF, VEGFR2, and smooth muscle actin in CNV lesions at post-laser day 7. This report suggests that TSA, and possibly HDACi’s in general, should be further evaluated for their therapeutic potential for the treatment of CNV. PMID:25807249

  17. Histone deacetylases inhibitors effects on Cryptococcus neoformans major virulence phenotypes.

    PubMed

    Brandão, Fabiana As; Derengowski, Lorena S; Albuquerque, Patrícia; Nicola, André M; Silva-Pereira, Ildinete; Poças-Fonseca, Marcio J

    2015-01-01

    Cryptococcus neoformans undergoes phenotypical changes during host infection in order to promote persistence and survival. Studies have demonstrated that such adaptations require alterations in gene transcription networks by distinct mechanisms. Drugs such as the histone deacetylases inhibitors (HDACi) Sodium Butyrate (NaBut) and Trichostatin A (TSA) can alter the chromatin conformation and have been used to modulate epigenetic states in the treatment of diseases such as cancer. In this work, we have studied the effect of NaBut and TSA on the expression of C. neoformans major virulence phenotypes and on the survival rate of an animal model infected with drugs-treated yeasts. Both drugs affected fungal growth at 37°C more intensely than at 30°C; nonetheless, drugs did not affect cell viability at the concentrations we studied. HDACi also provoked the reduction of the fungal capsule expansion. Phospholipases enzyme activity decreased; mating process and melanin synthesis were also affected by both inhibitors. NaBut led to an increase in the population of cells in G2/M. Treated yeast cells, which were washed in order to remove the drugs from the culture medium prior to the inoculation in the Galleria mellonela infection model, did not cause significant difference at the host survival curve when compared to non-treated cells. Overall, NaBut effects on the impairment of C. neoformans main virulence factors were more intense and stable than the TSA effects.

  18. Histone deacetylase inhibitors: understanding a new wave of anticancer agents.

    PubMed

    Villar-Garea, Ana; Esteller, Manel

    2004-11-01

    Cancer is as much an epigenetic disease as it is a genetic and cytogenetic disease. The discovery that drastic changes in DNA methylation and histone modifications are commonly found in human tumors has inspired various laboratories and pharmaceutical companies to develop and study epigenetic drugs. One of the most promising groups of agents is the inhibitors of histone deacetylases (HDACs), which have different biochemical and biologic properties but have a single common activity: induction of acetylation in histones, the key proteins in nucleosome and chromatin structure. One of the main mechanisms of action of HDAC inhibitors is the transcriptional reactivation of dormant tumor-suppressor genes, such as p21WAF1. However, their pleiotropic nature leaves open the possibility that their well-known differentiation, cell-cycle arrest and apoptotic properties are also involved in other functions associated with HDAC inhibition. Many phase I clinical trials indicate that HDAC inhibitors appear to be well-tolerated drugs. Thus, the field is ready for rigorous biologic and clinical scrutiny to validate the therapeutic potential of these drugs. Our current data indicate that the use of HDAC inhibitors, probably in association with classical chemotherapy drugs or in combination with DNA-demethylating agents, could be promising for cancer patients.

  19. Screening of selective histone deacetylase inhibitors by proteochemometric modeling

    PubMed Central

    2012-01-01

    Background Histone deacetylase (HDAC) is a novel target for the treatment of cancer and it can be classified into three classes, i.e., classes I, II, and IV. The inhibitors selectively targeting individual HDAC have been proved to be the better candidate antitumor drugs. To screen selective HDAC inhibitors, several proteochemometric (PCM) models based on different combinations of three kinds of protein descriptors, two kinds of ligand descriptors and multiplication cross-terms were constructed in our study. Results The results show that structure similarity descriptors are better than sequence similarity descriptors and geometry descriptors in the leftacterization of HDACs. Furthermore, the predictive ability was not improved by introducing the cross-terms in our models. Finally, a best PCM model based on protein structure similarity descriptors and 32-dimensional general descriptors was derived (R2 = 0.9897, Qtest2 = 0.7542), which shows a powerful ability to screen selective HDAC inhibitors. Conclusions Our best model not only predict the activities of inhibitors for each HDAC isoform, but also screen and distinguish class-selective inhibitors and even more isoform-selective inhibitors, thus it provides a potential way to discover or design novel candidate antitumor drugs with reduced side effect. PMID:22913517

  20. Histone deacetylases (HDACs): characterization of the classical HDAC family.

    PubMed Central

    de Ruijter, Annemieke J M; van Gennip, Albert H; Caron, Huib N; Kemp, Stephan; van Kuilenburg, André B P

    2003-01-01

    Transcriptional regulation in eukaryotes occurs within a chromatin setting, and is strongly influenced by the post-translational modification of histones, the building blocks of chromatin, such as methylation, phosphorylation and acetylation. Acetylation is probably the best understood of these modifications: hyperacetylation leads to an increase in the expression of particular genes, and hypoacetylation has the opposite effect. Many studies have identified several large, multisubunit enzyme complexes that are responsible for the targeted deacetylation of histones. The aim of this review is to give a comprehensive overview of the structure, function and tissue distribution of members of the classical histone deacetylase (HDAC) family, in order to gain insight into the regulation of gene expression through HDAC activity. SAGE (serial analysis of gene expression) data show that HDACs are generally expressed in almost all tissues investigated. Surprisingly, no major differences were observed between the expression pattern in normal and malignant tissues. However, significant variation in HDAC expression was observed within tissue types. HDAC inhibitors have been shown to induce specific changes in gene expression and to influence a variety of other processes, including growth arrest, differentiation, cytotoxicity and induction of apoptosis. This challenging field has generated many fascinating results which will ultimately lead to a better understanding of the mechanism of gene transcription as a whole. PMID:12429021

  1. Production of chitin deacetylase by Aspergillus flavus in submerged conditions.

    PubMed

    Narayanan, Karthik; Parameswaran, Binod; Pandey, Ashok

    2016-07-03

    Chitosan is a biopolymer obtained by deacetylation of chitin and has been proven to have various applications in industry and biomedicine. Deacetylation of chitin using the enzyme chitin deacetylase (CDA) is favorable in comparison to the hazardous chemical method involving strong alkali and high temperature. A fungal strain producing CDA was isolated from environmental samples collected from coastal regions of South Kerala, India. It was identified as Aspergillus flavus by morphological characteristics and ITS DNA analysis. Nutritional requirement for maximum production of CDA under submerged condition was optimized using statistical methods including Plackett-Burman and response surface methodology central composite design. A 5.98-fold enhancement in CDA production was attained in shake flasks when the fermentation process parameters were used at their optimum levels. The highest CDA activity was 57.69 ± 1.68 U under optimized bioprocess conditions that included 30 g L(-1) glucose, 40 g L(-1) yeast extract, 15 g L(-1) peptone, and 7 g L(-1) MgCl2 at initial media pH of 7 and incubation temperature of 32°C after 48 hr of incubation, while the unoptimized basal medium yielded 9.64 ± 2.04 U.

  2. Rectal Neuroendocrine Tumor G1 with a Solitary Hepatic Metastatic Lesion

    PubMed Central

    Nagata, Kohei; Tajiri, Kazuto; Shimada, Seitarou; Ando, Takayuki; Hosokawa, Ayumu; Matsui, Koshi; Imura, Joji; Sugiyama, Toshiro

    2017-01-01

    Rectal neuroendocrine tumor (NET) is a relatively rare tumor. NET is classified as G1, G2, or G3 according to the degree of mitosis or Ki-67 proliferation index, which reflect the malignant potential of the tumor, such as metastasis. Advanced cases with metastasis are indicated for chemotherapy treatment. However, the efficacy of chemotherapy is limited. Therefore, resection is considered, even in metastatic cases, if complete resection is possible. We herein report a case of small rectal NET discovered with hepatic metastasis classified as G1. The patient showed good progress with no recurrence after undergoing hepatectomy and endoscopic resection of rectal NET. PMID:28154272

  3. A Recombinant Fungal Chitin Deacetylase Produces Fully Defined Chitosan Oligomers with Novel Patterns of Acetylation

    PubMed Central

    Naqvi, Shoa; Cord-Landwehr, Stefan; Singh, Ratna; Bernard, Frank; Kolkenbrock, Stephan; El Gueddari, Nour Eddine

    2016-01-01

    ABSTRACT Partially acetylated chitosan oligosaccharides (paCOS) are potent biologics with many potential applications, and their bioactivities are believed to be dependent on their structure, i.e., their degrees of polymerization and acetylation, as well as their pattern of acetylation. However, paCOS generated via chemical N-acetylation or de-N-acetylation of GlcN or GlcNAc oligomers, respectively, typically display random patterns of acetylation, making it difficult to control and predict their bioactivities. In contrast, paCOS produced from chitin deacetylases (CDAs) acting on chitin oligomer substrates may have specific patterns of acetylation, as shown for some bacterial CDAs. However, compared to what we know about bacterial CDAs, we know little about the ability of fungal CDAs to produce defined paCOS with known patterns of acetylation. Therefore, we optimized the expression of a chitin deacetylase from the fungus Puccinia graminis f. sp. tritici in Escherichia coli. The best yield of functional enzyme was obtained as a fusion protein with the maltose-binding protein (MBP) secreted into the periplasmic space of the bacterial host. We characterized the MBP fusion protein from P. graminis (PgtCDA) and tested its activity on different chitinous substrates. Mass spectrometric sequencing of the products obtained by enzymatic deacetylation of chitin oligomers, i.e., tetramers to hexamers, revealed that PgtCDA generated paCOS with specific acetylation patterns of A-A-D-D, A-A-D-D-D, and A-A-D-D-D-D, respectively (A, GlcNAc; D, GlcN), indicating that PgtCDA cannot deacetylate the two GlcNAc units closest to the oligomer's nonreducing end. This unique property of PgtCDA significantly expands the so far very limited library of well-defined paCOS available to test their bioactivities for a wide variety of potential applications. IMPORTANCE We successfully achieved heterologous expression of a fungal chitin deacetylase gene from the basidiomycete Puccinia graminis f. sp

  4. Conversion of 11-hydroxy-O-methylsterigmatocystin to aflatoxin G1 in Aspergillus parasiticus

    USDA-ARS?s Scientific Manuscript database

    In aflatoxin biosynthesis, aflatoxins G1 (AFG1) and B1 (AFB1) are independently produced from a common precursor, O-methylsterigmatocystin (OMST). Recently, 11-hydroxy-O-methylsterigmatocystin (HOMST) was identified as a later precursor involved in the conversion of OMST to AFB1. However, the invo...

  5. Positive feedback of G1 cyclins ensures coherent cell cycle entry.

    PubMed

    Skotheim, Jan M; Di Talia, Stefano; Siggia, Eric D; Cross, Frederick R

    2008-07-17

    In budding yeast, Saccharomyces cerevisiae, the Start checkpoint integrates multiple internal and external signals into an all-or-none decision to enter the cell cycle. Here we show that Start behaves like a switch due to systems-level feedback in the regulatory network. In contrast to current models proposing a linear cascade of Start activation, transcriptional positive feedback of the G1 cyclins Cln1 and Cln2 induces the near-simultaneous expression of the approximately 200-gene G1/S regulon. Nuclear Cln2 drives coherent regulon expression, whereas cytoplasmic Cln2 drives efficient budding. Cells with the CLN1 and CLN2 genes deleted frequently arrest as unbudded cells, incurring a large fluctuation-induced fitness penalty due to both the lack of cytoplasmic Cln2 and insufficient G1/S regulon expression. Thus, positive-feedback-amplified expression of Cln1 and Cln2 simultaneously drives robust budding and rapid, coherent regulon expression. A similar G1/S regulatory network in mammalian cells, comprised of non-orthologous genes, suggests either conservation of regulatory architecture or convergent evolution.

  6. G1 arrest and differentiation can occur independently of Rb family function

    PubMed Central

    Wirt, Stacey E.; Adler, Adam S.; Gebala, Véronique; Weimann, James M.; Schaffer, Bethany E.; Saddic, Louis A.; Viatour, Patrick; Vogel, Hannes; Chang, Howard Y.; Meissner, Alex

    2010-01-01

    The ability of progenitor cells to exit the cell cycle is essential for proper embryonic development and homeostasis, but the mechanisms governing cell cycle exit are still not fully understood. Here, we tested the requirement for the retinoblastoma (Rb) protein and its family members p107 and p130 in G0/G1 arrest and differentiation in mammalian cells. We found that Rb family triple knockout (TKO) mouse embryos survive until days 9–11 of gestation. Strikingly, some TKO cells, including in epithelial and neural lineages, are able to exit the cell cycle in G0/G1 and differentiate in teratomas and in culture. This ability of TKO cells to arrest in G0/G1 is associated with the repression of key E2F target genes. Thus, G1 arrest is not always dependent on Rb family members, which illustrates the robustness of cell cycle regulatory networks during differentiation and allows for the identification of candidate pathways to inhibit the expansion of cancer cells with mutations in the Rb pathway. PMID:21059851

  7. Radio-Continuum Emission from the Young Galactic Supernova Remnant G1.9+0.3

    NASA Astrophysics Data System (ADS)

    De Horta, A. Y.; Filipovic, M. D.; Crawford, E. J.; Stootman, F. H.; Pannuti, T. G.; Bozzetto, L. M.; Collier, J. D.; Sommer, E. R.; Kosakowski, A. R.

    2014-12-01

    We present an analysis of a new Australia Telescope Compact Array (ATCA) radio-continuum observation of supernova remnant (SNR) G1.9+0.3, which at an age of ˜181±25 years is the youngest known in the Galaxy. We analysed all available radio-continuum observations at 6-cm from the ATCA and Very Large Array. Using this data we estimate an expansion rate for G1.9+0.3 of 0.563±0.078 percent per year between 1984 and 2009. We note that in the 1980's G1.9+0.3 expanded somewhat slower (0.484 percent per year) than more recently (0.641 percent per year). We estimate that the average spectral index between 20-cm and 6-cm, across the entire SNR is α=-0.72±0.26 which is typical for younger SNRs. At 6-cm, we detect an average of 6 percent fractionally polarised radio emission with a peak of 17±3 percent. The polarised emission follows the contours of the strongest of X-ray emission. Using the new equipartition formula we estimate a magnetic field strength of B≈273~μ G, which to date, is one of the highest magnetic field strength found for any SNR and consistent with G1.9+0.3 being a very young remnant.

  8. 17 CFR 275.202(a)(11)(G)-1 - Family offices.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 17 Commodity and Securities Exchanges 4 2014-04-01 2014-04-01 false Family offices. 275.202(a)(11... COMMISSION (CONTINUED) RULES AND REGULATIONS, INVESTMENT ADVISERS ACT OF 1940 § 275.202(a)(11)(G)-1 Family offices. (a) Exclusion. A family office, as defined in this section, shall not be considered to be an...

  9. 17 CFR 275.202(a)(11)(G)-1 - Family offices.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 17 Commodity and Securities Exchanges 3 2013-04-01 2013-04-01 false Family offices. 275.202(a)(11... COMMISSION (CONTINUED) RULES AND REGULATIONS, INVESTMENT ADVISERS ACT OF 1940 § 275.202(a)(11)(G)-1 Family offices. (a) Exclusion. A family office, as defined in this section, shall not be considered to be an...

  10. Lattice QCD results for the B --> D(*) l nu form factors: F(1) and G(1)

    SciTech Connect

    Van de Water, R.S.; /Fermilab

    2007-01-01

    I review the current status of lattice QCD calculations of the B {yields} D and B {yields} D* form factors and discuss prospects for their improvement. Successful calculations within the quenched approximation demonstrate the power of lattice methods for calculating F(1) and G(1), and the unquenched calculations in progress should soon allow for a 2-3% exclusive determination of |Vcb|.

  11. G1 cyclins link proliferation, pluripotency and differentiation of embryonic stem cells

    PubMed Central

    Liu, Lijun; Michowski, Wojciech; Inuzuka, Hiroyuki; Shimizu, Kouhei; Nihira, Naoe Taira; Chick, Joel M.; Li, Na; Geng, Yan; Meng, Alice Y.; Ordureau, Alban; Kolodziejczyk, Aleksandra; Ligon, Keith L.; Bronson, Roderick T.; Polyak, Kornelia; Harper, J. Wade; Gygi, Steven P.; Wei, Wenyi; Sicinski, Piotr

    2017-01-01

    Progression of mammalian cells through the G1 and S phases of the cell cycle is driven by D-type and E-type cyclins. According to the current models, at least one of these cyclin families must be present to allow cell proliferation. Here, we show that several cell types can proliferate in the absence of all G1 cyclins. However, upon ablation of G1 cyclins, embryonic stem (ES) cells attenuated their pluripotent characteristics, with majority of cells acquiring the trophectodermal cell fate. We established that G1 cyclins, together with their associated cyclin-dependent kinases (CDKs) phosphorylate and stabilize core pluripotency factors Nanog, Sox2 and Oct4. Treatment of murine ES cells, patient-derived glioblastoma tumor-initiating cells, or triple-negative breast cancer cells with a CDK-inhibitor strongly decreased Sox2 and Oct4 levels. Our findings suggest that CDK-inhibition might represent an attractive therapeutic strategy by targeting glioblastoma tumor-initiating cells, which depend on Sox2 to maintain their tumorigenic potential. PMID:28192421

  12. Positive feedback of G1 cyclins ensures coherent cell cycle entry

    PubMed Central

    Skotheim, Jan M.; Di Talia, Stefano; Siggia, Eric D.; Cross, Frederick R.

    2008-01-01

    In budding yeast, the Start checkpoint integrates multiple internal and external signals into an all-or-none decision to enter the cell cycle. Here, we show that Start behaves like a switch due to systems-level feedback in the regulatory network. In contrast to current models proposing a linear cascade of Start activation, transcriptional positive feedback of the G1 cyclins Cln1,2 induces the near-simultaneous expression of the ~200-gene G1/S regulon. Nuclear Cln2 drives coherent regulon expression, while cytoplasmic Cln2 drives efficient budding. cln1,2-deleted cells frequently arrest as unbudded cells, incurring a large fluctuation-induced fitness penalty due to both the lack of cytoplasmic Cln2 and insufficient G1/S regulon expression. Thus, positive-feedback-amplified expression of Cln1,2 simultaneously drives robust budding and rapid, coherent regulon expression. A similar G1/S regulatory network in mammalian cells, comprised of non-orthologous genes, suggests either the conservation of regulatory architecture or convergent evolution. PMID:18633409

  13. Human IgG1 antibodies suppress angiogenesis in a target-independent manner

    PubMed Central

    Bogdanovich, Sasha; Kim, Younghee; Mizutani, Takeshi; Yasuma, Reo; Tudisco, Laura; Cicatiello, Valeria; Bastos-Carvalho, Ana; Kerur, Nagaraj; Hirano, Yoshio; Baffi, Judit Z; Tarallo, Valeria; Li, Shengjian; Yasuma, Tetsuhiro; Arpitha, Parthasarathy; Fowler, Benjamin J; Wright, Charles B; Apicella, Ivana; Greco, Adelaide; Brunetti, Arturo; Ruvo, Menotti; Sandomenico, Annamaria; Nozaki, Miho; Ijima, Ryo; Kaneko, Hiroki; Ogura, Yuichiro; Terasaki, Hiroko; Ambati, Balamurali K; Leusen, Jeanette HW; Langdon, Wallace Y; Clark, Michael R; Armour, Kathryn L; Bruhns, Pierre; Verbeek, J Sjef; Gelfand, Bradley D; De Falco, Sandro; Ambati, Jayakrishna

    2016-01-01

    Aberrant angiogenesis is implicated in diseases affecting nearly 10% of the world’s population. The most widely used anti-angiogenic drug is bevacizumab, a humanized IgG1 monoclonal antibody that targets human VEGFA. Although bevacizumab does not recognize mouse Vegfa, it inhibits angiogenesis in mice. Here we show bevacizumab suppressed angiogenesis in three mouse models not via Vegfa blockade but rather Fc-mediated signaling through FcγRI (CD64) and c-Cbl, impairing macrophage migration. Other approved humanized or human IgG1 antibodies without mouse targets (adalimumab, alemtuzumab, ofatumumab, omalizumab, palivizumab and tocilizumab), mouse IgG2a, and overexpression of human IgG1-Fc or mouse IgG2a-Fc, also inhibited angiogenesis in wild-type and FcγR humanized mice. This anti-angiogenic effect was abolished by Fcgr1 ablation or knockdown, Fc cleavage, IgG-Fc inhibition, disruption of Fc-FcγR interaction, or elimination of FcRγ-initated signaling. Furthermore, bevacizumab’s Fc region potentiated its anti-angiogenic activity in humanized VEGFA mice. Finally, mice deficient in FcγRI exhibited increased developmental and pathological angiogenesis. These findings reveal an unexpected anti-angiogenic function for FcγRI and a potentially concerning off-target effect of hIgG1 therapies. PMID:26918197

  14. 26 CFR 25.2523(g)-1 - Special rule for charitable remainder trusts.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... (CONTINUED) ESTATE AND GIFT TAXES GIFT TAX; GIFTS MADE AFTER DECEMBER 31, 1954 Deductions § 25.2523(g)-1 Special rule for charitable remainder trusts. (a) In general. (1) With respect to gifts made after... the property under section 2523(f). (3) The donee spouse's interest need not be an interest for life...

  15. Regulation of p27Kip1 phosphorylation and G1 cell cycle progression by protein phosphatase PPM1G

    PubMed Central

    Sun, Chuang; Wang, Gaohang; Wrighton, Katharine H; Lin, Han; Songyang, Zhou; Feng, Xin-Hua; Lin, Xia

    2016-01-01

    The cell cycle, an essential process leading to the cell division, is stringently controlled by the key cell cycle regulators, cyclin-CDK complexes, whose activity is further regulated by a variety of mechanisms. p27Kip1 is a cyclin-CDK inhibitor that arrests the cell cycle at the G1 phase by blocking the activation of cyclin E-CDK2 complex, preventing the improper entry to the cell cycle. Dysfunction of p27 has been frequently observed in many types of human cancers, resulting from p27 protein degradation and cytoplasmic mislocalization, which are highly regulated by the phosphorylation status of p27. Although the kinases that phosphorylate p27 have been extensively studied, phosphatases that dephosphorylate p27 remain to be elucidated. By using genomic phosphatase screening, we identified a PPM family phosphatase, PPM1G, which could reduce p27 phosphorylation at T198. We further confirmed that PPM1G is a novel p27 phosphatase by demonstrating that PPM1G can interact with and dephosphorylate p27 in cells and in vitro. Functionally, ectopic expression of PPM1G enhanced p27 protein stability and delayed cell cycle progression from G1 to S phase. In accordance, knockdown of PPM1G accelerated p27 degradation during G1 phase and rendered cells resistant to the cell cycle arrest induced by serum deprivation. Mechanistically, PPM1G inhibited the interaction of p27 to 14-3-3θ, a chaperone protein that facilitates p27 nuclear export. Knockdown of PPM1G promoted the cytoplasmic localization of p27. Taken together, our studies identified PPM1G as a novel regulator of p27 that dephosphorylates p27 at T198 site and, together with p27 kinases, PPM1G controls cell cycle progression by maintaining the proper level of p27 protein. PMID:27822412

  16. Characterization and Expression Analysis of Common Bean Histone Deacetylase 6 during Development and Cold Stress Response

    PubMed Central

    Ligaba-Osena, Ayalew; Subramani, Mayavan; Brown, Adrianne; Melmaiee, Kalpalatha; Hossain, Khwaja

    2017-01-01

    Histone deacetylases (HDACs) are important regulators of gene transcription thus controlling multiple cellular processes. Despite its essential role in plants, HDA6 is yet to be validated in common bean. In this study, we show that HDA6 is involved in plant development and stress response. Differential expression of HDA6 was determined in various tissues and the expression was seen to be upregulated with plant age (seedling < flowering < maturity). Higher expression was observed in flowers and pods than in stem, leaf, and root. Upregulation of HDA6 gene during cold stress implies its prominent role in abiotic stress. Furthermore, the HDA6 gene was isolated from three common bean genotypes and sequence analyses revealed homology with functionally characterized homologs in model species. The 53 kDa translated product was detected using an HDA6 specific antibody and recombinant protein overexpressed in Escherichia coli showed HDAC activity in vitro. To our knowledge, this is the first report in the agriculturally important crop common bean describing the functional characterization and biological role of HDA6. PMID:28127547

  17. Inhibition of histone deacetylase in utero causes sociability deficits in postnatal mice.

    PubMed

    Moldrich, Randal X; Leanage, Gayeshika; She, David; Dolan-Evans, Elliot; Nelson, Michael; Reza, Nargis; Reutens, David C

    2013-11-15

    Exposure to sodium valproate (VPA) in utero increases the risk of language impairment and a diagnosis of autism spectrum disorder (ASD). Mice exposed to VPA while in utero have also shown postnatal social deficits. Inhibition of histone deacetylase (HDAC) is one of VPA's many biological effects. The main objective of this study was to test the hypothesis that HDAC inhibition causes these behavioral outcomes following prenatal VPA exposure in mice. We exposed embryonic mice to VPA, the HDAC inhibitor trichostatin A (TSA), or vehicle controls. TSA (1mg/kg) inhibited HDAC in embryonic tissue at a level comparable to 600 mg/kg VPA, resulting in significant increases in histone H3 and H4 acetylation, and histone H3 lysine 4 tri-methylation. Postnatally, decreases in ultrasonic vocalization, olfactory motivation and sociability were observed in TSA and VPA-exposed pups. Treated mice exhibited elevated digging and grooming suggestive of mild restrictive and repetitive behaviors. Olfactory social preference, social novelty and habituation were normal. Together, these data indicate that embryonic HDAC inhibition alone can cause abnormal social behaviors in mice. This result serves as a molecular understanding of infant outcomes following mild VPA exposure in utero.

  18. Emerging roles of SIRT1 deacetylase in regulating cardiomyocyte survival and hypertrophy

    PubMed Central

    Sundaresan, Nagalingam R; Pillai, Vinodkumar B.; Gupta, Mahesh P.

    2011-01-01

    Calorie restriction is considered to be the best environmental intervention providing health benefits to mammals. The underlying mechanism of this intervention seems to be controlled by a group of NAD-dependent deacetylases, collectively called sirtuins. In mammals there are seven sirutuin analogues, SIRT1 to SIRT7. The founding member of this family, SIRT1 is shown to protect cardiomyocytes from apoptosis and age-dependent degeneration in a dose dependent manner—protecting cells at low doses but showing detrimental effects at high doses. Studies performed with over expression or knockdown of SIRT1 indicated that though it protects cells from oxidative stress and ischemia-reperfusion injury, it promotes hypertrophy of cardiomyocytes. Activation of endogenous SIRT1 by resveratrol also displayed pro-survival and pro-hypertrophic activity of SIRT1. In this article we review recent findings documenting the role of SIRT1 in regulating cardiac myocyte growth and survival under stress, and the proposed mechanism behind its cardio protective effects. We also briefly discuss two other sirtuin analogues which have been shown to have cardioprotective effects. PMID:21276800

  19. Glycolysis-dependent histone deacetylase 4 degradation regulates inflammatory cytokine production

    PubMed Central

    Wang, Bin; Liu, Ting-yu; Lai, Chun-Hsiang; Rao, Yan-hua; Choi, Moon-Chang; Chi, Jen-Tsan; Dai, Jian-wu; Rathmell, Jeffrey C.; Yao, Tso-Pang

    2014-01-01

    Activation of the inflammatory response is accompanied by a metabolic shift to aerobic glycolysis. Here we identify histone deacetylase 4 (HDAC4) as a new component of the immunometabolic program. We show that HDAC4 is required for efficient inflammatory cytokine production activated by lipopolysaccharide (LPS). Surprisingly, prolonged LPS treatment leads to HDAC4 degradation. LPS-induced HDAC4 degradation requires active glycolysis controlled by GSK3β and inducible nitric oxide synthase (iNOS). Inhibition of GSK3β or iNOS suppresses nitric oxide (NO) production, glycolysis, and HDAC4 degradation. We present evidence that sustained glycolysis induced by LPS treatment activates caspase-3, which cleaves HDAC4 and triggers its degradation. Of importance, a caspase-3–resistant mutant HDAC4 escapes LPS-induced degradation and prolongs inflammatory cytokine production. Our findings identify the GSK3β-iNOS-NO axis as a critical signaling cascade that couples inflammation to metabolic reprogramming and a glycolysis-driven negative feedback mechanism that limits inflammatory response by triggering HDAC4 degradation. PMID:25187650

  20. Glycolysis-dependent histone deacetylase 4 degradation regulates inflammatory cytokine production.

    PubMed

    Wang, Bin; Liu, Ting-Yu; Lai, Chun-Hsiang; Rao, Yan-hua; Choi, Moon-Chang; Chi, Jen-Tsan; Dai, Jian-wu; Rathmell, Jeffrey C; Yao, Tso-Pang

    2014-11-01

    Activation of the inflammatory response is accompanied by a metabolic shift to aerobic glycolysis. Here we identify histone deacetylase 4 (HDAC4) as a new component of the immunometabolic program. We show that HDAC4 is required for efficient inflammatory cytokine production activated by lipopolysaccharide (LPS). Surprisingly, prolonged LPS treatment leads to HDAC4 degradation. LPS-induced HDAC4 degradation requires active glycolysis controlled by GSK3β and inducible nitric oxide synthase (iNOS). Inhibition of GSK3β or iNOS suppresses nitric oxide (NO) production, glycolysis, and HDAC4 degradation. We present evidence that sustained glycolysis induced by LPS treatment activates caspase-3, which cleaves HDAC4 and triggers its degradation. Of importance, a caspase-3-resistant mutant HDAC4 escapes LPS-induced degradation and prolongs inflammatory cytokine production. Our findings identify the GSK3β-iNOS-NO axis as a critical signaling cascade that couples inflammation to metabolic reprogramming and a glycolysis-driven negative feedback mechanism that limits inflammatory response by triggering HDAC4 degradation. © 2014 Wang et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  1. HDAC6: A Novel Histone Deacetylase Implicated in Pulmonary Arterial Hypertension.

    PubMed

    Boucherat, Olivier; Chabot, Sophie; Paulin, Roxane; Trinh, Isabelle; Bourgeois, Alice; Potus, François; Lampron, Marie-Claude; Lambert, Caroline; Breuils-Bonnet, Sandra; Nadeau, Valérie; Paradis, Renée; Goncharova, Elena A; Provencher, Steeve; Bonnet, Sébastien

    2017-07-03

    Pulmonary arterial hypertension (PAH) is a vascular remodeling disease with limited therapeutic options. Although exposed to stressful conditions, pulmonary artery (PA) smooth muscle cells (PASMCs) exhibit a "cancer-like" pro-proliferative and anti-apoptotic phenotype. HDAC6 is a cytoplasmic histone deacetylase regulating multiple pro-survival mechanisms and overexpressed in response to stress in cancer cells. Due to the similarities between cancer and PAH, we hypothesized that HDAC6 expression is increased in PAH-PASMCs to face stress allowing them to survive and proliferate, thus contributing to vascular remodeling in PAH. We found that HDAC6 is significantly up-regulated in lungs, distal PAs, and isolated PASMCs from PAH patients and animal models. Inhibition of HDAC6 reduced PAH-PASMC proliferation and resistance to apoptosis in vitro sparing control cells. Mechanistically, we demonstrated that HDAC6 maintains Ku70 in a hypoacetylated state, blocking the translocation of Bax to mitochondria and preventing apoptosis. In vivo, pharmacological inhibition of HDAC6 improved established PAH in two experimental models and can be safely given in combination with currently approved PAH therapies. Moreover, Hdac6 deficient mice were partially protected against chronic hypoxia-induced pulmonary hypertension. Our study shows for the first time that HDAC6 is implicated in PAH development and represents a new promising target to improve PAH.

  2. Novel Class IIa-Selective Histone Deacetylase Inhibitors Discovered Using an in Silico Virtual Screening Approach.

    PubMed

    Hsu, Kai-Cheng; Liu, Chang-Yi; Lin, Tony Eight; Hsieh, Jui-Hua; Sung, Tzu-Ying; Tseng, Hui-Ju; Yang, Jinn-Moon; Huang, Wei-Jan

    2017-06-12

    Histone deacetylases (HDAC) contain eighteen isoforms that can be divided into four classes. Of these isoform enzymes, class IIa (containing HDAC4, 5, 7 and 9) target unique substrates, some of which are client proteins associated with epigenetic control. Class IIa HDACs are reportedly associated with some neuronal disorders, making HDACs therapeutic targets for treating neurodegenerative diseases. Additionally, some reported HDAC inhibitors contain hydroxamate moiety that chelates with zinc ion to become the cofactor of HDAC enzymes. However, the hydroxamate functional group is shown to cause undesirable effects and has poor pharmacokinetic profile. This study used in silico virtual screening methodology to identify several nonhydroxamate compounds, obtained from National Cancer Institute database, which potentially inhibited HDAC4. Comparisons of the enzyme inhibitory activity against a panel of HDAC isoforms revealed these compounds had strong inhibitory activity against class IIa HDACs, but weak inhibitory activity against class I HDACs. Further analysis revealed that a single residue affects the cavity size between class I and class IIa HDACs, thus contributing to the selectivity of HDAC inhibitors discovered in this study. The discovery of these inhibitors presents the possibility of developing new therapeutic treatments that can circumvent the problems seen in traditional hydroxamate-based drugs.

  3. Selective transcription and cellular proliferation induced by PDGF require histone deacetylase activity

    SciTech Connect

    Catania, Annunziata; Iavarone, Carlo; Carlomagno, Stella M.; Chiariello, Mario . E-mail: chiariel@unina.it

    2006-05-05

    Histone deacetylases (HDACs) are key regulatory enzymes involved in the control of gene expression and their inhibition by specific drugs has been widely correlated to cell cycle arrest, terminal differentiation, and apoptosis. Here, we investigated whether HDAC activity was required for PDGF-dependent signal transduction and cellular proliferation. Exposure of PDGF-stimulated NIH3T3 fibroblasts to the HDAC inhibitor trichostatin A (TSA) potently repressed the expression of a group of genes correlated to PDGF-dependent cellular growth and pro-survival activity. Moreover, we show that TSA interfered with STAT3-dependent transcriptional activity induced by PDGF. Still, neither phosphorylation nor nuclear translocation and DNA-binding in vitro and in vivo of STAT3 were affected by using TSA to interfere with PDGF stimulation. Finally, TSA treatment resulted in the suppression of PDGF-dependent cellular proliferation without affecting cellular survival of NIH3T3 cells. Our data indicate that inhibition of HDAC activity antagonizes the mitogenic effect of PDGF, suggesting that these drugs may specifically act on the expression of STAT-dependent, PDGF-responsive genes.

  4. Histone Deacetylase 4 promotes cholestatic liver injury in the absence of Prohibitin-1

    PubMed Central

    Barbier-Torres, Lucía; Beraza, Naiara; Fernández-Tussy, Pablo; Lopitz-Otsoa, Fernando; Fernández-Ramos, David; Zubiete-Franco, Imanol; Varela-Rey, Marta; Delgado, Teresa C; Gutiérrez, Virginia; Anguita, Juan; Pares, Albert; Banales, Jesús M; Villa, Erica; Caballería, Juan; Alvarez, Luis; Lu, Shelly C; Mato, Jose M; Martínez-Chantar, María Luz

    2015-01-01

    Prohibitin 1 (PHB1) is an evolutionary conserved pleiotropic protein that participates in diverse processes depending on its subcellular localization and interactome. Recent data have indicated a diverse role for PHB1 in the pathogenesis of obesity, cancer and inflammatory bowel disease, among others. Data presented here suggest that PHB1 is also linked to cholestatic liver disease. PHB1 expression is markedly reduced in patients with primary billiary cirrhosis and biliary atresia and Alagille syndrome, two major pediatric cholestatic conditions. In the experimental model of bile duct ligation, silencing of PHB1 induced liver fibrosis, reduced animal survival and induced bile duct proliferation. Importantly, the modulatory effect of PHB1 is not dependent on its known mitochondrial function. Importantly, d PHB1 interacts with Histone Deacetylase 4 (HDAC4) in the presence of bile acids. Hence, PHB1 depletion leads to increased nuclear HDAC4 content and its associated epigenetic changes. Remarkably, HDAC4 silencing and the administration of the HDAC inhibitor parthenolide during obstructive cholestasis in vivo promote genomic reprogramming leading to the regression of the fibrotic phenotype in the liver-specific Phb1 KO mice. Conclusion Our data identify PHB1 as an important mediator of cholestatic liver injury regulating the activity of HDAC4, which controls specific epigenetic marks. These results identify potential novel strategies to treat liver injury and fibrosis, particularly as a consequence of chronic cholestasis. PMID:26109312

  5. RPD3 histone deacetylase and nutrition have distinct but interacting effects on Drosophila longevity.

    PubMed

    Frankel, Stewart; Woods, Jared; Ziafazeli, Tahereh; Rogina, Blanka

    2015-12-01

    Single-gene mutations that extend longevity have revealed regulatory pathways related to aging and longevity. RPD3 is a conserved histone deacetylase (Class I HDAC). Previously we showed that Drosophila rpd3 mutations increase longevity. Here we tested the longevity effects of RPD3 on multiple nutrient levels. Dietary restriction (DR) has additive effects on RPD3-mediated longevity extension, but the effect may be modestly attenuated relative to controls. RPD3 and DR therefore appear to operate by distinct but interacting mechanisms. Since RPD3 regulates transcription, the mRNA levels for two proteins involved in nutrient signaling, 4E-BP and Tor, were examined in rpd3 mutant flies. 4E-BP mRNA was reduced under longevity-increasing conditions. Epistasis between RPD3 and 4E-BP with regard to longevity was then tested. Flies only heterozygous for a mutation in Thor, the 4E-BP gene, have modestly decreased life spans. Flies mutant for both rpd3 and Thor show a superposition of a large RPD3-mediated increase and a small Thor-mediated decrease in longevity at all food levels, consistent with each gene product having distinct effects on life span. However, DR-mediated extension was absent in males carrying both mutations and lessened in females. Our results support the view that multiple discrete but interacting mechanisms regulate longevity.

  6. Histone deacetylase inhibitors: Future therapeutics for insulin resistance and type 2 diabetes.

    PubMed

    Sharma, Sorabh; Taliyan, Rajeev

    2016-11-01

    Insulin resistance is a common feature of obesity and predisposes the affected individuals to a variety of pathologies, including type 2 diabetes mellitus (T2DM), dyslipidemias, hypertension, cardiovascular disease etc. Insulin resistance is the primary cause of T2DM and it occurs many years before the disease onset. Although Thiazolidinediones (TZDs) such as rosiglitazone and pioglitazone are outstanding insulin sensitizers and are in clinical use since 1990s, however, their serious side effects such as heart attack and bladder cancer have limited their utilization. Thus, there is an unmet need to identify a new class of drugs with insulin sensitizing activity and minimal side effects. In the recent years, Histone deacetylase (HDAC) has emerged as a new molecular target in the control of insulin resistance and T2DM. The level of histone acetylation/deacetylation has been found to be altered during insulin resistance and T2DM conditions. HDAC inhibitors have been found to effectively manage insulin resistance and T2DM in various preclinical models and clinical trials. In this review we will focus on various aspects related to regulation of insulin signalling by HDACs and the future scope of HDAC inhibitors as therapeutics for insulin resistance.

  7. Antitumor Action of a Novel Histone Deacetylase Inhibitor, YF479, in Breast Cancer1

    PubMed Central

    Zhang, Tao; Chen, Yihua; Li, Jingjie; Yang, Feifei; Wu, Haigang; Dai, Fujun; Hu, Meichun; Lu, Xiaoling; Peng, Yi; Liu, Mingyao; Zhao, Yongxiang; Yi, Zhengfang

    2014-01-01

    Accumulating evidence demonstrates important roles for histone deacetylase in tumorigenesis (HDACs), highlighting them as attractive targets for antitumor drug development. Histone deactylase inhibitors (HDACIs), which have shown favorable anti-tumor activity with low toxicity in clinical investigations, are a promising class of anticancer therapeutics. Here, we screened our compound library to explore small molecules that possess anti-HDAC activity and identified a novel HDACI, YF479. Suberoylanilide hydroxamic acid (SAHA), which was the first approved HDAC inhibitor for clinical treatment by the FDA, was as positive control in our experiments. We further demonstrated YF479 abated cell viability, suppressed colony formation and tumor cell motility in vitro. To investigate YF479 with superior pharmacodynamic properties, we developed spontaneous and experimental breast cancer animal models. Our results showed YF479 significantly inhibited breast tumor growth and metastasis in vivo. Further study indicated YF479 suppressed both early and end stages of metastatic progression. Subsequent adjuvant chemotherapy animal experiment revealed the elimination of local-regional recurrence (LRR) and distant metastasis by YF479. More important, YF479 remarkably prolonged the survival of tumor-bearing mice. Intriguingly, YF479 displayed more potent anti-tumor activity in vitro and in vivo compared with SAHA. Together, our results suggest that YF479, a novel HDACI, inhibits breast tumor growth, metastasis and recurrence. In light of these results, YF479 may be an effective therapeutic option in clinical trials for patients burdened by breast cancer. PMID:25220594

  8. Histone Deacetylases and Mood Disorders: Epigenetic Programming in Gene-Environment Interactions

    PubMed Central

    Machado-Vieira, Rodrigo; Ibrahim, Lobna; Zarate, Carlos A.

    2010-01-01

    Epigenetics involves molecular mechanisms related to gene expression independent of DNA sequence, mostly mediated by modification of chromatin histones. It has recently been suggested that these transcriptional changes may be implicated in the pathophysiology of mood disorders. In addition, histone deacetylase (HDAC) inhibitors have been shown to control epigenetic programming associated with the regulation of cognition and behavior, and may reverse dysfunctional epigenetic regulation associated with early life events in preclinical models. In this context, the active and continuous adaptation of chromatin, and the access of gene promoters to transcription factor mechanisms may represent a potential therapeutic target in the treatment of mood disorders such as bipolar disorder (BD) and major depressive disorder (MDD). Notably, the standard mood stabilizer valproate (VPA) has been shown to modulate the epigenome by inhibiting HDACs. However, several potential limitations are associated with this class of agents, including lack of selectivity for specific HDAC isoforms as well as risk of potentially serious side effects. Further studies regarding the potential role of chromatin remodeling in the mechanism of action of antidepressants and mood stabilizers are necessary to clarify the potential role of this class of agents as therapeutics for mood disorders. PMID:20961400

  9. Unliganded Thyroid Hormone Receptor Regulates Metamorphic Timing via the Recruitment of Histone Deacetylase Complexes

    PubMed Central

    2014-01-01

    Anuran metamorphosis involves a complex series of tissue transformations that change an aquatic tadpole to a terrestrial frog and resembles the postembryonic perinatal period in mammals. Thyroid hormone (TH) plays a causative role in amphibian metamorphosis and its effect is mediated by TH receptors (TRs). Molecular analyses during Xenopus development have shown that unliganded TR recruits histone deacetylase (HDAC)-containing N-CoR/SMRT complexes and causes histone deacetylation at target genes while liganded TR leads to increased histone acetylations and altered histone methylations at target genes. Transgenic studies involving mutant TR-cofactors have shown that corepressor recruitment by unliganded TR is required to ensure proper timing of the onset of metamorphosis while coactivator levels influence the rate of metamorphic progression. In addition, a number of factors that can influence cellular free TH levels appear to contribute the timing of metamorphic transformations of different organs by regulating the levels of unliganded vs. liganded TR in an organ-specific manner. Thus, the recruitment of HDAC-containing corepressor complexes by unliganded TR likely controls both the timing of the initiation of metamorphosis and the temporal regulation of organ-specific transformations. Similar mechanisms likely mediate TR function in mammals as the maturation of many organs during postembryonic development is dependent upon TH and resembles organ metamorphosis in amphibians. PMID:23962846

  10. Histone deacetylase 8 regulates cortactin deacetylation and contraction in smooth muscle tissues.

    PubMed

    Li, Jia; Chen, Shu; Cleary, Rachel A; Wang, Ruping; Gannon, Olivia J; Seto, Edward; Tang, Dale D

    2014-08-01

    Histone deacetylases (HDACs) are a family of enzymes that mediate nucleosomal histone deacetylation and gene expression. Some members of the HDAC family have also been implicated in nonhistone protein deacetylation, which modulates cell-cycle control, differentiation, and cell migration. However, the role of HDACs in smooth muscle contraction is largely unknown. Here, HDAC8 was localized both in the cytoplasm and the nucleus of mouse and human smooth muscle cells. Knockdown of HDAC8 by lentivirus-encoding HDAC8 shRNA inhibited force development in response to acetylcholine. Treatment of smooth muscle tissues with HDAC8 inhibitor XXIV (OSU-HDAC-44) induced relaxation of precontracted smooth muscle tissues. In addition, cortactin is an actin-regulatory protein that undergoes deacetylation during migration of NIH 3T3 cells. In this study, acetylcholine stimulation induced cortactin deacetylation in mouse and human smooth muscle tissues, as evidenced by immunoblot analysis using antibody against acetylated lysine. Knockdown of HDAC8 by RNAi or treatment with the inhibitor attenuated cortactin deacetylation and actin polymerization without affecting myosin activation. Furthermore, expression of a charge-neutralizing cortactin mutant inhibited contraction and actin dynamics during contractile activation. These results suggest a novel mechanism for the regulation of smooth muscle contraction. In response to contractile stimulation, HDAC8 may mediate cortactin deacetylation, which subsequently promotes actin filament polymerization and smooth muscle contraction.

  11. Boric acid inhibits embryonic histone deacetylases: A suggested mechanism to explain boric acid-related teratogenicity

    SciTech Connect

    Di Renzo, Francesca; Cappelletti, Graziella; Broccia, Maria L.; Giavini, Erminio; Menegola, Elena . E-mail: elena.menegola@unimi.it

    2007-04-15

    Histone deacetylases (HDAC) control gene expression by changing histonic as well as non histonic protein conformation. HDAC inhibitors (HDACi) are considered to be among the most promising drugs for epigenetic treatment for cancer. Recently a strict relationship between histone hyperacetylation in specific tissues of mouse embryos exposed to two HDACi (valproic acid and trichostatin A) and specific axial skeleton malformations has been demonstrated. The aim of this study is to verify if boric acid (BA), that induces in rodents malformations similar to those valproic acid and trichostatin A-related, acts through similar mechanisms: HDAC inhibition and histone hyperacetylation. Pregnant mice were treated intraperitoneally with a teratogenic dose of BA (1000 mg/kg, day 8 of gestation). Western blot analysis and immunostaining were performed with anti hyperacetylated histone 4 (H4) antibody on embryos explanted 1, 3 or 4 h after treatment and revealed H4 hyperacetylation at the level of somites. HDAC enzyme assay was performed on embryonic nuclear extracts. A significant HDAC inhibition activity (compatible with a mixed type partial inhibition mechanism) was evident with BA. Kinetic analyses indicate that BA modifies substrate affinity by a factor {alpha} = 0.51 and maximum velocity by a factor {beta} = 0.70. This work provides the first evidence for HDAC inhibition by BA and suggests such a molecular mechanism for the induction of BA-related malformations.

  12. Inhibition of class I histone deacetylase with an apicidin derivative prevents cardiac hypertrophy and failure.

    PubMed

    Gallo, Pasquale; Latronico, Michael V G; Gallo, Paolo; Grimaldi, Serena; Borgia, Francesco; Todaro, Matilde; Jones, Philip; Gallinari, Paola; De Francesco, Raffaele; Ciliberto, Gennaro; Steinkühler, Christian; Esposito, Giovanni; Condorelli, Gianluigi

    2008-12-01

    Recent studies have demonstrated the importance of chromatin remodelling via histone acetylation/deacetylation for the control of cardiac gene expression. Specific histone deacetylases (HDACs) can, in fact, play a positive or negative role in determining cardiac myocyte (CM) size. Here, we report on the effect on hypertrophy development of three inhibitors (HDACi) of class I HDACs. The compounds were first analysed in vitro by scoring hypertrophy, expression of foetal genes, and apoptosis of neonatal rat CMs stimulated with phenylephrine, an alpha1-adrenergic agonist. This initial screening indicated that a truncated derivative of apicidin with class I HDAC specificity, denoted API-D, had the highest efficacy to toxicity ratio, and was thus selected for further analysis in vivo. Administration of this drug significantly decreased myocardial hypertrophy and foetal gene expression after 1 week of pressure overload induced by thoracic aortic constriction (TAC) in mice. After 9 weeks of TAC, when manifest heart failure is encountered, mice treated with API-D presented with significantly improved echocardiographic and haemodynamic parameters of cardiac function when compared with untreated TAC-operated mice. The apicidin derivative, API-D, is capable of reducing hypertrophy and, consequently, the transition to heart failure in mice subjected to TAC. Treatment with this substance, therefore, holds promise as an important therapeutic option for heart failure.

  13. Clinical use and applications of histone deacetylase inhibitors in multiple myeloma

    PubMed Central

    Tandon, Nidhi; Ramakrishnan, Vijay; Kumar, Shaji K

    2016-01-01

    The incorporation of various novel therapies has resulted in a significant survival benefit in newly diagnosed and relapsed patients with multiple myeloma (MM) over the past decade. Despite these advances, resistance to therapy leads to eventual relapse and fatal outcomes in the vast majority of patients. Hence, there is an unmet need for new safe and efficacious therapies for continued improvement in outcomes. Given the role of epigenetic aberrations in the pathogenesis and progression of MM and the success of histone deacetylase inhibitors (HDACi) in other malignancies, many HDACi have been tried in MM. Various preclinical studies helped us to understand the antimyeloma activity of different HDACi in MM as a single agent or in combination with conventional, novel, and immune therapies. The early clinical trials of HDACi depicted only modest single-agent activity, but recent studies have revealed encouraging clinical response rates in combination with other antimyeloma agents, especially proteasome inhibitors. This led to the approval of the combination of panobinostat and bortezomib for the treatment of relapsed/refractory MM patients with two prior lines of treatment by the US Food and Drug Administration. However, it remains yet to be defined how we can incorporate HDACi in the current therapeutic paradigms for MM that will help to achieve longer disease control and significant survival benefits. In addition, isoform-selective and/or class-selective HDAC inhibition to reduce unfavorable side effects needs further evaluation. PMID:27226735

  14. Histone deacetylase inhibitors mediate DNA damage repair in ameliorating hemorrhagic cystitis

    PubMed Central

    Haldar, Subhash; Dru, Christopher; Mishra, Rajeev; Tripathi, Manisha; Duong, Frank; Angara, Bryan; Fernandez, Ana; Arditi, Moshe; Bhowmick, Neil A.

    2016-01-01

    Hemorrhagic cystitis is an inflammatory and ulcerative bladder condition associated with systemic chemotherapeutics, like cyclophosphomide. Earlier, we reported reactive oxygen species resulting from cyclophosphamide metabolite, acrolein, causes global methylation followed by silencing of DNA damage repair genes. Ogg1 (8-oxoguanine DNA glycosylase) is one such silenced base excision repair enzyme that can restore DNA integrity. The accumulation of DNA damage results in subsequent inflammation associated with pyroptotic death of bladder smooth muscle cells. We hypothesized that reversing inflammasome-induced imprinting in the bladder smooth muscle could prevent the inflammatory phenotype. Elevated recruitment of Dnmt1 and Dnmt3b to the Ogg1 promoter in acrolein treated bladder muscle cells was validated by the pattern of CpG methylation revealed by bisulfite sequencing. Knockout of Ogg1 in detrusor cells resulted in accumulation of reactive oxygen mediated 8-Oxo-dG and spontaneous pyroptotic signaling. Histone deacetylase (HDAC) inhibitor, suberoylanilide hydroxamic acid (SAHA), restored Ogg1 expression in cells treated with acrolein and mice treated with cyclophosphamide superior to the standard of care, mesna or nicotinamide-induced DNA demethylation. SAHA restored cyclophosphamide-induced bladder pathology to that of untreated control mice. The observed epigenetic imprinting induced by inflammation suggests a new therapeutic target for the treatment of hemorrhagic cystitis. PMID:27995963

  15. Fasting and high-fat diet alter histone deacetylase expression in the medial hypothalamus.

    PubMed

    Funato, Hiromasa; Oda, Satoko; Yokofujita, Junko; Igarashi, Hiroaki; Kuroda, Masaru

    2011-04-15

    Increasing attention is now being given to the epigenetic regulation of animal and human behaviors including the stress response and drug addiction. Epigenetic factors also influence feeding behavior and metabolic phenotypes, such as obesity and insulin sensitivity. In response to fasting and high-fat diets, the medial hypothalamus changes the expression of neuropeptides regulating feeding, metabolism, and reproductive behaviors. Histone deacetylases (HDACs) are involved in the epigenetic control of gene expression and alter behavior in response to a variety of environmental factors. Here, we examined the expression of HDAC family members in the medial hypothalamus of mice in response to either fasting or a high-fat diet. In response to fasting, HDAC3 and -4 expression levels increased while HDAC10 and -11 levels decreased. Four weeks on a high-fat diet resulted in the increased expression of HDAC5 and -8. Moreover, fasting decreased the number of acetylated histone H3- and acetylated histone H4-positive cells in the ventrolateral subdivision of the ventromedial hypothalamus. Therefore, HDACs may be implicated in altered gene expression profiles in the medial hypothalamus under different metabolic states.

  16. Histone deacetylase inhibitors impair innate immune responses to Toll-like receptor agonists and to infection.

    PubMed

    Roger, Thierry; Lugrin, Jérôme; Le Roy, Didier; Goy, Geneviève; Mombelli, Matteo; Koessler, Thibaud; Ding, Xavier C; Chanson, Anne-Laure; Reymond, Marlies Knaup; Miconnet, Isabelle; Schrenzel, Jacques; François, Patrice; Calandra, Thierry

    2011-01-27

    Regulated by histone acetyltransferases and deacetylases (HDACs), histone acetylation is a key epigenetic mechanism controlling chromatin structure, DNA accessibility, and gene expression. HDAC inhibitors induce growth arrest, differentiation, and apoptosis of tumor cells and are used as anticancer agents. Here we describe the effects of HDAC inhibitors on microbial sensing by macrophages and dendritic cells in vitro and host defenses against infection in vivo. HDAC inhibitors down-regulated the expression of numerous host defense genes, including pattern recognition receptors, kinases, transcription regulators, cytokines, chemokines, growth factors, and costimulatory molecules as assessed by genome-wide microarray analyses or innate immune responses of macrophages and dendritic cells stimulated with Toll-like receptor agonists. HDAC inhibitors induced the expression of Mi-2β and enhanced the DNA-binding activity of the Mi-2/NuRD complex that acts as a transcriptional repressor of macrophage cytokine production. In vivo, HDAC inhibitors increased the susceptibility to bacterial and fungal infections but conferred protection against toxic and septic shock. Thus, these data identify an essential role for HDAC inhibitors in the regulation of the expression of innate immune genes and host defenses against microbial pathogens.

  17. Histone Deacetylase 3 Depletion in Osteo/Chondroprogenitor Cells Decreases Bone Density and Increases Marrow Fat

    PubMed Central

    Casper, Michelle E.; McGee-Lawrence, Meghan E.; Stensgard, Bridget A.; Li, Xiaodong; Secreto, Frank J.; Knutson, Sarah K.; Hiebert, Scott W.; Westendorf, Jennifer J.

    2010-01-01

    Histone deacetylase (Hdac)3 is a nuclear enzyme that contributes to epigenetic programming and is required for embryonic development. To determine the role of Hdac3 in bone formation, we crossed mice harboring loxP sites around exon 7 of Hdac3 with mice expressing Cre recombinase under the control of the osterix promoter. The resulting Hdac3 conditional knockout (CKO) mice were runted and had severe deficits in intramembranous and endochondral bone formation. Calvarial bones were significantly thinner and trabecular bone volume in the distal femur was decreased 75% in the Hdac3 CKO mice due to a substantial reduction in trabecular number. Hdac3-CKO mice had fewer osteoblasts and more bone marrow adipocytes as a proportion of tissue area than their wildtype or heterozygous littermates. Bone formation rates were depressed in both the cortical and trabecular regions of Hdac3 CKO femurs. Microarray analyses revealed that numerous developmental signaling pathways were affected by Hdac3-deficiency. Thus, Hdac3 depletion in osterix-expressing progenitor cells interferes with bone formation and promotes bone marrow adipocyte differentiation. These results demonstrate that Hdac3 inhibition is detrimental to skeletal health. PMID:20628553

  18. Low Dose Histone Deacetylase Inhibitor, Depsipeptide (FR901228), Promotes Adenoviral Transduction in Human Rhabdomyosarcoma Cell Lines.

    PubMed

    Navid, Fariba; Mischen, Blaine T; Helman, Lee J

    2004-01-01

    Purpose. Transduction of rhabdomyosarcoma (RMS) cells with adenoviral vectors for in vivo and in vitro applications has been limited by the low to absent levels of coxackie and adenovirus receptor (CAR). This study investigates the potential use of low doses of a histone deacetylase inhibitor, depsipeptide (FR901228), currently in Phase II human trials, to enhance adenoviral uptake in six rhabdomyosarcoma cell lines.Methods. Differences in adenoviral uptake in the presence and absence of depsipeptide (FR901228) were assessed using an adenoviral construct tagged with green fluorescent protein. Changes in CAR and alpha(v) integrin expression RMS in response to pretreatment with depsipeptide (FR901128) was determined using RT-PCR.Results. Pretreatment of five of six RMS cell lines with 0.5 ng/ml of depsipeptide (FR901228) for 72 h resulted in increased viral uptake as assessed by green fluorescent protein expression. RT-PCR analysis for CAR showed that in four of these five cell lines, CAR expression was increased 2.8-8.1-fold in cells treated with depsipeptide (FR901228) as compared to control. alpha(v) integrin expression was substantially increased in the one cell line, RH5, which showed increased GFP expression in response to depsipeptide (FR901228) pretreatment but a minimal increase in CAR expression.Conclusions. Depsipeptide (FR901228) can be used as a vehicle to enhance adenoviral transduction in a majority of RMS cells. The mechanism of increased viral uptake appears to mediate via upregulation of CAR.

  19. Low Dose Histone Deacetylase Inhibitor, Depsipeptide (FR901228), Promotes Adenoviral Transduction in Human Rhabdomyosarcoma Cell Lines

    PubMed Central

    Mischen, Blaine T.; Helman, Lee J.

    2004-01-01

    Purpose. Transduction of rhabdomyosarcoma (RMS) cells with adenoviral vectors for in vivo and in vitro applications has been limited by the low to absent levels of coxackie and adenovirus receptor (CAR). This study investigates the potential use of low doses of a histone deacetylase inhibitor, depsipeptide (FR901228), currently in Phase II human trials, to enhance adenoviral uptake in six rhabdomyosarcoma cell lines. Methods. Differences in adenoviral uptake in the presence and absence of depsipeptide (FR901228) were assessed using an adenoviral construct tagged with green fluorescent protein. Changes in CAR and αv integrin expression RMS in response to pretreatment with depsipeptide (FR901128) was determined using RT-PCR. Results. Pretreatment of five of six RMS cell lines with 0.5 ng/ml of depsipeptide (FR901228) for 72 h resulted in increased viral uptake as assessed by green fluorescent protein expression. RT-PCR analysis for CAR showed that in four of these five cell lines, CAR expression was increased 2.8–8.1-fold in cells treated with depsipeptide (FR901228) as compared to control. αv integrin expression was substantially increased in the one cell line, RH5, which showed increased GFP expression in response to depsipeptide (FR901228) pretreatment but a minimal increase in CAR expression. Conclusions. Depsipeptide (FR901228) can be used as a vehicle to enhance adenoviral transduction in a majority of RMS cells. The mechanism of increased viral uptake appears to mediate via upregulation of CAR. PMID:18521390

  20. ATM modulates transcription in response to histone deacetylase inhibition as part of its DNA damage response.

    PubMed

    Jang, Eun Ryoung; Choi, Jae Duk; Park, Mi Ae; Jeong, Gajin; Cho, Hyeseong; Lee, Jong-Soo

    2010-03-31

    Chromatin structure has a crucial role in a diversity of physiological processes, including development, differentiation and stress responses, via regulation of transcription, DNA replication and DNA damage repair. Histone deacetylase (HDAC) inhibitors regulate chromatin structure and activate the DNA damage checkpoint pathway involving Ataxia-telangiectasia mutated (ATM). Herein, we investigated the impact of histone acetylation/deacetylation modification on the ATM-mediated transcriptional modulation to provide a better understanding of the transcriptional function of ATM. The prototype HDAC inhibitor trichostain A (TSA) reprograms expression of the myeloid cell leukemia-1 (MCL1) and Gadd45 genes via the ATM-mediated signal pathway. Transcription of MCL1 and Gadd45alpha is enhanced following TSA treatment in ATM(+) cells, but not in isogenic ATM(-) or kinase-dead ATM expressing cells, in the ATM-activated E2F1 or BRCA1- dependent manner, respectively. These findings suggest that ATM and its kinase activity are essential for the TSA-induced regulation of gene expression. In summary, ATM controls the transcriptional upregulation of MCL1 and Gadd45 through the activation of the ATM-mediated signal pathway in response to HDAC inhibition. These findings are important in helping to design combinatory treatment schedules for anticancer radio- or chemo-therapy with HDAC inhibitors.

  1. A Novel Continuous Assay for the Deacylase Sirtuin 5 and Other Deacetylases.

    PubMed

    Roessler, Claudia; Tüting, Christian; Meleshin, Marat; Steegborn, Clemens; Schutkowski, Mike

    2015-09-24

    Sirtuins are NAD(+) dependent lysine deacylases involved in many regulatory processes like control of metabolic pathways, DNA repair, and stress response. Modulators of sirtuin activity are needed as tools for uncovering the biological function of these enzymes and as potential therapeutics. Systematic discovery of such modulators is hampered by the lack of efficient and simple continuous activity assays running at low sirtuin concentrations in microtiter plates. Here we describe an improved continuous sirtuin 5 assay based on the coupling of the sirtuin reaction to a proteolytic cleavage using internally fluorescence-quenched substrates. Systematic optimization of a carbamoyl phosphate synthetase 1 derived, glutarylated peptide yielded a Sirt5 substrate with k(cat)/K(M) value of 337,000 M(-1) s(-1), which represents the best sirtuin substrate described so far. These extraordinary substrate properties allowed reliable determination of Ki values for different inhibitors in the presence of only 10 nM sirtuin in microtiter plate format. Assay conditions could be transferred effectively to other lysine deacetylases, like sirtuin 2 and sirtuin 3, which now enables more efficient development of sirtuin targeting drugs.

  2. The Histone Deacetylase Gene Rpd3 Is Required for Starvation Stress Resistance

    PubMed Central

    Nakajima, Ei; Shimaji, Kouhei; Umegawachi, Takanari; Tomida, Saki; Yoshida, Hideki; Yoshimoto, Nana; Izawa, Shingo; Kimura, Hiroshi; Yamaguchi, Masamitsu

    2016-01-01

    Epigenetic regulation in starvation is important but not fully understood yet. Here we identified the Rpd3 gene, a Drosophila homolog of histone deacetylase 1, as a critical epigenetic regulator for acquiring starvation stress resistance. Immunostaining analyses of Drosophila fat body revealed that the subcellular localization and levels of Rpd3 dynamically changed responding to starvation stress. In response to starvation stress, the level of Rpd3 rapidly increased, and it accumulated in the nucleolus in what appeared to be foci. These observations suggest that Rpd3 plays a role in regulation of rRNA synthesis in the nucleolus. The RT-qPCR and ChIP-qPCR analyses clarified that Rpd3 binds to the genomic region containing the rRNA promoters and activates rRNA synthesis in response to starvation stress. Polysome analyses revealed that the amount of polysomes was decreased in Rpd3 knockdown flies under starvation stress compared with the control flies. Since the autophagy-related proteins are known to be starvation stress tolerance proteins, we examined autophagy activity, and it was reduced in Rpd3 knockdown flies. Taken together, we conclude that Rpd3 accumulates in the nucleolus in the early stage of starvation, upregulates rRNA synthesis, maintains the polysome amount for translation, and finally increases stress tolerance proteins, such as autophagy-related proteins, to acquire starvation stress resistance. PMID:27907135

  3. Inhibition of histone deacetylase activity increases chromosomal instability by the aberrant regulation of mitotic checkpoint activation.

    PubMed

    Shin, Hyun-Jin; Baek, Kwan-Hyuck; Jeon, Ae-Hwa; Kim, So-Jung; Jang, Kyung-Lib; Sung, Young-Chul; Kim, Chang-Min; Lee, Chang-Woo

    2003-06-19

    Histone modification through acetylation and deacetylation is a key process in transcription, DNA replication, and chromosome segregation. During mitosis, histones are highly acetylated and chromatin is condensed. Here, we investigate the mechanistic involvement of histone deacetylase (HDAC) activity in the regulation of mitotic checkpoint activation. Inhibition of HDAC activity was found to cause the improper kinetochore localization of the mitotic checkpoint proteins, and to prolong mitotic arrest, and thus to lead to chromosomal instability due to aberrant exit from the mitotic cell cycle arrest. In addition, treatment with HDAC inhibitor attenuated the activations of p38 and ERK kinases, and increased the expression levels of cIAP-1, suggesting that the observed increased adaptation and chromosomal instability induced by inhibiting HDAC activity might be directly connected with the activations of cell survival and/or antiapoptotic signals. Moreover, the treatment of cells with mitotic defects with HDAC inhibitor increased their susceptibility to chromosomal instability. These results support the notion that HDAC activity plays an important role in the regulation of mitotic checkpoint activation, and thus the aberrant control of HDAC activity contributes to chromosomal instability.

  4. Boric acid inhibits embryonic histone deacetylases: a suggested mechanism to explain boric acid-related teratogenicity.

    PubMed

    Di Renzo, Francesca; Cappelletti, Graziella; Broccia, Maria L; Giavini, Erminio; Menegola, Elena

    2007-04-15

    Histone deacetylases (HDAC) control gene expression by changing histonic as well as non histonic protein conformation. HDAC inhibitors (HDACi) are considered to be among the most promising drugs for epigenetic treatment for cancer. Recently a strict relationship between histone hyperacetylation in specific tissues of mouse embryos exposed to two HDACi (valproic acid and trichostatin A) and specific axial skeleton malformations has been demonstrated. The aim of this study is to verify if boric acid (BA), that induces in rodents malformations similar to those valproic acid and trichostatin A-related, acts through similar mechanisms: HDAC inhibition and histone hyperacetylation. Pregnant mice were treated intraperitoneally with a teratogenic dose of BA (1000 mg/kg, day 8 of gestation). Western blot analysis and immunostaining were performed with anti hyperacetylated histone 4 (H4) antibody on embryos explanted 1, 3 or 4 h after treatment and revealed H4 hyperacetylation at the level of somites. HDAC enzyme assay was performed on embryonic nuclear extracts. A significant HDAC inhibition activity (compatible with a mixed type partial inhibition mechanism) was evident with BA. Kinetic analyses indicate that BA modifies substrate affinity by a factor alpha=0.51 and maximum velocity by a factor beta=0.70. This work provides the first evidence for HDAC inhibition by BA and suggests such a molecular mechanism for the induction of BA-related malformations.

  5. Histone deacetylase 3 associates with MeCP2 to regulate FOXO and social behavior

    PubMed Central

    Nott, Alexi; Cheng, Jemmie; Gao, Fan; Lin, Yuan-Ta; Gjoneska, Elizabeta; Ko, Tak; Minhas, Paras; Zamudio, Alicia Viridiana; Meng, Jia; Zhang, Feiran; Jin, Peng; Tsai, Li-Huei

    2016-01-01

    Mutations in MECP2 cause the neurodevelopmental disorder Rett syndrome (RTT). The RTT missense MECP2R306C mutation prevents MeCP2 interaction with NCoR/Histone deacetylase 3 (HDAC3); however, the neuronal function of HDAC3 is incompletely understood. We report that neuronal deletion of Hdac3 in mice elicits abnormal locomotor coordination, sociability, and cognition. Transcriptional and chromatin profiling revealed HDAC3 positively regulates a subset of genes and is recruited to active gene promoters via MeCP2. HDAC3-associated promoters are enriched for the FOXO transcription factors, and FOXO acetylation is elevated in Hdac3 KO and Mecp2 KO neurons. Human RTT patient-derived MECP2R306C neural progenitor cells have deficits in HDAC3 and FOXO recruitment and gene expression. Gene editing of MECP2R306C cells to generate isogenic controls rescued HDAC3-FOXO-mediated impairments in gene expression. Our data suggests that HDAC3 interaction with MeCP2 positively regulates a subset of neuronal genes through FOXO deacetylation, and disruption of HDAC3 contributes to cognitive and social impairment. PMID:27428650

  6. Fasting and High-Fat Diet Alter Histone Deacetylase Expression in the Medial Hypothalamus

    PubMed Central

    Funato, Hiromasa; Oda, Satoko; Yokofujita, Junko; Igarashi, Hiroaki; Kuroda, Masaru

    2011-01-01

    Increasing attention is now being given to the epigenetic regulation of animal and human behaviors including the stress response and drug addiction. Epigenetic factors also influence feeding behavior and metabolic phenotypes, such as obesity and insulin sensitivity. In response to fasting and high-fat diets, the medial hypothalamus changes the expression of neuropeptides regulating feeding, metabolism, and reproductive behaviors. Histone deacetylases (HDACs) are involved in the epigenetic control of gene expression and alter behavior in response to a variety of environmental factors. Here, we examined the expression of HDAC family members in the medial hypothalamus of mice in response to either fasting or a high-fat diet. In response to fasting, HDAC3 and −4 expression levels increased while HDAC10 and −11 levels decreased. Four weeks on a high-fat diet resulted in the increased expression of HDAC5 and −8. Moreover, fasting decreased the number of acetylated histone H3- and acetylated histone H4-positive cells in the ventrolateral subdivision of the ventromedial hypothalamus. Therefore, HDACs may be implicated in altered gene expression profiles in the medial hypothalamus under different metabolic states. PMID:21526203

  7. Improving allogeneic islet transplantation by suppressing Th17 and enhancing Treg with histone deacetylase inhibitors.

    PubMed

    Sugimoto, Koji; Itoh, Takeshi; Takita, Morihito; Shimoda, Masayuki; Chujo, Daisuke; SoRelle, Jeff A; Naziruddin, Bashoo; Levy, Marlon F; Shimada, Mitsuo; Matsumoto, Shinichi

    2014-04-01

    Islet transplantation is a new treatment for achieving insulin independence for patients with severe diabetes. However, major drawbacks of this treatment are the long graft survival, the necessity for immunosuppressive drugs, and the efficacy of transplantation. Donor-specific transfusion (DST) has been shown to reduce rejection after organ transplantation, potentially through enhanced regulatory T-cell (Treg) activity. However, recent findings have shown that activated Treg can be converted into Th17 cells. We focused on histone deacetylase inhibitors (HDACi) because it was reported that inhibition of HDAC activity prevented Treg differentiation into IL17-producing cells. We therefore sought to enhance Treg while suppressing Th17 cells using DST with HDACi to prolong graft survival. To stimulate Treg by DST, we used donor splenocytes. In DST with HDACi group, Foxp3 mRNA expression and Treg population increased in the thymus and spleen, whereas Th17 population decreased. qPCR analysis of lymphocyte mRNA indicated that Foxp3, IL-10, and TGF-b expression increased. However, interleukin 17a, Stat3 (Th17), and IFN-g expression decreased in DST + HDACi group, relative to DST alone. Moreover, DST treated with HDACi prolonged graft survival relative to controls in mice islet transplantation. DST with HDACi may therefore have utility in islet transplantation.

  8. Histone deacetylase inhibitors mediate DNA damage repair in ameliorating hemorrhagic cystitis.

    PubMed

    Haldar, Subhash; Dru, Christopher; Mishra, Rajeev; Tripathi, Manisha; Duong, Frank; Angara, Bryan; Fernandez, Ana; Arditi, Moshe; Bhowmick, Neil A

    2016-12-20

    Hemorrhagic cystitis is an inflammatory and ulcerative bladder condition associated with systemic chemotherapeutics, like cyclophosphomide. Earlier, we reported reactive oxygen species resulting from cyclophosphamide metabolite, acrolein, causes global methylation followed by silencing of DNA damage repair genes. Ogg1 (8-oxoguanine DNA glycosylase) is one such silenced base excision repair enzyme that can restore DNA integrity. The accumulation of DNA damage results in subsequent inflammation associated with pyroptotic death of bladder smooth muscle cells. We hypothesized that reversing inflammasome-induced imprinting in the bladder smooth muscle could prevent the inflammatory phenotype. Elevated recruitment of Dnmt1 and Dnmt3b to the Ogg1 promoter in acrolein treated bladder muscle cells was validated by the pattern of CpG methylation revealed by bisulfite sequencing. Knockout of Ogg1 in detrusor cells resulted in accumulation of reactive oxygen mediated 8-Oxo-dG and spontaneous pyroptotic signaling. Histone deacetylase (HDAC) inhibitor, suberoylanilide hydroxamic acid (SAHA), restored Ogg1 expression in cells treated with acrolein and mice treated with cyclophosphamide superior to the standard of care, mesna or nicotinamide-induced DNA demethylation. SAHA restored cyclophosphamide-induced bladder pathology to that of untreated control mice. The observed epigenetic imprinting induced by inflammation suggests a new therapeutic target for the treatment of hemorrhagic cystitis.

  9. Regulation of androgen receptor and histone deacetylase 1 by Mdm2-mediated ubiquitylation.

    PubMed

    Gaughan, Luke; Logan, Ian R; Neal, David E; Robson, Craig N

    2005-01-01

    The androgen receptor (AR) is a member of the nuclear hormone receptor family of transcription factors and plays a critical role in regulating the expression of genes involved in androgen-dependent and -independent tumour formation. Regulation of the AR is achieved by alternate binding of either histone acetyltransferase (HAT)-containing co-activator proteins, or histone deacetylase 1 (HDAC1). Factors that control AR stability may also constitute an important regulatory mechanism, a notion that has been confirmed with the finding that the AR is a direct target for Mdm2-mediated ubiquitylation and proteolysis. Using chromatin immunoprecipitation (ChIP) and re-ChIP analyses, we show that Mdm2 associates with AR and HDAC1 at the active androgen-responsive PSA promoter in LNCaP prostate cancer cells. Furthermore, we demonstrate that Mdm2-mediated modification of AR and HDAC1 catalyses protein destabilization and attenuates AR sactivity, suggesting that ubiquitylation of the AR and HDAC1 may constitute an additional mechanism for regulating AR function. We also show that HDAC1 and Mdm2 function co-operatively to reduce AR-mediated transcription that is attenuated by the HAT activity of the AR co-activator Tip60, suggesting interplay between acetylation status and receptor ubiquitylation in AR regulation. In all, our data indicates a novel role for Mdm2 in regulating components of the AR transcriptosome.

  10. MicroRNA-381 Regulates Chondrocyte Hypertrophy by Inhibiting Histone Deacetylase 4 Expression.

    PubMed

    Chen, Weishen; Sheng, Puyi; Huang, Zhiyu; Meng, Fangang; Kang, Yan; Huang, Guangxin; Zhang, Zhiqi; Liao, Weiming; Zhang, Ziji

    2016-08-23

    Chondrocyte hypertrophy, regulated by Runt-related transcription factor 2 (RUNX2) and matrix metalloproteinase 13 (MMP13), is a crucial step in cartilage degeneration and osteoarthritis (OA) pathogenesis. We previously demonstrated that microRNA-381 (miR-381) promotes MMP13 expression during chondrogenesis and contributes to cartilage degeneration; however, the mechanism underlying this process remained unclear. In this study, we observed divergent expression of miR-381 and histone deacetylase 4 (HDAC4), an enzyme that directly inhibits RUNX2 and MMP13 expression, during late-stage chondrogenesis of ATDC5 cells, as well as in prehypertrophic and hypertrophic chondrocytes during long bone development in E16.5 mouse embryos. We therefore investigated whether this miRNA regulates HDAC4 expression during chondrogenesis. Notably, overexpression of miR-381 inhibited HDAC4 expression but promoted RUNX2 expression. Moreover, transfection of SW1353 cells with an miR-381 mimic suppressed the activity of a reporter construct containing the 3'-untranslated region (3'-UTR) of HDAC4. Conversely, treatment with a miR-381 inhibitor yielded increased HDAC4 expression and decreased RUNX2 expression. Lastly, knockdown of HDAC4 expression resulted in increased RUNX2 and MMP13 expression in SW1353 cells. Collectively, our results indicate that miR-381 epigenetically regulates MMP13 and RUNX2 expression via targeting of HDAC4, thereby suggesting the possibilities of inhibiting miR-381 to control chondrocyte hypertrophy and cartilage degeneration.

  11. Identification of G1-Regulated Genes in Normally Cycling Human Cells

    PubMed Central

    Beyrouthy, Maroun J.; Alexander, Karen E.; Baldwin, Amy; Whitfield, Michael L.; Bass, Hank W.; McGee, Dan; Hurt, Myra M.

    2008-01-01

    Background Obtaining synchronous cell populations is essential for cell-cycle studies. Methods such as serum withdrawal or use of drugs which block cells at specific points in the cell cycle alter cellular events upon re-entry into the cell cycle. Regulatory events occurring in early G1 phase of a new cell cycle could have been overlooked. Methodology and Findings We used a robotic mitotic shake-off apparatus to select cells in late mitosis for genome-wide gene expression studies. Two separate microarray experiments were conducted, one which involved isolation of RNA hourly for several hours from synchronous cell populations, and one experiment which examined gene activity every 15 minutes from late telophase of mitosis into G1 phase. To verify synchrony of the cell populations under study, we utilized methods including BrdU uptake, FACS, and microarray analyses of histone gene activity. We also examined stress response gene activity. Our analysis enabled identification of 200 early G1-regulated genes, many of which currently have unknown functions. We also confirmed the expression of a set of genes candidates (fos, atf3 and tceb) by qPCR to further validate the newly identified genes. Conclusion and Significance Genome-scale expression analyses of the first two hours of G1 in naturally cycling cells enabled the discovery of a unique set of G1-regulated genes, many of which currently have unknown functions, in cells progressing normally through the cell division cycle. This group of genes may contain future targets for drug development and treatment of human disease. PMID:19079774

  12. Targeting GPR30 with G-1: a new therapeutic target for castration-resistant prostate cancer

    PubMed Central

    Lam, Hung-Ming; Ouyang, Bin; Chen, Jing; Ying, Jun; Wang, Jiang; Wu, Chin-Lee; Li, Jia; Medvedovic, Mario; Vessella, Robert L.; Ho, Shuk-Mei

    2014-01-01

    Castration-resistant prostate cancer (CRPC) is an advanced-stage prostate cancer (PC) associated with high mortality. We reported that G-1, a selective agonist of G protein-coupled receptor 30 (GPR30), inhibited PC cell growth by inducing G2 cell cycle arrest and arrested PC-3 xenograft growth. However, therapeutic actions of G-1 and their relationships with androgen in vivo are unclear. Using the LNCaP xenograft to model PC growth during androgen-sensitive (AS) versus castration-resistant (CR) phase, we found that G-1 inhibited growth of CR but not AS tumors with no observable toxicity to the host. Substantial necrosis (~65%) accompanied by marked intratumoral infiltration of neutrophils was observed only in CR tumors. Global transcriptome profiling of human genes identified 99 differential expressed genes with “interplay between innate and adaptive immune response” as the top pathway. Quantitative-PCR confirmed upregulation of neutrophil-related chemokines and inflammation-mediated cytokines only in the G-1-treated CR tumors. Expression of murine neutrophil-related cytokines also was elevated in these tumors. GPR30 expression was significantly higher in CR than AS tumors. In cell-based experiments, androgen repressed GPR30 expression, a response reversible by anti-androgen or siRNA-induced androgen receptor silencing. Finally, in clinical specimens, 80% of CRPC metastases (n=123) express a high level of GPR30 whereas only 54% of the primary PCs (n=232) showed high GPR30 expression. Together, these results provide the first evidence that GPR30 is an androgen-repressed target and G-1 mediates the anti-tumor effect via neutrophil infiltration-associated necrosis in CRPC. Additional studies are warranted to firmly establish GPR30 as a therapeutic target in CRPC. PMID:25287069

  13. Structural and functional characterization of a putative polysaccharide deacetylase of the human parasite Encephalitozoon cuniculi.

    PubMed

    Urch, Jonathan E; Hurtado-Guerrero, Ramon; Brosson, Damien; Liu, Zhanliang; Eijsink, Vincent G H; Texier, Catherine; van Aalten, Daan M F

    2009-06-01

    The microsporidian Encephalitozoon cuniculi is an intracellular eukaryotic parasite considered to be an emerging opportunistic human pathogen. The infectious stage of this parasite is a unicellular spore that is surrounded by a chitin containing endospore layer and an external proteinaceous exospore. A putative chitin deacetylase (ECU11_0510) localizes to the interface between the plasma membrane and the endospore. Chitin deacetylases are family 4 carbohydrate esterases in the CAZY classification, and several bacterial members of this family are involved in evading lysis by host glycosidases, through partial de-N-acetylation of cell wall peptidoglycan. Similarly, ECU11_0510 could be important for E. cuniculi survival in the host, by protecting the chitin layer from hydrolysis by human chitinases. Here, we describe the biochemical, structural, and glycan binding properties of the protein. Enzymatic analyses showed that the putative deacetylase is unable to deacetylate chitooligosaccharides or crystalline beta-chitin. Furthermore, carbohydrate microarray analysis revealed that the protein bound neither chitooligosaccharides nor any of a wide range of other glycans or chitin. The high resolution crystal structure revealed dramatic rearrangements in the positions of catalytic and substrate binding residues, which explain the loss of deacetylase activity, adding to the unusual structural plasticity observed in other members of this esterase family. Thus, it appears that the ECU11_0510 protein is not a carbohydrate deacetylase and may fulfill an as yet undiscovered role in the E. cuniculi parasite.

  14. G1-4A, a Polysaccharide from Tinospora cordifolia Inhibits the Survival of Mycobacterium tuberculosis by Modulating Host Immune Responses in TLR4 Dependent Manner

    PubMed Central

    Gupta, Pramod Kumar; Chakraborty, Pampi; Kumar, Santosh; Singh, Prafull Kumar; Rajan, M. G. R.; Sainis, Krishna B.; Kulkarni, Savita

    2016-01-01

    Rapid emergence of drug resistance in Mycobacterium tuberculosis (MTB) is a major health concern and demands the development of novel adjunct immunotherapeutic agents capable of modulating the host immune responses in order to control the pathogen. In the present study, we sought to investigate the immunomodulatory effects of G1-4A, a polysaccharide derived from the Indian medicinal plant Tinospora cordifolia, in in-vitro and aerosol mouse models of MTB infection. G1-4A treatment of MTB infected RAW264.7 macrophages significantly induced the surface expression of MHC-II and CD-86 molecules, secretion of proinflammatory cytokines (TNF-α, IL-β, IL-6, IL-12, IFN-γ) and nitric oxide leading to reduced intracellular survival of both drug sensitive (H37Rv) as well as multi drug resistant strains (Beijing and LAM) of MTB, which was partially attributed to G1-4A induced NO production in TLR4-MyD88 dependent manner. Similarly, bacillary burden was significantly reduced in the lungs of MTB infected BALB/c mice treated with G1-4A, with simultaneous up-regulation of the expression of TNF-α, INF-γ and NOS2 in the mouse lung along with increased levels of Th1 cytokines like IFN-γ, IL-12 and decreased levels of Th2 cytokine like IL-4 in the serum. Furthermore, combination of G1-4A with Isoniazid (INH) exhibited better protection against MTB compared to that due to INH or G1-4A alone, suggesting its potential as adjunct therapy. Our results demonstrate that modulation of host immune responses by G1-4A might improve the therapeutic efficacy of existing anti-tubercular drugs and provide an attractive strategy for the development of alternative therapies to control tuberculosis. PMID:27148868

  15. Histone deacetylases: unique players in shaping the epigenetic histone code.

    PubMed

    Thiagalingam, Sam; Cheng, Kuang-Hung; Lee, Hyunjoo J; Mineva, Nora; Thiagalingam, Arunthathi; Ponte, Jose F

    2003-03-01

    The epigenome is defined by DNA methylation patterns and the associated posttranslational modifications of histones. This histone code determines the expression status of individual genes dependent upon their localization on the chromatin. The silencing of gene expression is associated with deacetylated histones, which are often found to be associated with regions of DNA methylation as well as methylation at the lysine 4 residue of histone 3. In contrast, the activation of gene expression is associated with acetylated histones and methylation at the lysine 9 residue of histone 3. The histone deactylases play a major role in keeping the balance between the acetylated and deacetylated states of chromatin. Histone deacetylases (HDACs) are divided into three classes: class I HDACs (HDACs 1, 2, 3, and 8) are similar to the yeast RPD3 protein and localize to the nucleus; class II HDACs (HDACs 4, 5, 6, 7, 9, and 10) are homologous to the yeast HDA1 protein and are found in both the nucleus and cytoplasm; and class III HDACs form a structurally distinct class of NAD-dependent enzymes that are similar to the yeast SIR2 proteins. Since inappropriate silencing of critical genes can result in one or both hits of tumor suppressor gene (TSG) inactivation in cancer, theoretically the reactivation of affected TSGs could have an enormous therapeutic value in preventing and treating cancer. Indeed, several HDAC inhibitors are currently being developed and tested for their potency in cancer chemotherapy. Importantly, these agents are also potentially applicable to chemoprevention if their toxicity can be minimized. Despite the toxic side effects and lack of specificity of some of the inhibitors, progress is being made. With the elucidation of the structures, functions and modes of action of HDACs, finding agents that may be targeted to specific HDACs and potentially reactivate expression of only a defined set of affected genes in cancer will be more attainable.

  16. Inhibitors of Histone Deacetylases Enhance Neurotoxicity of DNA Damage

    PubMed Central

    Vashishta, A.

    2014-01-01

    The nonselective inhibitors of class I/II histone deacetylases (HDACs) including trichostatin A and the clinically used suberoylanilide hydroxamic acid (SAHA, vorinostat) are neuroprotective in several models of neuronal injury. Here, we report that in cultured cortical neurons from newborn rats and in the cerebral cortex of whole neonate rats, these HDAC inhibitors exacerbated cytotoxicity of the DNA double-strand break (DSB)-inducing anticancer drug etoposide by enhancing apoptosis. Similar neurotoxic interactions were also observed in neurons that were treated with other DNA damaging drugs including cisplatin and camptothecin. In addition, in rat neonates, SAHA increased cortical neuron apoptosis that was induced by a single injection of the NMDA receptor antagonist dizocilpine (MK801). In etoposide-treated neurons, the nonselective HDAC inhibition resulted in more DSBs. It also potentiated etoposide-induced accumulation and phosphorylation of the pro-apoptotic transcription factor p53. Moreover, nonselective HDAC inhibition exacerbated neuronal apoptosis that was induced by the overexpressed p53. Importantly, such effects cannot be fully explained by inhibition of HDAC1, which is known to play a role in DSB repair and regulation of p53. The specific HDAC1 inhibitor MS275 only moderately enhanced etoposide-induced neuronal death. Although in etoposide-treated neurons MS275 increased DSBs, it did not affect activation of p53. Our findings suggest that besides HDAC1, there are other class I/II HDACs that participate in neuronal DNA damage response attenuating neurotoxic consequences of genotoxic insults to the developing brain. PMID:25063076

  17. Human arylacetamide deacetylase is a principal enzyme in flutamide hydrolysis.

    PubMed

    Watanabe, Akinobu; Fukami, Tatsuki; Nakajima, Miki; Takamiya, Masataka; Aoki, Yasuhiro; Yokoi, Tsuyoshi

    2009-07-01

    Flutamide, an antiandrogen drug, is widely used for the treatment of prostate cancer. The initial metabolic pathways of flutamide are hydroxylation and hydrolysis. It was recently reported that the hydrolyzed product, 4-nitro-3-(trifluoromethyl)phenylamine (FLU-1), is further metabolized to N-hydroxy FLU-1, an assumed hepatotoxicant. However, the esterase responsible for the flutamide hydrolysis has not been characterized. In the present study, we found that human arylacetamide deacetylase (AADAC) efficiently hydrolyzed flutamide using recombinant AADAC expressed in COS7 cells. In contrast, carboxylesterase1 (CES1) and CES2, which are responsible for the hydrolysis of many drugs, could not hydrolyze flutamide. AADAC is specifically expressed in the endoplasmic reticulum. Flutamide hydrolase activity was highly detected in human liver microsomes (K(m), 794 +/- 83 microM; V(max), 1.1 +/- 0.0 nmol/min/mg protein), whereas the activity was extremely low in human liver cytosol. The flutamide hydrolase activity in human liver microsomes was strongly inhibited by bis-(p-nitrophenyl)phosphate [corrected], diisopropylphosphorofluoride, and physostigmine sulfate (eserine) but moderately inhibited by sodium fluoride, phenylmethylsulfonyl fluoride, and disulfiram. The same inhibition pattern was obtained with the recombinant AADAC. Moreover, human liver and jejunum microsomes showing AADAC expression could hydrolyze flutamide, but human pulmonary and renal microsomes, which do not express AADAC, showed slight activity. In human liver microsomal samples (n = 50), the flutamide hydrolase activities were significantly correlated with the expression levels of AADAC protein (r = 0.66, p < 0.001). In conclusion, these results clearly showed that flutamide is exclusively hydrolyzed by AADAC. AADAC would be an important enzyme responsible for flutamide-induced hepatotoxicity.

  18. Alcohol-induced serotonergic modulation: the role of histone deacetylases.

    PubMed

    Agudelo, Marisela; Yoo, Changwon; Nair, Madhavan P

    2012-11-01

    Previous studies have demonstrated that alcohol use disorders (AUDs) are regulated by multiple mechanisms such as neurotransmitters and enzymes. The neurotransmitter, serotonin (5-hydroxytryptamine, 5-HT) may contribute to alcohol effects and serotonin receptors, including 5-HT3, play an important role in AUDs. Recent studies have also implicated histone deacetylases (HDACs) and acetyltransferases (HATS) in regulation of drug addiction, and HDAC inhibitors (HDACi) have been reported as transcriptional modulators of monoaminergic neurotransmission. Therefore, we hypothesize that HDACs may play a role in ethanol-induced serotonergic modulation. The effects of ethanol on serotonin and 5-HT3, and the role HDACs, HDAC activity and the HDACi, trichostatin A (TSA), play in alcohol-induced serotonergic effects were studied. Human SK-N-MC and neurons, were treated with ethanol (0.05, 0.1 and 0.2%), and/or TSA (50 nM), and 5-HT3 levels were assessed at 24-72 h. Gene expression was evaluated by qRT-PCR and protein by western blot and flow cytometry. Serotonin release was assessed by ELISA and HDAC activity by fluorometric assay. Our results show an increase in 5-HT3 gene after ethanol treatment. Further, ethanol significantly increased HDACs 1 and 3 genes accompanied by an increased in HDAC activity while TSA significantly inhibited HDACs. Studies with TSA show a significant upregulation of ethanol effects on 5-HT3, while surprisingly TSA inhibited ethanol-induced serotonin production. These results suggest that ethanol affects 5-HT3 and serotonin through mechanisms involving HDACs and HATs. In summary, our studies demonstrate some of the novel properties of HDAC inhibitors and contribute to the understanding of the mechanisms involve in alcohol-serotonergic modulation in the CNS.

  19. Histone Deacetylases in Bone Development and Skeletal Disorders

    PubMed Central

    Bradley, Elizabeth W.; Carpio, Lomeli R.; van Wijnen, Andre J.; McGee-Lawrence, Meghan E.; Westendorf, Jennifer J.

    2015-01-01

    Histone deacetylases (Hdacs) are conserved enzymes that remove acetyl groups from lysine side chains in histones and other proteins. Eleven of the 18 Hdacs encoded by the human and mouse genomes depend on Zn2+ for enzymatic activity, while the other 7, the sirtuins (Sirts), require NAD2+. Collectively, Hdacs and Sirts regulate numerous cellular and mitochondrial processes including gene transcription, DNA repair, protein stability, cytoskeletal dynamics, and signaling pathways to affect both development and aging. Of clinical relevance, Hdacs inhibitors are United States Food and Drug Administration-approved cancer therapeutics and are candidate therapies for other common diseases including arthritis, diabetes, epilepsy, heart disease, HIV infection, neurodegeneration, and numerous aging-related disorders. Hdacs and Sirts influence skeletal development, maintenance of mineral density and bone strength by affecting intramembranous and endochondral ossification, as well as bone resorption. With few exceptions, inhibition of Hdac or Sirt activity though either loss-of-function mutations or prolonged chemical inhibition has negative and/or toxic effects on skeletal development and bone mineral density. Specifically, Hdac/Sirt suppression causes abnormalities in physiological development such as craniofacial dimorphisms, short stature, and bone fragility that are associated with several human syndromes or diseases. In contrast, activation of Sirts may protect the skeleton from aging and immobilization-related bone loss. This knowledge may prolong healthspan and prevent adverse events caused by epigenetic therapies that are entering the clinical realm at an unprecedented rate. In this review, we summarize the general properties of Hdacs/Sirts and the research that has revealed their essential functions in bone forming cells (e.g., osteoblasts and chondrocytes) and bone resorbing osteoclasts. Finally, we offer predictions on future research in this area and the utility of

  20. Identification and characterization of histone deacetylases in tomato (Solanum lycopersicum)

    PubMed Central

    Zhao, Linmao; Lu, Jingxia; Zhang, Jianxia; Wu, Pei-Ying; Yang, Songguang; Wu, Keqiang

    2015-01-01

    Histone acetylation and deacetylation at the N-terminus of histone tails play crucial roles in the regulation of eukaryotic gene activity. Histone acetylation and deacetylation are catalyzed by histone acetyltransferases and histone deacetylases (HDACs), respectively. A growing number of studies have demonstrated the importance of histone deacetylation/acetylation on genome stability, transcriptional regulation, development and response to stress in Arabidopsis. However, the biological functions of HDACs in tomato have not been investigated previously. Fifteen HDACs identified from tomato (Solanum lycopersicum) can be grouped into RPD3/HDA1, SIR2 and HD2 families based on phylogenetic analysis. Meanwhile, 10 members of the RPD3/HDA1 family can be further subdivided into four groups, namely Class I, Class II, Class III, and Class IV. High similarities of protein sequences and conserved domains were identified among SlHDACs and their homologs in Arabidopsis. Most SlHDACs were expressed in all tissues examined with different transcript abundance. Transient expression in Arabidopsis protoplasts showed that SlHDA8, SlHDA1, SlHDA5, SlSRT1 and members of the HD2 family were localized to the nucleus, whereas SlHDA3 and SlHDA4 were localized in both the cytoplasm and nucleus. The difference in the expression patterns and subcellular localization of SlHDACs suggest that they may play distinct functions in tomato. Furthermore, we found that three members of the RPD3/HDA1 family, SlHDA1, SIHDA3 and SlHDA4, interacted with TAG1 (TOMATO AGAMOUS1) and TM29 (TOMATO MADS BOX29), two MADS-box proteins associated with tomato reproductive development, indicating that these HDACs may be involved in gene regulation in reproductive development. PMID:25610445

  1. Histone Deacetylase 8: Characterization of Physiological Divalent Metal Catalysis.

    PubMed

    Nechay, Michael R; Gallup, Nathan M; Morgenstern, Amanda; Smith, Quentin A; Eberhart, Mark E; Alexandrova, Anastassia N

    2016-07-07

    Histone deacetylases (HDACs) are responsible for the removal of acetyl groups from histones, resulting in gene silencing. Overexpression of HDACs is associated with cancer, and their inhibitors are of particular interest as chemotherapeutics. However, HDACs remain a target of mechanistic debate. HDAC class 8 is the most studied HDAC, and of particular importance due to its human oncological relevance. HDAC8 has traditionally been considered to be a Zn-dependent enzyme. However, recent experimental assays have challenged this assumption and shown that HDAC8 is catalytically active with a variety of different metals, and that it may be a Fe-dependent enzyme in vivo. We studied two opposing mechanisms utilizing a series of divalent metal ions in physiological abundance (Zn(2+), Fe(2+), Co(2+), Mn(2+), Ni(2+), and Mg(2+)). Extensive sampling of the entire protein with different bound metals was done with the mixed quantum-classical QM/DMD method. Density functional theory (DFT) on an unusually large cluster model was used to describe the active site and reaction mechanism. We have found that the reaction profile of HDAC8 is similar among all metals tested, and follows one of the previously published mechanisms, but the rate-determining step is different from the one previously claimed. We further provide a scheme for estimating the metal binding affinities to the protein. We use the quantum theory of atoms in molecules (QTAIM) to understand the different binding affinities for each metal in HDAC8 as well as the ability of each metal to bind and properly orient the substrate for deacetylation. The combination of this data with the catalytic rate constants is required to reproduce the experimentally observed trend in metal-depending performance. We predict Co(2+) and Zn(2+) to be the most active metals in HDAC8, followed by Fe(2+), and Mn(2+) and Mg(2+) to be the least active.

  2. Activation and inhibition of histone deacetylase 8 by monovalent cations.

    PubMed

    Gantt, Stephanie L; Joseph, Caleb G; Fierke, Carol A

    2010-02-26

    The metal-dependent histone deacetylases (HDACs) catalyze hydrolysis of acetyl groups from acetyllysine side chains and are targets of cancer therapeutics. Two bound monovalent cations (MVCs) of unknown function have been previously observed in crystal structures of HDAC8; site 1 is near the active site, whereas site 2 is located > 20 A from the catalytic metal ion. Here we demonstrate that one bound MVC activates catalytic activity (K(1/2) = 3.4 mM for K(+)), whereas the second, weaker-binding MVC (K(1/2) = 26 mM for K(+)) decreases catalytic activity by 11-fold. The weaker binding MVC also enhances the affinity of the HDAC inhibitor suberoylanilide hydroxamic acid by 5-fold. The site 1 MVC is coordinated by the side chain of Asp-176 that also forms a hydrogen bond with His-142, one of two histidines important for catalytic activity. The D176A and H142A mutants each increase the K(1/2) for potassium inhibition by > or = 40-fold, demonstrating that the inhibitory cation binds to site 1. Furthermore, the MVC inhibition is mediated by His-142, suggesting that this residue is protonated for maximal HDAC8 activity. Therefore, His-142 functions either as an electrostatic catalyst or a general acid. The activating MVC binds in the distal site and causes a time-dependent increase in activity, suggesting that the site 2 MVC stabilizes an active conformation of the enzyme. Sodium binds more weakly to both sites and activates HDAC8 to a lesser extent than potassium. Therefore, it is likely that potassium is the predominant MVC bound to HDAC8 in vivo.

  3. Electrocardiographic effects of class 1 selective histone deacetylase inhibitor romidepsin

    PubMed Central

    Sager, Philip T; Balser, Barbara; Wolfson, Julie; Nichols, Jean; Pilot, Richard; Jones, Suzanne; Burris, Howard A

    2015-01-01

    Romidepsin is a histone deacetylase inhibitor approved by the FDA for the treatment of patients with cutaneous or peripheral T-cell lymphoma who have received prior systemic therapy. The objective of this analysis was to evaluate the potential QTc effects of romidepsin. Patients with advanced malignancy received 4-h infusions of 14 mg/m2 romidepsin on days 1, 8, and 15 of a 28-day cycle. In cycle 2, a subset of patients received 1-h infusions of 8–12 mg/m2 romidepsin. Patients were administered antiemetics before each romidepsin dose and electrolyte supplementation as needed. Electrocardiogram readings were performed prior to antiemetic administration, prior to romidepsin administration, and at specified time points over the subsequent 24 h. Romidepsin exposure and heart rate were also assessed. In the electrocardiogram-evaluable population, 26 patients received romidepsin at 14 mg/m2 over 4 h. The maximum mean increases from the preantiemetic baseline for QTcF and heart rate were 10.1 msec (upper 90% CI, 14.5 msec) and 18.2 beats per minute, respectively. No patient in this study had an absolute QTcF value >450 msec and only one patient had an increase from the preantiemetic baseline of >60 msec. There was a mild reduction in the PR interval and no meaningful changes in the QRS interval. Despite the use of QT-prolonging antiemetics, treatment with romidepsin did not markedly prolong the QTc interval through 24 h. Increases in calculated QTc may have been exaggerated as a consequence of transient increases in heart rate. PMID:25914207

  4. Electrocardiographic effects of class 1 selective histone deacetylase inhibitor romidepsin.

    PubMed

    Sager, Philip T; Balser, Barbara; Wolfson, Julie; Nichols, Jean; Pilot, Richard; Jones, Suzanne; Burris, Howard A

    2015-08-01

    Romidepsin is a histone deacetylase inhibitor approved by the FDA for the treatment of patients with cutaneous or peripheral T-cell lymphoma who have received prior systemic therapy. The objective of this analysis was to evaluate the potential QTc effects of romidepsin. Patients with advanced malignancy received 4-h infusions of 14 mg/m(2) romidepsin on days 1, 8, and 15 of a 28-day cycle. In cycle 2, a subset of patients received 1-h infusions of 8-12 mg/m(2) romidepsin. Patients were administered antiemetics before each romidepsin dose and electrolyte supplementation as needed. Electrocardiogram readings were performed prior to antiemetic administration, prior to romidepsin administration, and at specified time points over the subsequent 24 h. Romidepsin exposure and heart rate were also assessed. In the electrocardiogram-evaluable population, 26 patients received romidepsin at 14 mg/m(2) over 4 h. The maximum mean increases from the preantiemetic baseline for QTcF and heart rate were 10.1 msec (upper 90% CI, 14.5 msec) and 18.2 beats per minute, respectively. No patient in this study had an absolute QTcF value >450 msec and only one patient had an increase from the preantiemetic baseline of >60 msec. There was a mild reduction in the PR interval and no meaningful changes in the QRS interval. Despite the use of QT-prolonging antiemetics, treatment with romidepsin did not markedly prolong the QTc interval through 24 h. Increases in calculated QTc may have been exaggerated as a consequence of transient increases in heart rate. © 2015 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  5. Overexpression of a glycosyltransferase gene SrUGT74G1 from Stevia improved growth and yield of transgenic Arabidopsis by catechin accumulation.

    PubMed

    Guleria, Praveen; Yadav, Sudesh Kumar

    2014-03-01

    Steviol glycoside and gibberellin biosynthetic routes are known as divergent branches of a common origin in Stevia. A UDP-glycosyltransferase encoded by SrUGT74G1 catalyses the conversion of steviolbioside into stevioside in Stevia rebaudiana leaves. In the present study, transgenic Arabidopsis thaliana overexpressing SrUGT74G1 cDNA from Stevia were developed to check the probability of stevioside biosynthesis in them. However, stevioside accumulation was not evident in transgenics. Also, the transgenic Arabidopsis showed no change in GA3 content on SrUGT74G1 overexpression. Surprisingly, significant accumulation of catechin was noticed in transgenics. The transgenics showed a considerable increase in shoot length, root length and rosette area. An increase in free radical scavenging activity of transgenics was noticed. Moreover, the seed yield of transgenics was also increased by 6-15% than control. Additionally, variation in trichome branching pattern on leaf surface of transgenics was observed. The trichome branching pattern was also validated by exogenous catechin exposure (10, 50, 100 ng ml(-1)) to control plants. Hence, present study reports the probable role of SrUGT74G1 from Stevia in catechin accumulation of transgenic Arabidopsis thaliana. Thus, detailed study in present perspective has revealed the role of Stevia SrUGT74G1 gene in trichome branching pattern, improved vegetative growth, scavenging potential and seed yield by catechin accumulation in transgenic Arabidopsis.

  6. Skewed Fc glycosylation profiles of anti-proteinase 3 immunoglobulin G1 autoantibodies from granulomatosis with polyangiitis patients show low levels of bisection, galactosylation, and sialylation.

    PubMed

    Wuhrer, Manfred; Stavenhagen, Kathrin; Koeleman, Carolien A M; Selman, Maurice H J; Harper, Lorraine; Jacobs, Bart C; Savage, Caroline O S; Jefferis, Roy; Deelder, André M; Morgan, Matthew

    2015-04-03

    Granulomatosis with polyangiitis (GPA) is associated with circulating immunoglobulin (Ig) G anti-proteinase 3 specific (anti-PR3) anti-neutrophil cytoplasm antibodies (ANCA), which activate cytokine primed neutrophils via Fcgamma receptors. ANCA are class switched IgG antibodies implying T cell help in their production. Glycosylation of IgG Fc, under the control of T cell cytokines, determines the interaction between IgG and its receptors. Previous studies have reported aberrant glycosylation of Ig Fc in GPA patients. We investigated whether aberrant Fc glycosylation was present on anti-PR3 ANCA as well as whole IgG subclass preparations compared to healthy controls and whether this correlated with Birmingham vasculitis activity scores (BVAS), serum cytokines, and time to remission. Here, IgG Fc glycosylation of GPA patients and controls and anti-PR3 ANCA Fc glycosylation were determined by mass spectrometry of glycopeptides. IgG1 and IgG2 subclasses from GPA patients showed reduced galactosylation, sialylation, and bisection compared to healthy controls. Anti-PR3 IgG1 ANCA Fc galactosylation, sialylation, and bisection were reduced compared to total IgG1 in GPA. Galactosylation of anti-PR3 ANCA Fc correlated with inflammatory cytokines and time to remission but not BVAS. Bisection of anti-PR3 ANCA Fc correlated with BVAS. Total IgG1 and anti-PR3 IgG1 Fc galactosylation were weakly correlated, while bisection of IgG1 and anti-PR3 showed no correlation. Our data indicate that aberrant ANCA galactosylation may be driven in an antigen-specific manner.

  7. 26 CFR 6a.6652(g)-1 - Failure to make return or furnish statement required under section 6039C.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... required under section 6039C. 6a.6652(g)-1 Section 6a.6652(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... OMNIBUS RECONCILIATION ACT OF 1980 § 6a.6652(g)-1 Failure to make return or furnish statement required... limitation under § 6a.6652(g)-1(b)(3) with respect to failure to meet the requirements of section 6039C(c), U...

  8. Deacetylase inhibitors dissociate the histone-targeting ING2 subunit from the Sin3 complex

    PubMed Central

    Smith, Karen T.; Martin-Brown, Skylar A.; Florens, Laurence; Washburn, Michael P.; Workman, Jerry L.

    2010-01-01

    Summary Histone deacetylase (HDAC) inhibitors are in clinical development for several diseases, including cancers and neurodegenerative disorders. HDACs1 and 2 are among the targets of these inhibitors and are part of multisubunit protein complexes. HDAC inhibitors (HDACi) block the activity of HDACs by chelating a zinc molecule in their catalytic sites. It is not known if the inhibitors have any additional functional effects on the multisubunit HDAC complexes. Here, we find that suberoylanilide hydroxamic acid (SAHA), the recently FDA approved HDACi, causes the dissociation of the PHD-finger containing ING2 subunit from the Sin3 deacetylase complex. Loss of ING2 disrupts the in vivo binding of the Sin3 complex to the p21 promoter, an important target gene for cell growth inhibition by SAHA. Our findings reveal a new molecular mechanism by which HDAC inhibitors disrupt deacetylase function. PMID:20142042

  9. Inhibition of class IIb histone deacetylase significantly improves cloning efficiency in mice.

    PubMed

    Ono, Tetsuo; Li, Chong; Mizutani, Eiji; Terashita, Yukari; Yamagata, Kazuo; Wakayama, Teruhiko

    2010-12-01

    Since the first mouse clone was produced by somatic cell nuclear transfer, the success rate of cloning in mice has been extremely low. Some histone deacetylase inhibitors, such as trichostatin A and scriptaid, have improved the full-term development of mouse clones significantly, but the mechanisms allowing for this are unclear. Here, we found that two other specific inhibitors, suberoylanilide hydroxamic acid and oxamflatin, could also reduce the rate of apoptosis in blastocysts, improve the full-term development of cloned mice, and increase establishment of nuclear transfer-generated embryonic stem cell lines significantly without leading to obvious abnormalities. However, another inhibitor, valproic acid, could not improve cloning efficiency. Suberoylanilide hydroxamic acid, oxamflatin, trichostatin A, and scriptaid are inhibitors for classes I and IIa/b histone deacetylase, whereas valproic acid is an inhibitor for classes I and IIa, suggesting that inhibiting class IIb histone deacetylase is an important step for reprogramming mouse cloning efficiency.

  10. Implications of intermediate mass black hole in globular cluster G1 on dark matter detection.

    SciTech Connect

    Zaharijas, G.; High Energy Physics

    2008-07-01

    Recently there has been growing evidence in favor of the presence of an intermediate mass black hole in the globular cluster G1, in Andromeda Galaxy. Under the assumption that formation of this globular cluster occurred within a dark matter halo, we explore whether the presence of a black hole could result in an observable gamma ray signal due to dark matter annihilation in this globular cluster. Starting from an initial Navarro-Frenk-White matter profile, with density parameters consistent with G1 observations, we find that indeed, if the spike in the density has been formed and has survived until the present, the signal could be observed by GLAST and current atmospheric Cerenkov telescope detectors.

  11. Permanent draft genome sequence of the gliding predator Saprospira grandis strain Sa g1 (= HR1)

    SciTech Connect

    Mavromatis, K; Chertkov, Olga; Lapidus, Alla L.; Nolan, Matt; Lucas, Susan; Tice, Hope; Glavina Del Rio, Tijana; Cheng, Jan-Fang; Han, Cliff; Tapia, Roxanne; Bruce, David; Goodwin, Lynne A.; Pitluck, Sam; Huntemann, Marcel; Liolios, Konstantinos; Pagani, Ioanna; Ivanova, N; Mikhailova, Natalia; Pati, Amrita; Chen, Amy; Palaniappan, Krishna; Land, Miriam L; Brambilla, Evelyne-Marie; Rohde, Manfred; Spring, Stefan; Goker, Markus; Detter, J. Chris; Bristow, James; Eisen, Jonathan; Markowitz, Victor; Hugenholtz, Philip; Kyrpides, Nikos C; Klenk, Hans-Peter; Woyke, Tanja

    2012-01-01

    Saprospira grandis Gross et al. 1911 is a member of the Saprospiraceae, a family in the class 'Sphingobacteria' that remains poorly characterized at the genomic level. The species is known for preying on other marine bacteria via 'ixotrophy'. S. grandis strain Sa g1 was isolated from decaying crab carapace in France and was selected for genome sequencing because of its isolated location in the tree of life. Only one type strain genome has been published so far from the Saprospiraceae, while the sequence of strain Sa g1 represents the second genome to be published from a non-type strain of S. grandis. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 4,495,250 bp long Improved-High-Quality draft of the genome with its 3,536 protein-coding and 62 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

  12. G1 to S phase cell cycle transition in somatic and embryonic stem cells

    PubMed Central

    Neganova, Irina; Lako, Majlinda

    2008-01-01

    It is well known that G1 to S phase transition is tightly regulated by the expression and phosphorylation of a number of well-characterized cyclins, cyclin-dependent kinases and members of the retinoblastoma gene family. In this review we discuss the role of these components in regulation of G1 to S phase transition in somatic cells and human embryonic stem cells. Most importantly, we discuss some new tenable links between maintenance of pluripotency and cell cycle regulation in embryonic stem cells by describing the role that master transcription factors play in this process. Finally, the differences in cell cycle regulation between murine and human embryonic stem cells are highlighted, raising interesting questions regarding their biology and stages of embryonic development from which they have been derived. PMID:18638068

  13. Four free cysteine residues found in human IgG1 of healthy donors.

    PubMed

    Gevondyan, N M; Volynskaia, A M; Gevondyan, V S

    2006-03-01

    Modifications with different thiol reagents demonstrated that 28 of 32 cysteine residues of human IgG1 are involved in the formation of disulfide bonds, and four cysteines remain free. So IgG1 is a protein possessing both free SH-groups and disulfide bonds. Only one of the four SH-groups is accessible for silver or mercury ions and hydrophobic reagents, whereas the remaining three SH-groups are masked and can be revealed only after deep denaturation of the protein. Detection of the masked cysteine residues was shown to depend on the kinetics of intramolecular changes occurring during denaturation of the protein and on the method of the assay of the SH-groups.

  14. EXPANSION OF THE YOUNGEST GALACTIC SUPERNOVA REMNANT G1.9+0.3

    SciTech Connect

    Carlton, Ashley K.; Borkowski, Kazimierz J.; Reynolds, Stephen P.; Hwang, Una; Petre, Robert; Krishnamurthy, Kalyani; Willett, Rebecca

    2011-08-10

    We present a measurement of the expansion and brightening of G1.9 + 0.3, the youngest Galactic supernova remnant (SNR), comparing Chandra X-ray images obtained in 2007 and 2009. A simple uniform-expansion model describes the data well, giving an expansion rate of 0.642% {+-} 0.049% yr{sup -1} and a flux increase of 1.7% {+-} 1.0% yr{sup -1}. Without deceleration, the remnant age would then be 156 {+-} 11 yr, consistent with earlier results. Since deceleration must have occurred, this age is an upper limit; we estimate an age of about 110 yr or an explosion date of about 1900. The flux increase is comparable to reported increases at radio wavelengths. G1.9+0.3 is the only Galactic SNR increasing in flux, with implications for the physics of electron acceleration in shock waves.

  15. Asymmetric Fab glycosylation in guinea-pig IgG1 and IgG2.

    PubMed Central

    Malan Borel, I; Gentile, T; Angelucci, J; Margni, R A; Binaghi, R A

    1990-01-01

    The presence of asymmetric antibody molecules has been investigated in both IgG1 and IgG2 subclasses of guinea-pig immunoglobulins. It was found that about 20% of the IgG1 and 10% of the IgG2 were of asymmetric type. The proportion was essentially the same in the sera of normal animals, animals hyperimmunized with dinitrophenyl-bovine gamma globulin (DNP-BGG) and Freund's adjuvant, and animals infected with Trichinella spiralis. In the case of animals immunized with DNP-BGG, no differences were observed in the proportion of asymmetric molecules between the specific antibodies and the IgG not specific for the immunizing antigen. It is concluded that the asymmetric glycosylation occurs to a different extent in each subclass and that it is not affected by the antigen specificity of the antibodies studied. PMID:2379937

  16. PBOV1 promotes prostate cancer proliferation by promoting G1/S transition

    PubMed Central

    Pan, Tiejun; Wu, Rongpei; Liu, Bo; Wen, Handong; Tu, Zhong; Guo, Jun; Yang, Jiarong; Shen, Guoqiu

    2016-01-01

    Prostate cancer (PC) is one of the leading causes of cancer death in men, and thus, finding new regulators is critical for PC therapy. Prostate and breast cancer overexpressed 1 (PBOV1) is overexpressed in breast, prostate, and bladder cancers, as it is upregulated in the serum of patients with PC, but the role of PBOV1 in PC has not been studied. In this article, we found that PBOV1 was indeed overexpressed in PC cells; PBOV1 overexpression promoted cell proliferation and colony formation ability and arrested cell cycle in the G0/G1 phase and tumorigenicity ability in vitro, whereas knockdown of PBOV1 reduced these effects. Further analysis of PBOV1 overexpression inhibited cell cycle inhibitors, P21 and P27, and increased the phosphorylation level of Rb and cyclin D1 expression, suggesting that PBOV1 promoted cell proliferation through promoting G1/S transition. PMID:26937201

  17. Weighted G0-and G1-multi-degree reduction of Bézier curves

    NASA Astrophysics Data System (ADS)

    Rababah, Abedallah; Ibrahim, Salisu

    2016-06-01

    In this paper, weighted G0-and G1-multi-degree reduction of Bézier curves are considered. The degree reduction of a given Bézier curve of degree n is used to write it as a Bézier curve of degree m, m < n. Exact degree reduction is not possible, and, therefore approximation methods are used. The weight function w[t] = 2t(1 - t), t ∈ [0, 1] is used with the L2 -norm in multi degree reduction with G0- and G1- continuity at the end points of the curve. Numerical results and comparisons show that the new methods suggests smaller approximation errors in the interior of the domain and proves to be competative in applications.

  18. Global Structure of HIV-1 Neutralizing Antibody IgG1 b12 is Asymmetric

    SciTech Connect

    Ashish, F.; Solanki, A; Boone, C; Krueger, J

    2010-01-01

    Human antibody IgG1 b12 is one of the four antibodies known to neutralize a broad range of human immunodeficiency virus-1. The crystal structure of this antibody displayed an asymmetric disposition of the Fab arms relative to its Fc portion. Comparison of structures solved for other IgG1 antibodies led to a notion that crystal packing forces entrapped a 'snap-shot' of different conformations accessible to this antibody. To elucidate global structure of this unique antibody, we acquired small-angle X-ray scattering data from its dilute solution. Data analysis indicated that b12 adopts a bilobal globular structure in solution with a radius of gyration and a maximum linear dimension of {approx}54 and {approx}180 {angstrom}, respectively. Extreme similarity between its solution and crystal structure concludes that non-flexible, asymmetric shape is an inherent property of this rare antibody.

  19. 26 CFR 1.1402(g)-1 - Treatment of certain remuneration erroneously reported as net earnings from self-employment.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... reported as net earnings from self-employment. 1.1402(g)-1 Section 1.1402(g)-1 Internal Revenue INTERNAL...-Employment Income § 1.1402(g)-1 Treatment of certain remuneration erroneously reported as net earnings from... request, and should indicate clearly that it is a request that, pursuant to section 1402(g) of the Code...

  20. 26 CFR 31.3406(g)-1 - Exception for payments to certain payees and certain other payments.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 15 2010-04-01 2010-04-01 false Exception for payments to certain payees and certain other payments. 31.3406(g)-1 Section 31.3406(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... TAXES AND COLLECTION OF INCOME TAX AT SOURCE Collection of Income Tax at Source § 31.3406(g)-1...

  1. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  2. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  3. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  4. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  5. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  6. 17 CFR 270.17g-1 - Bonding of officers and employees of registered management investment companies.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 17 Commodity and Securities Exchanges 3 2011-04-01 2011-04-01 false Bonding of officers and employees of registered management investment companies. 270.17g-1 Section 270.17g-1 Commodity and... ACT OF 1940 § 270.17g-1 Bonding of officers and employees of registered management...

  7. Aircraft measurements of aerosol properties during GoAmazon - G1 and HALO inter-comparison

    NASA Astrophysics Data System (ADS)

    Mei, F.; Cecchini, M. A.; Wang, J.; Tomlinson, J. M.; Comstock, J. M.; Hubbe, J. M.; Pekour, M. S.; Machado, L.; Wendisch, M.; Longo, K.; Martin, S. T.; Schmid, B.; Weinzierl, B.; Krüger, M. L.; Zöger, M.

    2015-12-01

    Currently, the indirect effects of atmospheric aerosols remain the most uncertain components in forcing of climate change over the industrial period (IPCC, 2013). This large uncertainty is partially a result of our incomplete understanding of the ability of particles to form cloud droplets under atmospherically relevant supersaturations. One objective of the US Department of Energy (DOE) Green Ocean Amazon Project (GoAmazon2014/5) is to understand the influence of the emission from Manaus, a tropical megacity, on aerosol size, concentration, and chemical composition, and their impact on cloud condensation nuclei (CCN) spectrum. The GoAmazon2014/5 study was an international campaign with the collaboration efforts from US, Brazil and Germany. During the intensive operation period, in the dry season (Sep. 1st - Oct. 10th, 2014), aerosol concentration, size distributions, and CCN spectra, both under pristine conditions and inside the Manaus plume, were characterized in-situ from the DOE Gulfstream-1 (G-1) research aircraft and German HALO aircraft during 4 coordinated flights on Sep. 9th, Sep. 16th, Sep 21st and Oct. 1st, 2014. During those four flights, aerosol number concentrations and CCN concentrations at two supersaturations (0.25% and 0.5%) were measured by condensation particle counters (CPCs) and a DMT dual column CCN counter onboard both G-1 and HALO. Aerosol size distribution was also measured by a Fast Integrated Mobility Spectrometer (FIMS) aboard the G-1 and is compared with the size distribution from Ultra High Sensitivity Aerosol Spectrometer - Airborne (UHSAS-A, DMT), which were deployed both on the G-1 and the HALO. Good agreement between the aerosol properties measured from the two aircraft has been achieved. The vertical profiles of aerosol size distribution and CCN spectrum will be discussed.

  8. M/G/1 Subject to an Initial Quorum of Customers.

    DTIC Science & Technology

    1986-11-01

    size" for : i M/G/1(m). (Note: In that prior report m stands for the current m+1. The change simplifies somewhat the typography of many formulas.) We...2*A+x)+ 4)(A+2 x)+ 4(3*x)a + p (w.) ". where w. is the delay conditioned upon the server working. We simplify the typography of (1.4) and (1.4) by

  9. Structural Changes and Aggregation Mechanisms for Anti-Streptavidin IgG1 at Elevated Concentration.

    PubMed

    Barnett, Gregory V; Qi, Wei; Amin, Samiul; Lewis, E Neil; Razinkov, Vladimir I; Kerwin, Bruce A; Liu, Yun; Roberts, Christopher J

    2015-12-10

    Non-native protein aggregation may occur during manufacturing and storage of protein therapeutics, and this may decrease drug efficacy or jeopardize patient safety. From a regulatory perspective, changes in higher order structure due to aggregation are of particular interest but can be difficult to monitor directly at elevated protein concentrations. The present report focuses on non-native aggregation of antistreptavidin (AS) IgG1 at 30 mg/mL under solution conditions that prior work at dilute concentrations (e.g., 1 mg/mL) indicated would result in different aggregation mechanisms. Time-dependent aggregation and structural changes were monitored in situ with dynamic light scattering, small-angle neutron scattering, and Raman scattering and ex situ with far-UV circular dichroism and second-derivative UV spectroscopy. The effects of adding 0.15 M (∼5 w/w %) sucrose were also assessed. The addition of sucrose decreased monomer loss rates but did not change protein-protein interactions, aggregation mechanism(s), or aggregate structure and morphology. Consistent with prior results, altering the pD or salt concentration had the primary effect of changing the aggregation mechanism. Overall, the results provide a comparison of aggregate structure and morphology created via different growth mechanisms using orthogonal techniques and show that the techniques agree at least qualitatively. Interestingly, AS-IgG1 aggregates created at pD 5.3 with no added salt formed the smallest aggregates but had the largest structural changes compared to other solution conditions. The observation that the larger aggregates were also those with less structural perturbation compared to folded AS-IgG1 might be expected to extend to other proteins if the same strong electrostatic repulsions that mediate aggregate growth also mediate structural changes of the constituent proteins within aggregates.

  10. NONUNIFORM EXPANSION OF THE YOUNGEST GALACTIC SUPERNOVA REMNANT G1.9+0.3

    SciTech Connect

    Borkowski, Kazimierz J.; Reynolds, Stephen P.; Green, David A.; Hwang, Una; Petre, Robert; Willett, Rebecca

    2014-08-01

    We report measurements of the X-ray expansion of the youngest Galactic supernova remnant, G1.9+0.3, using Chandra observations in 2007, 2009, and 2011. The measured rates strongly deviate from uniform expansion, decreasing radially by about 60% along the X-ray bright SE-NW axis from 0.84% ± 0.06% yr{sup –1} to 0.52% ± 0.03% yr{sup –1}. This corresponds to undecelerated ages of 120-190 yr, confirming the young age of G1.9+0.3 and implying a significant deceleration of the blast wave. The synchrotron-dominated X-ray emission brightens at a rate of 1.9% ± 0.4% yr{sup –1}. We identify bright outer and inner rims with the blast wave and reverse shock, respectively. Sharp density gradients in either the ejecta or ambient medium are required to produce the sudden deceleration of the reverse shock or the blast wave implied by the large spread in expansion ages. The blast wave could have been decelerated recently by an encounter with a modest density discontinuity in the ambient medium, such as may be found at a wind termination shock, requiring strong mass loss in the progenitor. Alternatively, the reverse shock might have encountered an order-of-magnitude density discontinuity within the ejecta, such as may be found in pulsating delayed-detonation Type Ia models. We demonstrate that the blast wave is much more decelerated than the reverse shock in these models for remnants at ages similar to G1.9+0.3. Similar effects may also be produced by dense shells possibly associated with high-velocity features in Type Ia spectra. Accounting for the asymmetry of G1.9+0.3 will require more realistic three-dimensional Type Ia models.

  11. On the first G 1 stiff fluid spike solution in General Relativity

    NASA Astrophysics Data System (ADS)

    Coley, A. A.; Gregoris, D.; Lim, W. C.

    2016-11-01

    Using the Geroch transformation we obtain the first example of an exact stiff fluid spike solution to the Einstein field equations in a closed form exhibiting a spacelike G 1 group of symmetries (i.e., with a single isometry). This new solution is of Petrov type I and exhibits a spike crossing which persists to the past, which allows us to better understand spike crossings in the context of structure formation.

  12. Nonuniform Expansion of the Youngest Galactic Supernova Remnant G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Borkowski, Kazimierz J.; Reynolds, Stephen P.; Green, David A.; Hwang, Una; Petre, Robert; Krishnamurthy, Kalyani; Willett, Rebecca

    2014-08-01

    We report measurements of the X-ray expansion of the youngest Galactic supernova remnant, G1.9+0.3, using Chandra observations in 2007, 2009, and 2011. The measured rates strongly deviate from uniform expansion, decreasing radially by about 60% along the X-ray bright SE-NW axis from 0.84% ± 0.06% yr-1 to 0.52% ± 0.03% yr-1. This corresponds to undecelerated ages of 120-190 yr, confirming the young age of G1.9+0.3 and implying a significant deceleration of the blast wave. The synchrotron-dominated X-ray emission brightens at a rate of 1.9% ± 0.4% yr-1. We identify bright outer and inner rims with the blast wave and reverse shock, respectively. Sharp density gradients in either the ejecta or ambient medium are required to produce the sudden deceleration of the reverse shock or the blast wave implied by the large spread in expansion ages. The blast wave could have been decelerated recently by an encounter with a modest density discontinuity in the ambient medium, such as may be found at a wind termination shock, requiring strong mass loss in the progenitor. Alternatively, the reverse shock might have encountered an order-of-magnitude density discontinuity within the ejecta, such as may be found in pulsating delayed-detonation Type Ia models. We demonstrate that the blast wave is much more decelerated than the reverse shock in these models for remnants at ages similar to G1.9+0.3. Similar effects may also be produced by dense shells possibly associated with high-velocity features in Type Ia spectra. Accounting for the asymmetry of G1.9+0.3 will require more realistic three-dimensional Type Ia models.

  13. Localization of the X-ray source in the globular cluster G1 with Chandra

    NASA Astrophysics Data System (ADS)

    Kong, A. K. H.; Heinke, C. O.; di Stefano, R.; Cohn, H. N.; Lugger, P. M.; Barmby, P.; Lewin, W. H. G.; Primini, F. A.

    2010-09-01

    We report the most accurate X-ray position of the X-ray source in the giant globular cluster G1 in M31 by using the Chandra X-ray Observatory, Hubble Space Telescope (HST) and Canada-France-Hawaii Telescope (CFHT). G1 is clearly detected with Chandra and by cross-registering with HST and CFHT images, we derive a 1σ error radius of 0.15arcsec, significantly smaller than the previous measurement by XMM-Newton. We conclude that the X-ray emission of G1 is likely to come from within the core radius of the cluster. We have considered a number of possibilities for the origin of the X-ray emission but can rule all but two scenarios out: it could be due to either accretion on to a central intermediate-mass black hole (IMBH) or an ordinary low-mass X-ray binary (LMXB). Based on the X-ray luminosity and the Bondi accretion rate, an IMBH accreting from the cluster gas seems unlikely and we suggest that the X-rays are due to accretion from a companion. Alternatively, the probability that a 1.5 Msolar cluster LMXB lies within the 95 per cent X-ray error circle is about 0.7. Therefore we cannot rule out a single LMXB as the origin of the X-ray emission. While we cannot distinguish between different models with current observations, future high-resolution and high-sensitivity radio imaging observations will reveal whether there is an IMBH at the centre of G1.

  14. The Absence of Radio Emission from the Globular Cluster G1

    NASA Astrophysics Data System (ADS)

    Miller-Jones, J. C. A.; Wrobel, J. M.; Sivakoff, G. R.; Heinke, C. O.; Miller, R. E.; Plotkin, R. M.; Di Stefano, R.; Greene, J. E.; Ho, L. C.; Joseph, T. D.; Kong, A. K. H.; Maccarone, T. J.

    2012-08-01

    The detections of both X-ray and radio emission from the cluster G1 in M31 have provided strong support for existing dynamical evidence for an intermediate-mass black hole (IMBH) of mass (1.8 ± 0.5) × 104 M ⊙ at the cluster center. However, given the relatively low significance and astrometric accuracy of the radio detection, and the non-simultaneity of the X-ray and radio measurements, this identification required further confirmation. Here we present deep, high angular resolution, strictly simultaneous X-ray and radio observations of G1. While the X-ray emission (L X = 1.74+0.53 -0.44 × 1036 (d/750 kpc)2 erg s-1 in the 0.5-10 keV band) remained fully consistent with previous observations, we detected no radio emission from the cluster center down to a 3σ upper limit of 4.7 μJy beam-1. Our favored explanation for the previous radio detection is flaring activity from a black hole low-mass X-ray binary (LMXB). We performed a new regression of the "Fundamental Plane" of black hole activity, valid for determining black hole mass from radio and X-ray observations of sub-Eddington black holes, finding log M BH = (1.638 ± 0.070)log L R - (1.136 ± 0.077)log L X - (6.863 ± 0.790), with an empirically determined uncertainty of 0.44 dex. This constrains the mass of the X-ray source in G1, if a black hole, to be <9.7 × 103 M ⊙ at 95% confidence, suggesting that it is a persistent LMXB. This annuls what was previously the most convincing evidence from radiation for an IMBH in the Local Group, though the evidence for an IMBH in G1 from velocity dispersion measurements remains unaffected by these results.

  15. Insecticidal activities of histone deacetylase inhibitors against a dipteran parasite of sheep, Lucilia cuprina.

    PubMed

    Bagnall, Neil H; Hines, Barney M; Lucke, Andrew J; Gupta, Praveer K; Reid, Robert C; Fairlie, David P; Kotze, Andrew C

    2017-04-01

    Histone deacetylase inhibitors (HDACi) are being investigated for the control of various human parasites. Here we investigate their potential as insecticides for the control of a major ecto-parasite of sheep, the Australian sheep blowfly, Lucilia cuprina. We assessed the ability of HDACi from various chemical classes to inhibit the development of blowfly larvae in vitro, and to inhibit HDAC activity in nuclear protein extracts prepared from blowfly eggs. The HDACi prodrug romidepsin, a cyclic depsipeptide that forms a thiolate, was the most potent inhibitor of larval growth, with equivalent or greater potency than three commercial blowfly insecticides. Other HDACi with potent activity were hydroxamic acids (trichostatin, CUDC-907, AR-42), a thioester (KD5170), a disulphide (Psammaplin A), and a cyclic tetrapeptide bearing a ketone (apicidin). On the other hand, no insecticidal activity was observed for certain other hydroxamic acids, fatty acids, and the sesquiterpene lactone parthenolide. The structural diversity of the 31 hydroxamic acids examined here revealed some structural requirements for insecticidal activity; for example, among compounds with flexible linear zinc-binding extensions, greater potency was observed in the presence of branched capping groups that likely make multiple interactions with the blowfly HDAC enzymes. The insecticidal activity correlated with inhibition of HDAC activity in blowfly nuclear protein extracts, indicating that the toxicity was most likely due to inhibition of HDAC enzymes in the blowfly larvae. The inhibitor potencies against blowfly larvae are different from inhibition of human HDACs, suggesting some selectivity for human over blowfly HDACs, and a potential for developing compounds with the inverse selectivity. In summary, these novel findings support blowfly HDAC enzymes as new targets for blowfly control, and point to development of HDAC inhibitors as a promising new class of insecticides.

  16. Synergy between histone deacetylase inhibitors and DNA-damaging agents is mediated by histone deacetylase 2 in colorectal cancer

    PubMed Central

    Alzoubi, Samer; Brody, Leigh; Rahman, Sunniyat; Mahul-Mellier, Anne-Laure; Mercado, Nicolas; Ito, Kazuhiro; El-Bahrawy, Mona; Silver, Andrew; Boobis, Alan; Bell, Jimmy D.; Hajji, Nabil

    2016-01-01

    Previous studies have associated the overexpression of histone deacetylase 2 (HDAC2) and the presence of TP53 mutations with the progression to advanced stage drug resistant colorectal cancer (CRC). However, the mechanistic link between HDAC2 expression and the TP53 mutational status has remained unexplored. Here, we investigated the function of HDAC2 in drug resistance by assessing the synergistic effects of DNA-targeted chemotherapeutic agents and HDAC inhibitors (HDACis) on two TP53-mutated colorectal adenocarcinoma CRC cell lines (SW480 and HT-29) and on the TP53-wild type carcinoma cell line (HCT116 p53+/+) and its TP53 deficient sub-line (HCT116 p53−/−). We showed that in the untreated SW480 and HT-29 cells the steady-state level of HDAC2 was low compared to a TP53-wild type carcinoma cell line (HCT116 p53+/+). Increased expression of HDAC2 correlated with drug resistance, and depletion by shRNA sensitised the multi-drug resistance cell line HT-29 to CRC chemotherapeutic drugs such as 5-fluorouracil (5-FU) and oxaliplatin (Oxa). Combined treatment with the HDACi suberoylanilide hydroxamic acid plus 5-FU or Oxa reduced the level of HDAC2 expression, modified chromatin structure and induced mitotic cell death in HT-29 cells. Non-invasive bioluminescence imaging revealed significant reductions in xenograft tumour growth with HDAC2 expression level reduced to <50% in treated animals. Elevated levels of histone acetylation on residues H3K9, H4K12 and H4K16 were also found to be associated with resistance to VPA/Dox or SAHA/Dox treatment. Our results suggest that HDAC2 expression rather than the p53 mutation status influences the outcome of combined treatment with a HDACi and DNA-damaging agents in CRC. PMID:27283986

  17. HJURP interaction with the condensin II complex during G1 promotes CENP-A deposition

    PubMed Central

    Barnhart-Dailey, Meghan C.; Trivedi, Prasad; Stukenberg, P. Todd; Foltz, Daniel R.

    2017-01-01

    Centromeric chromatin is required for kinetochore assembly during mitosis and accurate chromosome segregation. A unique nucleosome containing the histone H3–specific variant CENP-A is the defining feature of centromeric chromatin. In humans, CENP-A nucleosome deposition occurs in early G1 just after mitotic exit at the time when the CENP-A deposition machinery localizes to centromeres. The mechanism by which CENP-A is deposited onto an existing, condensed chromatin template is not understood. Here we identify the selective association of the CENP-A chaperone HJURP with the condensin II complex and not condensin I. We show CAPH2 is present at centromeres during early G1 at the time when CENP-A deposition is occurring. CAPH2 localization to early G1 centromeres is dependent on HJURP. The CENP-A chaperone and assembly factor HJURP induces decondensation of a noncentromeric LacO array, and this decondensation is modulated by the condensin II complex. We show that condensin II function at the centromere is required for new CENP-A deposition in human cells. These data demonstrate that HJURP selectively recruits the condensin II chromatin-remodeling complex to facilitate CENP-A deposition in human cells. PMID:27807043

  18. Treatment of G1 Baskets at the CEA Marcoule Site - 12027

    SciTech Connect

    Fourquet, Line; Boya, Didier

    2012-07-01

    In the dismantling program for the first-generation French reactors in accordance with the nonproliferation treaty, the CEA is in charge of cleanup and dismantling operations for the facilities at Marcoule, including the decladding units. The G1 decladding was built between 1955 and 1957 in order to de-clad spent fuel elements from the G1 plutonium-producing reactor and prepare them for dissolution. The facility was also used for interim storage of G1, G2 and G3 fuel dissolution baskets, which had been used during plant operation for transfer (from the decladding facility to the UP1 plant) and/or dissolution of spent fuel elements. One of the cleanup projects involves recovery of the baskets, which will be cut up, sorted, and conditioned in metal bins. The bins will be immobilized with cement grout, then transferred to the onsite solid waste conditioning facility (CDS) and to the repository operated by the French National Radioactive Waste Management Agency (ANDRA). The project is now in progress, after special safety permits were issued and measurement stations and dedicated tools were developed to handle all types of baskets (which differed according to their origin and use). The disposal of all the baskets is scheduled to last 2 years and will produce 55 metal waste bins. (authors)

  19. Replication stress induces 53BP1-containing OPT domains in G1 cells

    PubMed Central

    Harrigan, Jeanine A.; Belotserkovskaya, Rimma; Coates, Julia; Dimitrova, Daniela S.; Polo, Sophie E.; Bradshaw, Charles R.; Fraser, Peter

    2011-01-01

    Chromosomal deletions and rearrangements in tumors are often associated with common fragile sites, which are specific genomic loci prone to gaps and breaks in metaphase chromosomes. Common fragile sites appear to arise through incomplete DNA replication because they are induced after partial replication inhibition by agents such as aphidicolin. Here, we show that in G1 cells, large nuclear bodies arise that contain p53 binding protein 1 (53BP1), phosphorylated H2AX (γH2AX), and mediator of DNA damage checkpoint 1 (MDC1), as well as components of previously characterized OPT (Oct-1, PTF, transcription) domains. Notably, we find that incubating cells with low aphidicolin doses increases the incidence and number of 53BP1-OPT domains in G1 cells, and by chromatin immunoprecipitation and massively parallel sequencing analysis of γH2AX, we demonstrate that OPT domains are enriched at common fragile sites. These findings invoke a model wherein incomplete DNA synthesis during S phase leads to a DNA damage response and formation of 53BP1-OPT domains in the subsequent G1. PMID:21444690

  20. SUMOylation of Rb enhances its binding with CDK2 and phosphorylation at early G1 phase.

    PubMed

    Meng, Fengxi; Qian, Jiang; Yue, Han; Li, Xiaofeng; Xue, Kang

    2016-07-02

    Retinoblastoma protein (Rb) is a prototypical tumor suppressor that is vital to the negative regulation of the cell cycle and tumor progression. Hypo-phosphorylated Rb is associated with G0/G1 arrest by suppressing E2F transcription factor activity, whereas Rb hyper-phosphorylation allows E2F release and cell cycle progression from G0/G1 to S phase. However, the factors that regulate cyclin-dependent protein kinase (CDK)-dependent hyper-phosphorylation of Rb during the cell cycle remain obscure. In this study, we show that throughout the cell cycle, Rb is specifically small ubiquitin-like modifier (SUMO)ylated at early G1 phase. SUMOylation of Rb stimulates its phosphorylation level by recruiting a SUMO-interaction motif (SIM)-containing kinase CDK2, leading to Rb hyper-phosphorylation and E2F-1 release. In contrast, a SUMO-deficient Rb mutant results in reduced SUMOylation and phosphorylation, weakened CDK2 binding, and attenuated E2F-1 sequestration. Furthermore, we reveal that Rb SUMOylation is required for cell proliferation. Therefore, our study describes a novel mechanism that regulates Rb phosphorylation during cell cycle progression.

  1. Tet1 is required for Rb phosphorylation during G1/S phase transition

    SciTech Connect

    Huang, Shengsong; Zhu, Ziqi; Wang, Yiqin; Wang, Yanru; Xu, Longxia; Chen, Xuemei; Xu, Qing; Zhang, Qimin; Zhao, Xin; Yu, Yi; Wu, Denglong

    2013-05-03

    Highlights: •Tet1 was required for NIT3T3 proliferation. •Tet1 depletion inhibited G1-S entry. •Cyclin D1 accumulation and Rb phosphorylation was blocked by Tet1 knockdown. -- Abstract: DNA methylation plays an important role in many biological processes, including regulation of gene expression, maintenance of chromatin conformation and genomic stability. TET-family proteins convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), which indicates that these enzymes may participate in DNA demethylation. The function of TET1 has not yet been well characterized in somatic cells. Here, we show that depletion of Tet1 in NIH3T3 cells inhibits cell growth. Furthermore, Tet1 knockdown blocks cyclin D1 accumulation in G1 phase, inhibits Rb phosphorylation and consequently delays entrance to G1/S phase. Taken together, this study demonstrates that Tet1 is required for cell proliferation and that this process is mediated through the Rb pathway.

  2. Acetyl-lysine analog peptides as mechanistic probes of protein deacetylases.

    PubMed

    Smith, Brian C; Denu, John M

    2007-12-21

    Class III histone deacetylases (Sir2 or sirtuins) catalyze the NAD+-dependent conversion of acetyl-lysine residues to nicotinamide, 2'-O-acetyl-ADP-ribose (OAADPr), and deacetylated lysine. Class I and II HDACs utilize a different deacetylation mechanism, utilizing an active site zinc to direct hydrolysis of acetyl-lysine residues to lysine and acetate. Here, using ten acetyl-lysine analog peptides, we have probed the substrate binding pockets of sirtuins and investigated the catalytic differences among sirtuins and class I and II deacetylases. For the sirtuin Hst2, acetyl-lysine analog peptide binding correlated with the hydrophobic substituent parameter pi with a slope of -0.35 from a plot of log Kd versus pi. Interestingly, propionyl- and butyryl-lysine peptides were found to bind tighter to Hst2 compared with acetyl-lysine peptide and showed measurable rates of catalysis with Hst2, Sirt1, Sirt2, and Sirt3, suggesting propionyl- and butyryl-lysine proteins may be sirtuin substrates in vivo. Unique among the acetyl-lysine analog peptides examined, homocitrulline peptide produced ADP-ribose instead of the corresponding OAADPr analog. The electron-withdrawing nature of each acetyl analog had a profound impact on the deacylation rate between deacetylase classes. The rate of catalysis with the acetyl-lysine analog peptides varied over five orders of magnitude with the class III deacetylase Hst2, revealing a linear free energy relationship with a slope of -1.57 when plotted versus the Taft constant, sigma*. HDAC8, a class I deacetylase, displayed the opposite trend with a slope of +0.79. These results are applicable toward the development of selective substrates and other mechanistic probes of protein deacetylases.

  3. Mycobacterium tuberculosis Rv1096 protein: gene cloning, protein expression, and peptidoglycan deacetylase activity

    PubMed Central

    2014-01-01

    Background Many bacteria modulate and evade the immune defenses of their hosts through peptidoglycan (PG) deacetylation. The PG deacetylases from Streptococcus pneumonia, Listeria monocytogenes and Lactococcus lactis have been characterized. However, thus far, the PG deacetylase of Mycobacterium tuberculosis has not been identified. Results In this study, we cloned the Rv1096 gene from the M. tuberculosis H37Rv strain and expressed Rv1096 protein in both Escherichia coli and M. smegmatis. The results showed that the purified Rv1096 protein possessed metallo-dependent PG deacetylase activity, which increased in the presence of Co2+. The kinetic parameters of the PG deacetylase towards M. smegmatis PG as a substrate were as follows: Km, 0.910 ± 0.007 mM; Vmax, 0.514 ± 0.038 μMmin-1; and Kcat = 0.099 ± 0.007 (S-1). Additionally, the viability of M. smegmatis in the presence of over-expressed Rv1096 protein was 109-fold higher than that of wild-type M. smegmatis after lysozyme treatment. Additionally, light microscopy and scanning electron microscopy showed that in the presence of over-expressed Rv1096 protein, M. smegmatis kept its regular shape, with an undamaged cell wall and smooth surface. These results indicate that Rv1096 caused deacetylation of cell wall PG, leading to lysozyme resistance in M. smegmatis. Conclusion We have determined that M. tuberculosis Rv1096 is a PG deacetylase. The PG deacetylase activity of Rv1096 contributed to lysozyme resistance in M. smegmatis. Our findings suggest that deacetylation of cell wall PG may be involved in evasion of host immune defenses by M. tuberculosis. PMID:24975018

  4. The human G1m1 allotype associates with CD4+ T-cell responsiveness to a highly conserved IgG1 constant region peptide and confers an asparaginyl endopeptidase cleavage site

    PubMed Central

    Stickler, M M; Reddy, A; Xiong, J M; Hinton, P R; DuBridge, R; Harding, F A

    2011-01-01

    The human G1m1 allotype comprises two amino acids, D12 and L14, in the CH3 domain of IGHG1. Although the G1m1 allotype is prevalent in human populations, ∼40% of Caucasiods are homozygous for the nG1m1 allotype corresponding to E12 and M14. Peptides derived from the G1m1 region were tested for their ability to induce CD4+ T-cell proliferative responses in vitro. A peptide immediately downstream from the G1m1 sequence was recognized by CD4+ T cells in a large percentage of donors (peptide CH315−29). CD4+ T-cell proliferative responses to CH315−29 were found at an increased frequency in nG1m1 homozygous donors. Homozygous nG1m1 donors possessing the HLA-DRB1*07 allele displayed the highest magnitudes of proliferation. CD4+ T cells from donors homozygous for nG1m1 proliferated to G1m1-carrying Fc-fragment proteins, whereas CD4+ T cells from G1m1 homozygous donors did not. The G1m1 sequence creates an enzymatic cleavage site for asparaginyl endopeptidase in vitro. Proteolytic activity at D12 may allow the presentation of the CH315−29 peptide, which in turn may result in the establishment of tolerance to this peptide in G1m1-positive donors. Homozygous nG1m1 patients may be more likely to develop CD4+ T-cell-mediated immune responses to therapeutic antibodies carrying the G1m1 allotype. PMID:21326320

  5. Histone deacetylase inhibitors: a potential epigenetic treatment for Duchenne muscular dystrophy.

    PubMed

    Consalvi, Silvia; Saccone, Valentina; Mozzetta, Chiara

    2014-01-01

    Duchenne muscular dystrophy (DMD) is a life-threatening genetic disease that currently has no available cure. A number of pharmacological strategies that aim to target events downstream of the genetic defect are currently under clinical investigation, and some of these are outlined in this report. In particular, we focus on the ability of histone deacetylase inhibitors to promote muscle regeneration and prevent the fibro-adipogenic degeneration of dystrophic mice. We describe the rationale behind the translation of histone deacetylase inhibitors into a clinical approach, which inspired the first clinical trial with an epigenetic drug as a potential therapeutic option for DMD patients.

  6. A novel G1-specific enhancer identified in the human heat shock protein 70 gene.

    PubMed Central

    Taira, T; Narita, T; Iguchi-Ariga, S M; Ariga, H

    1997-01-01

    Expression of the human heat shock protein 70 gene (hsp70) is induced by various kinds of stress and by oncogenes. In the absence of stress, hsp70 is mainly expressed in the G1and S phases of the cell cycle, but the elements contributing to cell cycle-dependent expression from the hsp70 promoter remain elusive. We have previously reported that two elements, named HSP-MYCA and HSP-MYCB, located approximately 200 bp upstream (-200) from the transcription start site (+1) of human hsp70, are important for initiation of DNA replication at the hsp70 locus. In this report we examine the effect of these two elements on transcriptional activity from the hsp70 promoter, especially in terms of cell cycle-dependent expression. Various segments of the hsp70 promoter region (up to -300) were linked to the luciferase gene and the constructs were transfected into mouse L cells to examine their transcriptional activity. A strong enhancer activity was defined in the HSP-MYCB element, but not in HSP-MYCA. Mutations introduced within HSP-MYCB abolished the transcriptional activation. In synchronized cells, pHB-Luc (a luciferase construct containing approximately 2.4 kb of the hsp70 promoter region) as well as endogenous hsp70 showed two peaks of expression; one in G1 and the other in the S phase. Site-directed mutagenesis of HSP-MYCB in pHB-Luc abolished the expression peak in G1, but not that in the S phase. To test promoter specificity, wild-type and mutant HSP-MYCB elements were then linked to the luciferase gene in combination with the hsp70 , the cyclin A or the PCNA promoter. Both in transient experiments and established cell lines, a strong peak of expression in mid-G1phase was observed with all the constructs containing wild-type HSP-MYCB, but not with the constructs containing the mutant sequence. These results suggest that the HSP-MYCB sequence is a G1-specific enhancer and is responsible for cell cycle-dependent expression of hsp70. PMID:9115365

  7. p67SRF is a constitutive nuclear protein implicated in the modulation of genes required throughout the G1 period.

    PubMed Central

    Gauthier-Rouvière, C; Cavadore, J C; Blanchard, J M; Lamb, N J; Fernandez, A

    1991-01-01

    Indirect immunofluorescence analysis, using antibodies directed against peptide sequences outside the DNA-binding domain of the 67-kDa serum response factor (p67SRF), revealed a punctuated nuclear staining, constant throughout the cell cycle and in all different cell lines tested. p67SRF was also tightly associated with chromatin through all stages of mitosis. Inhibition of p67SRF activity in vivo, through microinjection of anti-p67SRF antibodies, specifically suppressed DNA synthesis induced after serum addition or ras microinjection, suggesting that these antibodies were effective in preventing expression of serum response element (SRE)-regulated genes. A similar inhibition was also obtained in cells injected with oligonucleotides corresponding to the DNA binding sequence for p67SRF protein, SRE. Moreover, this inhibition of DNA synthesis by anti-p67SRF or SRE injection was still observed in cells injected during late G1, well after c-fos induction. These data imply that genes regulated by p67SRF are continuously involved in the proliferation pathway throughout G1 and that p67SRF forms an integral component of mammalian cell transcriptional control. Images PMID:1782216

  8. TopBP1 is required at mitosis to reduce transmission of DNA damage to G1 daughter cells

    PubMed Central

    Pedersen, Rune Troelsgaard; Kruse, Thomas; Nilsson, Jakob

    2015-01-01

    Genome integrity is critically dependent on timely DNA replication and accurate chromosome segregation. Replication stress delays replication into G2/M, which in turn impairs proper chromosome segregation and inflicts DNA damage on the daughter cells. Here we show that TopBP1 forms foci upon mitotic entry. In early mitosis, TopBP1 marks sites of and promotes unscheduled DNA synthesis. Moreover, TopBP1 is required for focus formation of the structure-selective nuclease and scaffold protein SLX4 in mitosis. Persistent TopBP1 foci transition into 53BP1 nuclear bodies (NBs) in G1 and precise temporal depletion of TopBP1 just before mitotic entry induced formation of 53BP1 NBs in the next cell cycle, showing that TopBP1 acts to reduce transmission of DNA damage to G1 daughter cells. Based on these results, we propose that TopBP1 maintains genome integrity in mitosis by controlling chromatin recruitment of SLX4 and by facilitating unscheduled DNA synthesis. PMID:26283799

  9. Ethanol extract of Innotus obliquus (Chaga mushroom) induces G1 cell cycle arrest in HT-29 human colon cancer cells

    PubMed Central

    Lee, Hyun Sook; Kim, Eun Ji

    2015-01-01

    BACKGROUND/OBJECTIVES Inonotus obliquus (I. obliquus, Chaga mushroom) has long been used as a folk medicine to treat cancer. In the present study, we examined whether or not ethanol extract of I. obliquus (EEIO) inhibits cell cycle progression in HT-29 human colon cancer cells, in addition to its mechanism of action. MATERIALS/METHODS To examine the effects of Inonotus obliquus on the cell cycle progression and the molecular mechanism in colon cancer cells, HT-29 human colon cancer cells were cultured in the presence of 2.5 - 10 µg/mL of EEIO, and analyzed the cell cycle arrest by flow cytometry and the cell cycle controlling protein expression by Western blotting. RESULTS Treatment cells with 2.5 - 10 µg/mL of EEIO reduced viable HT-29 cell numbers and DNA synthesis, increased the percentage of cells in G1 phase, decreased protein expression of CDK2, CDK4, and cyclin D1, increased expression of p21, p27, and p53, and inhibited phosphorylation of Rb and E2F1 expression. Among I. obliquus fractions, fraction 2 (fractionated by dichloromethane from EEIO) showed the same effect as EEIO treatment on cell proliferation and cell cycle-related protein levels. CONCLUSIONS These results demonstrate that fraction 2 is the major fraction that induces G1 arrest and inhibits cell proliferation, suggesting I. obliquus could be used as a natural anti-cancer ingredient in the food and/or pharmaceutical industry. PMID:25861415

  10. An APC/C-Cdh1 Biosensor Reveals the Dynamics of Cdh1 Inactivation at the G1/S Transition

    PubMed Central

    Ondracka, Andrej; Robbins, Jonathan A.; Cross, Frederick R.

    2016-01-01

    B-type cyclin-dependent kinase activity must be turned off for mitotic exit and G1 stabilization. B-type cyclin degradation is mediated by the anaphase-promoting complex/cyclosome (APC/C); during and after mitotic exit, APC/C is dependent on Cdh1. Cdh1 is in turn phosphorylated and inactivated by cyclin-CDK at the Start transition of the new cell cycle. We developed a biosensor to assess the cell cycle dynamics of APC/C-Cdh1. Nuclear exit of the G1 transcriptional repressor Whi5 is a known marker of Start; APC/C-Cdh1 is inactivated 12 min after Whi5 nuclear exit with little measurable cell-to-cell timing variability. Multiple phosphorylation sites on Cdh1 act in a redundant manner to repress its activity. Reducing the number of phosphorylation sites on Cdh1 can to some extent be tolerated for cell viability, but it increases variability in timing of APC/C-Cdh1 inactivation. Mutants with minimal subsets of phosphorylation sites required for viability exhibit striking stochasticity in multiple responses including budding, nuclear division, and APC/C-Cdh1 activity itself. Multiple cyclin-CDK complexes, as well as the stoichiometric inhibitor Acm1, contribute to APC/C-Cdh1 inactivation; this redundant control is likely to promote rapid and reliable APC/C-Cdh1 inactivation immediately following the Start transition. PMID:27410035

  11. G1 cell cycle arrest and apoptosis induction by nuclear Smad4/Dpc4: Phenotypes reversed by a tumorigenic mutation

    PubMed Central

    Dai, Jia Le; Bansal, Ravi K.; Kern, Scott E.

    1999-01-01

    The tumor suppressor Smad4/Dpc4 is a transcription activator that binds specific DNA sequences and whose nuclear localization is induced after exposure to type β transforming growth factor-like cytokines. We explored an inducible system in which Smad4 protein is activated by translocation to the nucleus when cell lines that stably express wild-type or mutant Smad4 proteins fused to a murine estrogen receptor domain are treated with 4-hydroxytamoxifen. This induced Smad4-mediated transcriptional activation and a decrease in growth rate, attributable to a cell cycle arrest at the G1 phase and an induction of apoptosis. A tumor-derived mutation (Arg-100 → Thr) affecting a residue critical for DNA-binding demonstrated an “oncogenic” phenotype, having decreases in both the G1 fraction and apoptosis and, consequently, an augmentation of population growth. This model should be useful in the exploration and control of components that lie further downstream in the Smad4 tumor-suppressor pathway. PMID:9990040

  12. An APC/C-Cdh1 Biosensor Reveals the Dynamics of Cdh1 Inactivation at the G1/S Transition.

    PubMed

    Ondracka, Andrej; Robbins, Jonathan A; Cross, Frederick R

    2016-01-01

    B-type cyclin-dependent kinase activity must be turned off for mitotic exit and G1 stabilization. B-type cyclin degradation is mediated by the anaphase-promoting complex/cyclosome (APC/C); during and after mitotic exit, APC/C is dependent on Cdh1. Cdh1 is in turn phosphorylated and inactivated by cyclin-CDK at the Start transition of the new cell cycle. We developed a biosensor to assess the cell cycle dynamics of APC/C-Cdh1. Nuclear exit of the G1 transcriptional repressor Whi5 is a known marker of Start; APC/C-Cdh1 is inactivated 12 min after Whi5 nuclear exit with little measurable cell-to-cell timing variability. Multiple phosphorylation sites on Cdh1 act in a redundant manner to repress its activity. Reducing the number of phosphorylation sites on Cdh1 can to some extent be tolerated for cell viability, but it increases variability in timing of APC/C-Cdh1 inactivation. Mutants with minimal subsets of phosphorylation sites required for viability exhibit striking stochasticity in multiple responses including budding, nuclear division, and APC/C-Cdh1 activity itself. Multiple cyclin-CDK complexes, as well as the stoichiometric inhibitor Acm1, contribute to APC/C-Cdh1 inactivation; this redundant control is likely to promote rapid and reliable APC/C-Cdh1 inactivation immediately following the Start transition.

  13. Histone deacetylase enzyme silencing using shRNAs enhances radiosensitivity of SW579 thyroid cancer cells

    PubMed Central

    Wang, Ye; Jin, Tao; Dai, Xueming; Yan, Dongwang; Peng, Zhihai

    2016-01-01

    The aim of the present study was to screen the enzymes that are associated with the radiosensitivity of SW579 thyroid cancer cells, and investigate whether radiation, combined with specific RNA interference on the screened enzymes, enhances radiosensitivity of SW579 thyroid cancer cells. Quantitative polymerase chain reaction (qPCR) was used to analyze epigenetic enzyme expression changes before and after radiotherapy, and four enzymes, histone deacetylase 1 (HDAC1), HDAC2, HDAC4 and HDAC6 were screened. Western blot analysis was performed to analyze the change in HDAC1, HDAC2, HDAC4 and HDAC6 protein expression following radiotherapy. Short hairpin RNA (ShRNA)-HDAC1, shRNA-HDAC2, shRNA-HDAC4 and shRNA-HDAC6 plasmids were constructed and SW579 cells were transfected with corresponding shRNA-HDACs. Reverse transcription-qPCR was used to detect whether downregulation of HDAC mRNAs had been effective. In addition, shRNA and shRNA negative control (NC) pools were established and transfected into the SW579 cells. The samples were divided into four groups; control, trichostatin A, shRNA pool and shRNA NC pool, to analyze the effective enhancement of specific shRNA on radiosensitivity in thyroid cancer cells. The morphological changes were observed in the SW579 cells, and the number of tumor cells decreased markedly in the shRNA pool group compared with that of the other three groups. Therefore, it was concluded that HDACs present a potential target for increasing the sensitivity of thyroid cancer cells to radiotherapy, and shRNA-HDAC interference combined with radiotherapy promotes the radiosensitivity of tumors. PMID:27600599

  14. Histone Deacetylase Inhibitor SAHA Attenuates TLR4 Signaling in LPS-Stimulated Mouse Macrophages

    PubMed Central

    Chong, Wei; Li, Yongqing; Liu, Baoling; Zhao, Ting; Fukudome, Eugene Y.; Liu, Zhengcai; Smith, William M.; Velmahos, George C.; deMoya, Marc; Alam, Hasan B.

    2012-01-01

    Objective We have previously demonstrated that pre- and post-treatment of animals with suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor (HDACI), can improve survival in a mouse model of lipopolysaccharides (LPS)-induced severe shock. This study was to assess whether SAHA affects LPS/Toll like receptor4 (TLR4) signaling through acetylation of HSP90 and degradation of its client protein interleukin-1 receptor associated kinase 1 (IRAK1). Methods and Results RAW264.7 cells were exposed to LPS (1 μg/ml) for two hours followed by treatment with SAHA (10 μM) or one of HSP90 inhibitors, geldanamycin (GA) (3 μM). Sham (no SAHA, no LPS) macrophages served as a control. The cells were harvested at different time points, and time zero served as the reference point. LPS dramatically increased protein expression of myeloid differentiation factor 88 (MyD88) and IRAK1, and stimulated nuclear translocation of nuclear factor kB (NF-kB), leading to increases of gene expression and protein production of TNF-α and IL-6. Treatment with SAHA significantly attenuated these LPS- stimulated alterations. LPS or SAHA did not change the levels of HSP90 protein, but immunoprecipitation studies demonstrated that SAHA treatment enhanced acetylation of HSP90, and increased the dissociation of IRAK1, compared to the LPS control. Conclusions SAHA suppresses LPS/TLR4 signaling in LPS-stimulated macrophages through multiple possible mechanisms. It inhibits the function of HSP90 through hyperacetylation of the chaperone protein, which results in dissociation and degradation of the client protein IRAK1 and, at least in part, leads to a resultant decrease in nuclear translocation of NF-κB and attenuation of key pro-inflammatory cytokine expression. PMID:22868051

  15. Class IIa histone deacetylases affect neuronal remodeling and functional outcome after stroke.

    PubMed

    Kassis, Haifa; Shehadah, Amjad; Li, Chao; Zhang, Yi; Cui, Yisheng; Roberts, Cynthia; Sadry, Neema; Liu, Xianshuang; Chopp, Michael; Zhang, Zheng Gang

    2016-06-01

    We have previously demonstrated that stroke induces nuclear shuttling of class IIa histone deacetylase 4 (HDAC4). Stroke-induced nuclear shuttling of HDAC4 is positively and significantly correlated with improved indices of neuronal remodeling in the peri-infarct cortex. In this study, using a rat model for middle cerebral artery occlusion (MCAO), we tested the effects of selective inhibition of class IIa HDACs on functional recovery and neuronal remodeling when administered 24hr after stroke. Adult male Wistar rats (n = 15-17/group) were subjected to 2 h MCAO and orally gavaged with MC1568 (a selective class IIa HDAC inhibitor), SAHA (a non-selective HDAC inhibitor), or vehicle-control for 7 days starting 24 h after MCAO. A battery of behavioral tests was performed. Lesion volume measurement and immunohistochemistry were performed 28 days after MCAO. We found that stroke increased total HDAC activity in the ipsilateral hemisphere compared to the contralateral hemisphere. Stroke-increased HDAC activity was significantly decreased by the administration of SAHA as well as by MC1568. However, SAHA significantly improved functional outcome compared to vehicle control, whereas selective class IIa inhibition with MC1568 increased mortality and lesion volume and did not improve functional outcome. In addition, MC1568 decreased microtubule associated protein 2 (MAP2, dendrites), phosphorylated neurofilament heavy chain (pNFH, axons) and myelin basic protein (MBP, myelination) immunoreactivity in the peri-infarct cortex. Quantitative RT-PCR of cortical neurons isolated by laser capture microdissection revealed that MC1568, but not SAHA, downregulated CREB and c-fos expression. Additionally, MC1568 decreased the expression of phosphorylated CREB (active) in neurons. Taken together, these findings demonstrate that selective inhibition of class IIa HDACs impairs neuronal remodeling and neurological outcome. Inactivation of CREB and c-fos by MC1568 likely contributes to

  16. Histone deacetylase activators: N-acetylthioureas serve as highly potent and isozyme selective activators for human histone deacetylase-8 on a fluorescent substrate.

    PubMed

    Singh, Raushan K; Mandal, Tanmay; Balsubramanian, Narayanaganesh; Viaene, Tajae; Leedahl, Travis; Sule, Nitesh; Cook, Gregory; Srivastava, D K

    2011-10-01

    We report, for the first time, that certain N-acetylthiourea derivatives serve as highly potent and isozyme selective activators for the recombinant form of human histone deacetylase-8 in the assay system containing Fluor-de-Lys as a fluorescent substrate. The experimental data reveals that such activating feature is manifested via decrease in the K(m) value of the enzyme's substrate and increase in the catalytic turnover rate of the enzyme.

  17. The H,G_1,G_2 photometric system with scarce observational data

    NASA Astrophysics Data System (ADS)

    Penttilä, A.; Granvik, M.; Muinonen, K.; Wilkman, O.

    2014-07-01

    The H,G_1,G_2 photometric system was officially adopted at the IAU General Assembly in Beijing, 2012. The system replaced the H,G system from 1985. The 'photometric system' is a parametrized model V(α; params) for the magnitude-phase relation of small Solar System bodies, and the main purpose is to predict the magnitude at backscattering, H := V(0°), i.e., the (absolute) magnitude of the object. The original H,G system was designed using the best available data in 1985, but since then new observations have been made showing certain features, especially near backscattering, to which the H,G function has troubles adjusting to. The H,G_1,G_2 system was developed especially to address these issues [1]. With a sufficient number of high-accuracy observations and with a wide phase-angle coverage, the H,G_1,G_2 system performs well. However, with scarce low-accuracy data the system has troubles producing a reliable fit, as would any other three-parameter nonlinear function. Therefore, simultaneously with the H,G_1,G_2 system, a two-parameter version of the model, the H,G_{12} system, was introduced [1]. The two-parameter version ties the parameters G_1,G_2 into a single parameter G_{12} by a linear relation, and still uses the H,G_1,G_2 system in the background. This version dramatically improves the possibility to receive a reliable phase-curve fit to scarce data. The amount of observed small bodies is increasing all the time, and so is the need to produce estimates for the absolute magnitude/diameter/albedo and other size/composition related parameters. The lack of small-phase-angle observations is especially topical for near-Earth objects (NEOs). With these, even the two- parameter version faces problems. The previous procedure with the H,G system in such circumstances has been that the G-parameter has been fixed to some constant value, thus only fitting a single-parameter function. In conclusion, there is a definitive need for a reliable procedure to produce

  18. Regulation of cyclin-dependent kinase (Cdk) 2 Thr-160 phosphorylation and activity by mitogen-activated protein kinase in late G1 phase.

    PubMed Central

    Chiariello, M; Gomez, E; Gutkind, J S

    2000-01-01

    Mitogen-activated protein (MAP) kinases, p42(MAPK) and p44(MAPK), are central components of growth-promoting signalling pathways. However, how stimulation of MAP kinases culminates in cell-cycle progression is still poorly understood. Here we show that mitogenic stimulation of NIH 3T3 cells causes a sustained activation of MAP kinases, which lasts until cells begin progressing through the G(1)/S boundary. Furthermore, we observed that disruption of the MAP-kinase pathway with a selective MEK (MAP kinase/extracellular-signal-regulated protein kinase kinase) inhibitor, PD98059, prevents the activation of cyclin-dependent kinase (Cdk) 2 and DNA synthesis, even when added during late G(1) phase, once the known mechanisms by which MAP kinase controls G(1) progression, accumulation of G(1) cyclins and degradation of Cdk inhibitors have already taken place. Moreover, we provide evidence indicating that MAP kinases control Cdk2 Thr-160 activating phosphorylation and function, possibly by regulating the activity of a Cdk-activating kinase, thus promoting the re-initiation of DNA synthesis. These findings suggest the existence of a novel mechanism whereby signal-transducing pathways converging on MAP kinases can affect the cell-cycle machinery and, ultimately, participate in cell-growth control. PMID:10903150

  19. Study of the genetic variability in a Parkinson's Disease gene: EIF4G1.

    PubMed

    Tucci, Arianna; Charlesworth, Gavin; Sheerin, Una-Marie; Plagnol, Vincent; Wood, Nicholas W; Hardy, John

    2012-06-14

    Chartier-Harlin and colleagues [2] recently reported mutations in the eukaryotic translation initiation factor 4-gamma (EIF4G1) gene in families with parkinsonism. Large-scale screening found two mutations (p.R1205H and p.A502V) only in affected individuals, although their relative frequency was very low. The aim of this study was to investigate EIF4G1 parkinsonism-related variants in two separate cohorts and study coding variability across the gene. We first screened a series of familial Parkinson's Disease (PD) patients in an attempt to confirm previous results by showing segregation. Then, to determine the extent of coding variation in the gene, we first screened a cohort of sub-Saharan African individuals from the Centre d'Etude du Polymorphisme Humain - Human Genome Diversity Cell Line Panel (HGDP) [1] and then analyzed data from 5350 individuals National Heart, Lung, and Blood Institute (NHLBI) exome sequencing project. We failed to identify any PD-related mutations in the familial samples. Conversely we found the p.A502V variant in the NHLBI population. We observed a high number of coding polymorphism in the exons where the two PD variants have been previously reported. We conclude that either EIF4G1 variants are an extremely rare cause of familial PD in Caucasian cohorts, or that A502V is in fact a rare benign variant not involved in PD aetiology. Our data also suggests that the protein can tolerate some extent of variability particularly at this point of the gene.

  20. AAF G1 and Millimeter Wavelength ARM Radar Observations and Analysis from the ACAPEX Field Campaign

    NASA Astrophysics Data System (ADS)

    Matthews, A. A.; Hardin, J. C.; Comstock, J. M.; Bharadwaj, N.; Mei, F.

    2015-12-01

    From late January to early March of 2015, the ARM Cloud Aerosol Precipitation Experiment (ACAPEX) deployed a variety of instruments to study atmospheric rivers as they made landfall on the western United States. These atmospheric rivers are an important source of rainfall for the drought-ridden regions of California, so it is important to understand the cloud characteristics and processes that occur during the events. For this study, the KAZR zenith pointing millimeter wavelength radar aboard the Ron Brown research vessel as well as two cloud probes, namely the Two-Dimensional Stereo Probe (2DS) and High Volume Precipitation Spectrometer-3 (HVPS3), flown on the ARM Aerial Facility Gulfstream-1 (AAF G1) aircraft were used to analyze the clouds associated with the atmospheric rivers. PyDisdrometer, an open-source software, is used to calculate radar reflectivity values based on the drop size distributions observed by the cloud probes so that the aircraft data may be directly compared to the KAZR data. This research will focus on two flight days. First, a coordinated flight flown on February 5, where the G1 flew a spiral pattern over the Ron Brown while the KAZR was running will be used to examine the clouds over the ocean. This will also allow for a direct comparison between the calculated radar reflectivity from the cloud probes and the KAZR measured reflectivity. The second case was chosen to be the atmospheric river landfall event on February 6. This case will be analyzed over the ocean using the KAZR, then over the shoreline, Sacramento, and the inland mountains using the cloud probe data, as well as other instrumentations onboard the G1, such as those that measure cloud thickness and liquid water content. In this way, the changes that occur to the clouds as they move from open-ocean to land may be examined in greater detail.