A comparison of two assessment tools used in overviews of systematic reviews: ROBIS versus AMSTAR-2.

BACKGROUND: AMSTAR-2 is a 16-item assessment tool to check the quality of a systematic review and establish whether the most important elements are reported. ROBIS is another assessment tool which was designed to evaluate the level of bias present within a systematic review. Our objective was to compare, contrast and establish both inter-rater reliability and usability of both tools as part of two overviews of systematic reviews. Strictly speaking, one tool assesses methodological quality (AMSTAR-2) and the other assesses risk of bias (ROBIS), but there is considerable overlap between the tools in terms of the signalling questions. METHODS: Three reviewers independently assessed 31 systematic reviews using both tools. The inter-rater reliability of all sub-sections using each instrument (AMSTAR-2 and ROBIS) was calculated using Gwet's agreement coefficient (AC1 for unweighted analysis and AC2 for weighted analysis). RESULTS: Thirty-one systematic reviews were included. For AMSTAR-2, the median agreement for all questions was 0.61. Eight of the 16 AMSTAR-2 questions had substantial agreement or higher (> 0.61). For ROBIS, the median agreement for all questions was also 0.61. Eleven of the 24 ROBIS questions had substantial agreement or higher. CONCLUSION: ROBIS is an effective tool for assessing risk of bias in systematic reviews and AMSTAR-2 is an effective tool at assessing quality. The median agreement between raters for both tools was identical (0.61). Reviews that included a meta-analysis were easier to rate with ROBIS; however, further developmental work could improve its use in reviews without a formal synthesis. AMSTAR-2 was more straightforward to use; however, more response options would be beneficial.

A survey of helminth control practices on sheep farms in Great Britain and Ireland.

A telephone survey of 600 farmers throughout Great Britain and Ireland was conducted in order to characterise helminth control practices, and identify factors correlated with perceived anthelmintic failure. Overall, 93% of surveyed farmers routinely treated their sheep against nematodes, 67% against liver fluke and 58% against tapeworms. Anthelmintic resistance in nematodes was perceived by farmers to be present on 10% of farms. Farmers who dosed ewes at mating were more likely to have observed anthelmintic failure, than those who were aware of national guidelines on parasite control. However, objective assessment of anthelmintic efficacy had only been undertaken on 19% of farms. Ewes were treated at mating and lambing on 63% and 62% of farms, respectively. On average, lambs were treated 3.6 times annually, depending on geographical region and on dates of lambing and finishing. Although 'quarantine' treatments were widely administered to bought-in stock, these were appropriately applied in only 3% of cases. This study provides baseline data against which the impact of future anthelmintic information campaigns can be assessed; it will facilitate the development of rational, farm-level mathematical models in support of sustainable parasite control, and will aid in the design of farm management practices that prolong the effective lifespan of novel classes of anthelmintic.

JIP1 binding to RBP-Jk mediates cross-talk between the Notch1 and JIP1-JNK signaling pathway.

Notch1 signaling has a critical function in maintaining a balance among cell proliferation, differentiation, and apoptosis. Our earlier work showed that the Notch1 intracellular domain interferes with the scaffolding function of c-Jun N-terminal kinase (JNK)-interacting protein-1 (JIP1), yet the effect of JIP1 for Notch1-recombining binding protein suppressor of hairless (RBP-Jk) signaling remains unknown. Here, we show that JIP1 suppresses Notch1 activity. JIP1 was found to physically associate with either intracellular domain of Notch1 or RBP-Jk and interfere with the interaction between them. Furthermore, we ascertained that JIP1 caused the cytoplasmic retention of RBP-Jk through an interaction between the C-terminal region of JIP1 including Src homology 3 domain and the proline-rich domain of RBP-Jk. We also found that RBP-Jk inhibits JIP1-mediated activation of the JNK1 signaling cascade and cell death. Our results suggest that direct protein-protein interactions coordinate cross-talk between the Notch1-RBP-Jk and JIP1-JNK pathways.

Number and ownership profiles of cats and dogs in the UK.

A random sample of 2980 households in the UK in 2007 showed that 26 per cent and 31 per cent of households owned cats and dogs, respectively. Households with gardens were more likely to own cats and dogs than households without gardens. Households in which someone was qualified to degree level were more likely to own cats and less likely to own dogs than other households. Cats were more likely to be owned by semi-urban/rural households and by female respondents. Dog ownership significantly decreased the likelihood of cat ownership, and respondents aged 65 years or more were less likely to report that their household owned a cat than younger respondents. Households with one or more dogs and children aged 11 to 15 years were more likely to own a cat than other households. The likelihood of dog ownership increased as household size increased. Dogs were more likely to be owned by rural households, and less likely to be owned by households with cats or children aged 10 years or younger. Female respondents and those aged less than 55 years were more likely to report dog ownership than other respondents. The estimated size (and 95 per cent confidence intervals) of the owned cat and dog populations in the UK in 2006 was 10,332,955 (9,395,642 to 11,270,269) cats and 10,522,186 (9,623,618 to 11,420,755) dogs.

Survey of the characteristics of cats owned by households in the UK and factors affecting their neutered status.

Data were collected on 1260 cats owned by a random sample of UK households in 2007, by means of telephone questionnaires, which were completed by 33 per cent of the households contacted. Cats were owned by 26 per cent of these households, 42 per cent of which owned more than one cat. The owners of 622 female cats reported a total of 110 unplanned litters of kittens. Excluding cats with an unknown neutered status 92 per cent of the cats aged six months or more were neutered, but only 66 per cent of the cats aged six to 12 months were neutered. Multivariable logistic regression, based on data collected for one randomly selected cat aged four months or more in each cat-owning household, was used to identify factors affecting the cats' neutered status. Analysis of 48 sexually entire cats and 501 neutered cats showed that cats aged 10 months or more, with indoor access, that had been vaccinated within the last year or were currently registered with a veterinary practice, were more likely to be neutered than cats aged less than 10 months, without indoor access, that had been vaccinated more than one year ago or were not registered with a veterinary practice.

Role of mitogen-activated protein kinase kinase 4 in cancer.

Mitogen-activated protein (MAP) kinase kinase 4 (MKK4) is a component of stress activated MAP kinase signaling modules. It directly phosphorylates and activates the c-Jun N-terminal kinase (JNK) and p38 families of MAP kinases in response to environmental stress, pro-inflammatory cytokines and developmental cues. MKK4 is ubiquitously expressed and the targeted deletion of the Mkk4 gene in mice results in early embryonic lethality. Further studies in mice have indicated a role for MKK4 in liver formation, the immune system and cardiac hypertrophy. In humans, it is reported that loss of function mutations in the MKK4 gene are found in approximately 5% of tumors from a variety of tissues, suggesting it may have a tumor suppression function. Furthermore, MKK4 has been identified as a suppressor of metastasis of prostate and ovarian cancers. However, the role of MKK4 in cancer development appears complex as other studies support a pro-oncogenic role for MKK4 and JNK. Here we review the biochemical and functional properties of MKK4 and discuss the likely mechanisms by which it may regulate the steps leading to the formation of cancers.

The JIP family of MAPK scaffold proteins.

The components of MAPK (mitogen-activated protein kinase) signalling pathways can assemble into complexes that are co-ordinated by regulatory proteins including scaffold proteins. There is increasing evidence that scaffold proteins (i) maintain signalling specificity and facilitate the activation of pathway components, (ii) localize pathway components to particular subcellular sites or to specific targets, and (iii) serve as a point of signal integration to allow regulation of MAPK pathways by other signalling events in the cell. One family of scaffold proteins that regulate signalling by stress-activated MAPKs are the JIPs [JNK (c-Jun N-terminal kinase)-interacting proteins]. JIP proteins have been demonstrated to form complexes with specific JNK and p38 MAPK signalling modules and to play important roles in brain development, neuronal trafficking, apoptosis, beta-cell function and insulin responses. Here, I briefly review our current understanding of the biochemical properties and physiological roles of JIP proteins.

Requirement of the JIP1 scaffold protein for stress-induced JNK activation.

The c-Jun N-terminal kinase (JNK) signal transduction pathway is activated in response to the exposure of cells to environmental stress. Components of the JNK signaling pathway interact with the JIP1 scaffold protein. JIP1 is located in the neurites of primary hippocampal neurons. However, in response to stress, JIP1 accumulates in the soma together with activated JNK and phosphorylated c-Jun. Disruption of the Jip1 gene in mice by homologous recombination prevented JNK activation caused by exposure to excitotoxic stress and anoxic stress in vivo and in vitro. These data show that the JIP1 scaffold protein is a critical component of a MAP-kinase signal transduction pathway.

Regulation of transcription factor function by phosphorylation.

Changes in protein phosphorylation represent a mechanism that is frequently employed by cells to regulate transcription factor activity. In response to alterations in the extracellular environment, signal transduction pathways target transcription factors, transcriptional coregulators and chromatin-modifying factors, leading to their phosphorylation by protein kinases or dephosphorylation by protein phosphatases. These modifications either positively or negatively regulate transcription factor activity to facilitate a program of gene expression that results in appropriate changes in cell behavior. Protein phosphorylation and dephosphorylation can directly regulate distinct aspects of transcription factor function, including cellular localization, protein stability, protein-protein interactions and DNA binding. The phosphorylation-dependent modulation of the activities of transcriptional coregulators and chromatin-modifying factors can also control transcription factor activity. Here we review recent studies that have led to a better understanding of the mechanisms by which protein phosphorylation and dephosphorylation governs transcription factor function.

JNK is required for effector T-cell function but not for T-cell activation.

The hallmark of T-cell activation is the production of interleukin 2 (IL-2). c-Jun amino-terminal kinase (JNK), a MAP kinase that phosphorylates c-Jun and other components of the AP-1 group of transcription factors, has been implicated in the activation of IL-2 expression. Previously, we found that T cells from mice deficient in the Jnk1 or Jnk2 gene can be activated and produce IL-2 normally, but are deficient in functional differentiation into Th1 or Th2 subsets. However, studies of mice with compound mutations indicate that JNK1 and JNK2 are redundant during mouse development. Here we use three new mouse models in which peripheral T cells completely lack JNK proteins or signalling, to test whether the JNK signalling pathway is crucial for IL-2 expression and T-cell activation. Unexpectedly, these T cells made more IL-2 and proliferated better than wild-type cells. However, production of effector T-cell cytokines did require JNK. Thus, JNK is necessary for T-cell differentiation but not for naive T-cell activation.

Conference highlight: do T cells care about the mitogen-activated protein kinase signalling pathways?

Mitogen-activated protein (MAP) kinases, which include the extracellular response kinases, p38 and c-Jun amino terminal kinases (JNK), play a significant role in mediating signals triggered by cytokines, growth factors and environmental stress. The JNK and p38 MAP kinases have been involved in growth, differentiation and cell death in different cell types. In the present paper, we describe how the JNK and p38 MAP kinase signalling pathways are regulated and their role during thymocyte development and the activation and differentiation of T cells in the peripheral immune system. The results from these studies demonstrate that the JNK and p38 MAP kinase signalling pathways regulate different aspects of T-cell mediated immune responses.

Regulation of c-Jun NH(2)-terminal kinase (Jnk) gene expression during T cell activation.

The c-Jun NH(2)-terminal kinases (JNKs) are a group of mitogen-activated protein (MAP) kinases that participate in signal transduction events mediating specific cellular functions. Activation of JNK is regulated by phosphorylation in response to cellular stress and inflammatory cytokines. Here, we demonstrate that JNK is regulated by a second, novel mechanism. Induction of Jnk gene expression is required in specific tissues before activation of this signaling pathway. The in vivo and in vitro ligation of the T cell receptor (TCR) leads to induction of JNK gene and protein expression. TCR signals are sufficient to induce JNK expression, whereas JNK phosphorylation also requires CD28-mediated costimulatory signals. Therefore, both expression and activation contribute to the regulation of the JNK pathway to ensure proper control during the course of an immune response.

The JIP group of mitogen-activated protein kinase scaffold proteins.

Activation of the c-Jun NH(2)-terminal kinase (JNK) group of mitogen-activated protein (MAP) kinases is mediated by a protein kinase cascade. This signaling mechanism may be coordinated by the interaction of components of the protein kinase cascade with scaffold proteins. The JNK-interacting protein (JIP) group of scaffold proteins selectively mediates signaling by the mixed-lineage kinase (MLK)-->MAP kinase kinase 7 (MKK7)-->JNK pathway. The scaffold proteins JIP1 and JIP2 interact to form oligomeric complexes that accumulate in peripheral cytoplasmic projections extended at the cell surface. The JIP proteins function by aggregating components of a MAP kinase module (including MLK, MKK7, and JNK) and facilitate signal transmission by the protein kinase cascade.

Mechanism-based inhibition of C5-cytosine DNA methyltransferases by 2-H pyrimidinone.

DNA duplexes in which the target cytosine base is replaced by 2-H pyrimidinone have previously been shown to bind with a significantly greater affinity to C5-cytosine DNA methyltransferases than unmodified DNA. Here, it is shown that 2-H pyrimidinone, when incorporated into DNA duplexes containing the recognition sites for M.HgaI-2 and M.MspI, elicits the formation of inhibitory covalent nucleoprotein complexes. We have found that although covalent complexes are formed between 2-H pyrimidinone-modified DNA and both M.HgaI-2 and M.MspI, the kinetics of complex formation are quite distinct in each case. Moreover, the formation of a covalent complex is still observed between 2-H pyrimidinone DNA and M.MspI in which the active-site cysteine residue is replaced by serine or threonine. Covalent complex formation between M.MspI and 2-H pyrimidinone occurs as a direct result of nucleophilic attack by the residue at the catalytic position, which is enhanced by the absence of the 4-amino function in the base. The substitution of the catalytic cysteine residue by tyrosine or chemical modification of the wild-type enzyme with N-ethylmaleimide, abolishes covalent interaction. Nevertheless the 2-H pyrimidinone-substituted duplex still binds to M.MspI with a greater affinity than a standard cognate duplex, since the 2-H pyrimidinone base is mis-paired with guanine.

c-Jun is a JNK-independent coactivator of the PU.1 transcription factor.

The ETS domain transcription factor PU.1 is necessary for the development of monocytes and regulates, in particular, the expression of the monocyte-specific macrophage colony-stimulating factor (M-CSF) receptor, which is critical for monocytic cell survival, proliferation, and differentiation. The bZIP transcription factor c-Jun, which is part of the AP-1 transcription factor complex, is also important for monocytic differentiation, but the monocyte-specific M-CSF receptor promoter has no AP-1 consensus binding sites. We asked the question of whether c-Jun could promote the induction of the M-CSF receptor by collaborating with PU.1. We demonstrate that c-Jun enhances the ability of PU.1 to transactivate the M-CSF receptor promoter as well as a minimal thymidine kinase promoter containing only PU.1 DNA binding sites. c-Jun does not directly bind to the M-CSF receptor promoter but associates via its basic domain with the ETS domain of PU.1. Consistent with our observation that AP-1 binding does not contribute to c-Jun coactivation is the observation that the activation of PU.1 by c-Jun is blocked by overexpression of c-Fos. Phosphorylation of c-Jun by c-Jun NH2-terminal kinase on Ser-63 and -73 does not alter the ability of c-Jun to enhance PU.1 transactivation. Activated Ras enhances the transcriptional activity of PU.1 by up-regulating c-Jun expression without changing the phosphorylation pattern of PU.1. The activation of PU.1 by Ras is blocked by a mutant c-Jun protein lacking the basic domain. The expression of this mutant form of c-Jun also completely blocks 12-O-tetradecanoylphorbol-13-acetate-induced M-CSF receptor promoter activity during monocytic differentiation. We propose therefore that c-Jun acts as a c-Jun NH2-terminal kinase-independent coactivator of PU.1, resulting in M-CSF receptor expression and development of the monocytic lineage.

The MKK7 gene encodes a group of c-Jun NH2-terminal kinase kinases.

The c-Jun NH2-terminal protein kinase (JNK) is a member of the mitogen-activated protein kinase (MAPK) group and is an essential component of a signaling cascade that is activated by exposure of cells to environmental stress. JNK activation is regulated by phosphorylation on both Thr and Tyr residues by a dual-specificity MAPK kinase (MAPKK). Two MAPKKs, MKK4 and MKK7, have been identified as JNK activators. Genetic studies demonstrate that MKK4 and MKK7 serve nonredundant functions as activators of JNK in vivo. We report here the molecular cloning of the gene that encodes MKK7 and demonstrate that six isoforms are created by alternative splicing to generate a group of protein kinases with three different NH2 termini (alpha, beta, and gamma isoforms) and two different COOH termini (1 and 2 isoforms). The MKK7alpha isoforms lack an NH2-terminal extension that is present in the other MKK7 isoforms. This NH2-terminal extension binds directly to the MKK7 substrate JNK. Comparison of the activities of the MKK7 isoforms demonstrates that the MKK7alpha isoforms exhibit lower activity, but a higher level of inducible fold activation, than the corresponding MKK7beta and MKK7gamma isoforms. Immunofluorescence analysis demonstrates that these MKK7 isoforms are detected in both cytoplasmic and nuclear compartments of cultured cells. The presence of MKK7 in the nucleus was not, however, required for JNK activation in vivo. These data establish that the MKK4 and MKK7 genes encode a group of protein kinases with different biochemical properties that mediate activation of JNK in response to extracellular stimuli.

Signal transduction by MAP kinases: regulation by phosphorylation-dependent switches.

The kinases of mitogen-activated protein (MAP) kinase cascades transmit signals through sequential phosphorylation and activation of the enzymes. However, recent evidence indicates that protein-protein interactions between the kinases themselves or with substrates or other components are also a critical means of regulation. Whitmarsh and Davis summarize these findings with emphasis on new evidence from yeast that, when phosphorylated, a MAP kinase kinase actually switches from a negative regulator that binds to and inhibits its target MAP kinase to a positive regulator of that same enzyme.