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1.
Mark T Uhlik Amy N Abell Bruce D Cuevas Kazuhiro Nakamura Gary L Johnson 《Biochimie et biologie cellulaire》2004,82(6):658-663
Mitogen-activated protein kinase (MAPK) pathways are activated by a plethora of stimuli. The literature is filled with papers describing the activation of different MAPKs by almost any stimulus or insult imaginable to cells. In this review, we use signal transduction wiring diagrams to illustrate putative upstream regulators for the MAPK kinase kinases, MEKK1, 2, and 3. Targeted gene disruption of MEKK1, 2, or 3 defined phenotypes for each MEKK associated with loss of specific MAPK regulation. Genetic analysis of MEKK function clearly defines specific components of the wiring diagram that require MEKK1, 2, or 3 for physiological responses. We propose that signal transduction network wiring diagrams are valuable tools for hypothesis building and filtering physiologically relevant phenotypic responses from less connected protein relations in the regulation of MAPK pathways. 相似文献
2.
Richard Stouthamer Robert F. Luck John D. Pinto Gary R. Platner Beth Stephens 《Entomologia Experimentalis et Applicata》1996,80(3):481-489
In non-reciprocal cross-incompatibility (NRCI), the crossing of a female of a strain A with a male of a strain B results in hybrid offspring, whereas the reciprocal cross produces few or no hybrids. Only females are of hybrid origin in Hymenoptera because they arise from fertilized eggs; males arise from unfertilized (haploid) eggs. Crosses between many strains of Trichogramma deion showed some degree of NRCI. Crosses between a T. deion culture collected in Seven Pines, California (SVP) with one from Marysville, California (MRY) showed an extreme form of NRCI in which practically no female offspring was produced when MRY females were crossed with SVP males. The reciprocal cross produced a close to normal proportion of female and male offspring. Detailed studied of this cross indicated that 1) the female offspring produced in the compatible interstrain cross were not the result of parthenogenesis but were true hybrids, 2) the incompatible interstrain cross did not produce female offspring because fertilized eggs died during development, 3) the death of these eggs could not be prevented by either antibiotic or temperature treatment, 4) cytoplasmically inherited factors causing NRCI could be discounted because backcrossed females with the genome of MRY and the cytoplasm of SVP, exhibit the NRCI relationship characteristic of their genome. Therefore the NRCI between these strains appears to be caused by a modification coded for by the nuclear genes of MRY that results in incompatibility when SVP sperm fertilizes MRY eggs. In addition the level of incompatibility in crosses between the SVP females and MRY males is temperature sensitive, the higher the rearing temperature the lower the level of compatibility. 相似文献
3.
Carol J. Maslansky Gary M. Williams 《In vitro cellular & developmental biology. Plant》1982,18(8):683-693
Summary Hepatocyte primary cultures (HPC) derived from rat, mouse, hamster, and rabbit liver were characterized for a variety of parameters.
The conditions that maximized recovery, attachment, and survival varied between species. Hepatocytes from all four species
were capable of attaching in serum-free Williams’ medium E (WME), but optimal attachment as monolayer cultures was achieved
for mouse and hamster HPC in medium receiving 1% calf serum supplementation. Hamster hepatocytes required additional cations,
whereas rabbit and rat hepatocytes displayed maximal attachment in medium supplemented with 10% calf serum. Survival of mouse
and rabbit hepatocytes after 24 h in serum supplemented media was in the order of 90%. Rat and hamster hepatocyte 24 h survival
was approximately 70 and 60%, respectively, and was not significantly affected by serum supplementation. Hepatocytes from
each species varied in their content of cytochromeP450 at the time of isolation and in the rate of reduction during culture. Mouse and rat hepatocytes demonstrated the most
rapid decline in content during the initial 24 h in culture, whereas concentrations in rabbit hepatocytes were virtually unchanged.
The rate of decline inP450 concentrations in hamster hepatocytes was intermediate between those displayed by rat and rabbit hepatocytes. These studies
have delineated conditions useful for the culture of hepatocytes from four species and have documented the status of an important
parameter of their functional capability.
This study was supported by EPA contract 68-01-6179. C. J. Maslansky was a recipient of a Monsanto Fund Fellowship in Toxicology. 相似文献
4.
Specific activities of eight enzymes involved in glycerol metabolism were determined in crude extracts of three strains ofNeurospora crassa after growth on six different carbon sources. One of the strains was wild type, which grew poorly on glycerol as sole carbon source; the other two were mutant strains which were efficient glycerol utilizers. A possible basis for this greater effeciency of glycerol utilization was catabolite repression of glyceraldehyde kinase by glycerol in wild type, and two-fold higher glycerate kinase activity in the mutant strains after growth on glycerol, thus apparently allowing two routes for glyceraldehyde to enter the glycolytic pathway in the mutant strains but only one in wild type. The preferential entry of glyceraldehyde to the glycolytic pathway through glycerate was suggested by the lack of glyceraldehyde kinase in all three strains after growth on one or more of the carbon sources and the generally higher levels of aldehyde dehydrogenase and of glycerate kinase than of glyceraldehyde kinase. 相似文献
5.
6.
Process for the isolation of preparative quantities of
Paul A. Kroon Maria T. Garcia Conesa Ian J. Colquhoun Gary Williamson 《Carbohydrate research》1997,300(4)
Specific enzymes were used to hydrolyse sugarbeet pulp to facilitate the isolation of
in preparative amounts. The feruloylated arabinose disaccharide was purified by chromatography on Sephadex LH-20 and Bio-Gel P-2 and the structure confirmed by NMR and UV spectroscopy and high-performance liquid chromatography. 相似文献
Full-size image
7.
Extremely high degrees of lead tolerance, measured by comparing rates of root extension in culture solutions, are reported from populations of Festuca ovina growing at two lead-mining sites (Westschacht and Keldenich-II) near Mechernich in the Eifel Mountains, Germany. Other populations from nearby heavy metal-contaminated areas show a considerably smaller degree of lead tolerance. Samples of Festuca ovina collected in the field at Westschacht and Keldenich-II contain higher levels of lead in their aerial organs than do those from other lead-contaminated sites. The main soil factor determining the high degree of lead tolerance is the high Pb/Ca ratio. Populations from soils with a low Pb/Ca ratio display a very low degree of tolerance. It is therefore concluded that in Westschacht and Keldenich-II plants, a genuine intracellular tolerance mechanism is present, allowing the accumulation of lead in aerial organs.Leaf samples of zinc-tolerant Festuca contain higher levels of zinc than do samples of non-tolerant plants. Lead and zinc amounts in leaves are correlated with the soil ratios of Pb/Ca and Zn/Ca, respectively, rather than with the absolute soil-metal levels.In a slightly lead-tolerant, but highly zinc-tolerant clone of Festuca ovina from a site contaminated with large amounts of lead and zinc (Plombières), lead was found to be the major factor affecting the inhibition of root extension with combined treatments of lead and zinc in culture solutions. In the highly lead-tolerant, zinc-sensitive population from Westschacht, zinc governs the response of root growth to combinations of the two metals. The results are discussed in terms of discriminating distinct types of heavy-metal tolerance. 相似文献
8.
Molecular and functional analysis of the ruv region of Escherichia coli K-12 reveals three genes involved in DNA repair and recombination 总被引：14，自引：0，他引：14
Gary J. Sharples Fiona E. Benson Graham T. Illing Robert G. Lloyd 《Molecular & general genetics : MGG》1990,221(2):219-226
Summary Recombinant plasmids carrying ruvA, ruvB, or both were constructed and used to investigate the genetic defects in a collection of UV-sensitive ruv mutants. The results revealed that efficient survival of UV-irradiated cells depends on both ruvA and ruvB, and on a third gene, ruvC, located upstream of the ruvAB operon. Southern blotting analysis was used to locate insertions in ruv and to examine putative deletion mutants. Two Tn10 insertions were located to the region encoding ruvA. Since these insertions caused a deficiency in the activities of both ruvA and ruvB, we concluded that they must exert a polar effect on ruvB. Two putative ruv deletion mutants were shown to be the result of deletion-inversion events mediated during imprecise excision of Tn10. The relevant inversion breakpoints in these mutants were located to ruvA and ruvC. 相似文献
9.
Mycorrhization helper bacteria, Paenibacillus sp. EJP73 and Burkholderia sp. EJP67, were used to study the importance of bacterial inoculum dose and bacterial derived soluble and volatile metabolites localization for enhancing mycorrhiza formation in the Pinus sylvestris-Lactarius rufus symbiosis, using a laboratory based microcosm. EJP73 and EJP67 produced different responses in relation to the inoculum dose; EJP73 significantly enhanced mycorrhiza formation to the same degree at all doses tested (10(5), 10(7), 10(9) and 10(10) CFU mL(-1)), whereas, EJP67 only stimulated mycorrhiza formation within a narrow range of inoculum densities (10(7) and 10(9) CFU mL(-1)). The importance of soluble bacterial metabolites was assessed by applying spent broth derived from exponential and stationary phase bacterial cultures to microcosms. No spent broth enhanced mycorrhiza formation over the control. As EJP73 produced the helper effect over a wide range of inoculum doses, this bacterium was chosen for further study. Physical separation of EJP73 from the fungal and plant symbiosis partners was carried out, in order to determine the contribution of constitutively produced bacterial volatile metabolites to the mycorrhization helper bacteria effect. When EJP73 was physically separated from the symbiosis, it had a significant negative effect on mycorrhiza formation. These results suggest that close proximity, or indeed cell contact, is required for the helper effect. Therefore, fluorescent in situ hybridization in conjunction with cryosectioning was used to determine the localization of EJP73 in mycorrhizal tissue. The cells were found to occur as rows or clusters ( approximately 10 cells) within the mycorrhizal mantle, both at the root tip and along the length of the mycorrhizal short roots. 相似文献
10.
Gary S Firestein 《Arthritis research & therapy》2014,16(3):114
Rheumatoid arthritis is a complex disease where predetermined and stochastic factors conspire to confer disease susceptibility. In light of the diverse responses to targeted therapies, rheumatoid arthritis might represent a final common clinical phenotype that reflects many pathogenic pathways. Therefore, it might be appropriate to begin thinking about rheumatoid arthritis as a syndrome rather than a disease. Use of genetics, epigenetics, microbiomics, and other unbiased technologies will probably permit stratification of patients based on mechanisms of disease rather than by clinical phenotype.Observer la nature, et suivez la route qu’elle vous trace.JJ Rousseau, quoted in [1].Over 150 years ago, Garrod coined the term ‘rheumatoid arthritis’ (RA) to distinguish it from other forms of arthritis, most notably gout and acute rheumatism [1]. Years later, disease subsets were further characterized based, in part, on clinical manifestations such as erosions and nodules or laboratory values such as autoantibodies in the blood. For instance, patients with rheumatoid factors and anti-citrullinated protein antibodies (ACPAs) tend to have more severe disease and worse long-term outcomes than seronegative patients.The broad range of genes associated with RA, the role of the environment in disease initiation, and the diversity of responses to targeted therapies necessitate a re-evaluation of time-honored stratification based on carefully documented clinical phenotypes. Moreover, we should reconsider whether RA should be viewed as the disease that Garrod described or whether it represents a final common pathway of divergent mechanisms in an organ (synovium) with a limited repertoire of responses. In this context, RA could be thought of as a syndrome with multiple etiologic events.RA susceptibility is determined, in part, by inherited risk factors that are predetermined. The single nucleotide polymorphisms (SNPs) associated with RA are dispersed widely across the genome, with notable concentration in genes that participate in adaptive and innate immune responses [2]. Multiple genome-wide association studies have identified scores of disease-associated SNPs. By far the greatest genetic risk is conferred by the class II major histocompatibility gene HLA-DR, which participates in antigen presentation to T lymphocytes [3]. The critical regions of the encoded protein have been well characterized and are located in and around the antigen-binding groove. However, the observation that identical twins only have perhaps a 15% concordance rate for RA indicates that inherited DNA sequences account for a minority of risk and might not be as important as other influences [4]. Put another way, full diploid genome sequencing of patients ignores over 80% of disease risk.Many SNPs outside the major histocompatibility complex also contribute to susceptibility, but their influence is much lower, with relative risks typically <1.2 [5]. One need not have all of these SNPs to develop RA; only a limited subset are probably needed in the presence of the proper environmental exposures. Individual and combinations of low-penetrance susceptibility genes have not offered major insights into the clinical phenotype, although it is still early days for these complex analyses. The fact that various combinations of genes and types of environmental stress lead to the same phenotype suggests that we are not looking at a single disease but at a process with multiple pathways.The “original sin” in ACPA-positive RA is probably due to an interaction between disease-associated HLA-DR genes and the environment, especially at mucosal surfaces (reviewed in [6]). The first steps could be viewed as a normal adaptive immune response against stress-induced modification of peptides, most notably by citrullination. Stochastic events such as smoking, infection, periodontitis, lung inflammation, or the gut microbiome thus lead to induce enzymes (for example, peptidyl arginine deiminases) that alter peptides and produce neo-epitopes not encountered by the thymus during early development. This concept is especially relevant since recent studies suggest that the gastrointestinal flora in early RA might be unique, with an overabundance of Provatella copri[7]. These environmental differences could potentially contribute also to altered polarization of T cells to the pathogenic T-helper type 17 phenotype [8].The autoreactive clones that recognize altered antigens were not deleted during development and can respond appropriately to the antigen. An array of citrullinated peptides fit avidly into the HLA-DR binding groove and activate T cells much more efficiently than the native protein [9]. These early steps probably represent a normal adaptive immune response against altered antigens rather than true autoimmunity. Production of ACPAs directed against a variety of peptides ensues. In the presence of a second hit, such as immune complexes or other mechanisms that activate innate immunity and prepare the synovium, ACPAs gain access to the joint, engage complement, and recruit inflammatory cells that amplify the response. Ultimately, breakdown of tolerance and true autoimmunity against the native proteins ensue, possibly by epitope spread. Interestingly, recently described novel antibody systems to other altered antigens associated with RA, such as through carbamylation rather than citrullination [10], could lead to a similar process.The most persuasive argument that RA has multiple pathways to the same phenotype is the diversity of responses to highly specific immunotherapies. T-cell co-stimulation blocker, B-cell depletion, tumor necrosis factor inhibitors, or interleukin-6 inhibitors demonstrate similar clinical response rates; that is, about one-half of patients treated with any single agent have a major benefit [11]. If a patient does not respond to one targeted agent, a good response to another agent with a distinct mechanism of action is only slightly less likely than in a biologic-naïve patient [12].Evaluation of genes or other analytes to stratify patients based on their underlying pathogenesis rather than on clinical phenotype could shed light on how the variable responses occur. Figure 1 shows an example (which is clearly a simplification), focusing only on gene associations. In this model, a patient with clusters of disease-associated SNPs enriched for tumor necrosis factor regulation, for example, might be expected to be a tumor necrosis factor responder. A B-cell genotype, a T-cell genotype, and so on, would also provide clues on how to treat a patient. If no particular clustering occurs and the gene associations are spread across multiple pathways, then any individual targeted therapy would have a low likelihood of success.Open in a separate windowFigure 1Simplified schema showing how genes might affect clinical responses to targeted therapies. Various genes with associated single nucleotide polymorphisms (SNPs) could be generally categorized into various pathogenic mechanisms (for example, tumor necrosis factor (TNF), T cells, B cells, others in this version). A particular individual might only inherit a subset of each of these SNPs. If the majority of inherited SNPs cluster in one mechanism, such as TNF blocker (see bottom rows), then the individual would have a response to the agent that targets this pathway. If the SNPs are not enriched for any particular pathway, then the patient would be a nonresponder. This schema only focuses on SNPs, but would be integrated with pathways that are enriched for epigenetic marks or other regulators of gene expression/function.As attractive as this notion might be, RA is not that simple and, despite individual studies with potential signals, we cannot reliably predict which patients will respond to a particular biologic despite evaluating many gene associations as well as studies of blood cytokines, synovial pathology, or serum autoantibody profiles. Success will probably require integrating more sophisticated datasets that also take into account many nongenetic influences, such as epigenomics, microbiomics, proteomics, metabolomics, or immunomics, to define the deep profile of a particular individual’s version of RA. Initial studies examining potential pathogenic pathways focusing on DNA methylation in RA synoviocytes or integrating DNA methylation and gene associations in peripheral blood cells provide insights into how this information might begin to identify previously unrecognized subsets [13-15]. Systems biology approaches to nongenetic and genetic influences also permit application of computational methods to test the effects of perturbing networks in silico. While this approach is still in its infancy, it could ultimately decrease the need for biologic validation of every potential target or could identify combinations of therapies that will be additive or synergistic.These observations suggest that RA might be thought of as a collection of distinct mechanisms rather than a single pathway; that is, as a syndrome rather than a disease. A similar conceptual evolution has occurred with other diseases, such as acute myelogenous leukemia, with a transition from phenotype or histologic diagnosis to segmenting the disease by genotype. We face the reverse of past progress in medicine, where a unifying cause ultimately links many clinical phenotypes, such as the great imitator syphilis. Instead, our understanding of RA as a clinical phenotype is devolving into multiple pathogenic pathways. RA might have a common entry point, such as adaptive immune responses to altered peptides followed by immune complexes and autoimmunity, but the subsequent byzantine pathway to the clinical phenotype is so convoluted and personalized that solving RA for a particular patient requires a systems approach using multiple emerging technologies.We have come a long way from “acute rheumatism”, but still have far to go before these pathogenic processes can be meaningfully dissected. The therapeutic successes with the average patient have been stunning, but we have reached the limit of this traditional approach. We must begin the process of deconvoluting RA using unbiased technology and carefully integrating predetermined and stochastic influences that lead to the syndrome we call RA. 相似文献