MAP Kinase Modules: The Excursion Model and the Steps that Count

MAP kinase modules propagate diverse extracellular signals to downstream effectors. The two dual phosphorylation reactions catalyzed by the modules are thought to control the switch behavior of the pathway. Here we review recent approaches to understand these pathways through signal-to-response studies in cells and in vitro. These data are reconciled with physical models as well as predictions made on mathematical and theoretical grounds. Biochemical analysis has shown recently that the dual phosphorylation reactions catalyzed by MAP kinase modules are sequential at both levels of the cascade. The observed order of phosphorylation events suggests an excursion from the Ser/Thr kinase activity of the MAP3K into Tyr kinase activity of the central dual specificity MAP2K. How the order of events might be encoded in the structures and interactions is discussed. The ordered mechanism confirms predictions that reactions should be sequential to generate the steep signal-to-response curves and delayed responses observed in cells.     

MAP Kinase Modules: The Excursion Model and the Steps that Count

MAP kinase modules propagate diverse extracellular signals to downstream effectors. The two dual phosphorylation reactions catalyzed by the modules are thought to control the switch behavior of the pathway. Here we review recent approaches to understand these pathways through signal-to-response studies in cells and in vitro. These data are reconciled with physical models as well as predictions made on mathematical and theoretical grounds. Biochemical analysis has shown recently that the dual phosphorylation reactions catalyzed by MAP kinase modules are sequential at both levels of the cascade. The observed order of phosphorylation events suggests an excursion from the Ser/Thr kinase activity of the MAP3K into Tyr kinase activity of the central dual specificity MAP2K. How the order of events might be encoded in the structures and interactions is discussed. The ordered mechanism confirms predictions that reactions should be sequential to generate the steep signal-to-response curves and delayed responses observed in cells.     

Variation in the synthesis of membrane proteins of human T lymphocytes during mitogenic activation: recognition of a 70,000 dalton protein with anti-p23,30 heteroantiserum and a novel 42,000 dalton protein with anti-p44,12 heteroantiserum

Membrane proteins of [35S]methionine-labeled, human T lymphocytes were analyzed by SDS polyacrylamide gradient slab gel electrophoresis and autoradiography each day during a 6-day period of activation with phytohemagglutinin or with concanavalin A. This process was characterized by the novel appearance and limited duration of synthesis of many proteins, in particular of 30, 35, 48, 50, and 55 kilodalton molecules in the early days of blast transformation and subsequently of 120, 125, 135, and 145 kilodalton proteins. The HLAA-A,-B antigens and beta 2-microglobulin, as recognized by anti-p44,12 serum, were synthesized by both resting and mitogen-activated T cells on each day of culture. But, an additional 42-kilodalton protein was recognized with this same antiserum on days 4 and 5 of activation. A 70-kilodalton protein, immunoprecipitated by anti-p23,30 (anti-HLA-protein, immunoprecipitated by anti-p23,30 (anti-HLA-DR) heteroantiserum, was synthesized principally on days 2 and 4 of mitogenic transformation. This molecule was absent from normal, resting T cells, the T cell line, CCRF-CEM, and the B cell line, Raji. In a parallel test, the same anti-p23,30 serum detected the conventional HLA-DR bimolecular glycoprotein complex of 29 and 34 kilodaltons in nonionic detergent solubilized Raji B cell membrane preparations. This study described in detail the molecular changes in the membrane proteins of activated T lymphocytes and included the definition of novel forms of HLA-A,B and HLA-DR associated molecules.     

Genetic Differentiation in Response to Selection for Water-Soluble Carbohydrate Content in Perennial Ryegrass (<Emphasis Type="Italic">Lolium perenne</Emphasis> L.)

Plant carbohydrates are of increasing interest as renewable feedstocks to replace petrochemicals in the generation of fuels and production of high-value chemicals. Greater understanding of the genetic control of diversity in fructan synthesis and accumulation would facilitate more directed channelling of feedstock to process in a ryegrass biorefinery. Divergent populations produced by phenotypic selection for water-soluble carbohydrate content have been used to investigate relationships between traits, and to identify patterns of genetic differentiation which indicate genomic regions under high and low selection pressure. Selection for high water-soluble carbohydrate content was associated with increased synthesis of large fructan polymers and increased accumulation of above-ground plant biomass, particularly during spring. Three rounds of selection and two rounds of recombination resulted in widespread genetic differentiation across the whole genome, causing reduced allelic richness and increasing homozygosity at some loci. A smaller number of loci were shown to be subject to high selection pressure. Breeding material subjected to many years of selection for water-soluble carbohydrate also showed allelic differences which may reflect the consequences of high selection pressure at some of these same loci. However, some of the loci unaffected in the divergent selection experiment showed similar effects. This might arise from differences in linkage disequilibrium in these two sets of plant materials, but more likely from the different genetic background of the germplasm. This illustrates the complex nature of the water-soluble carbohydrate trait in perennial ryegrass.     

Generation and Characterization of Antibodies against Asian Elephant (Elephas maximus) IgG,IgM, and IgA

Asian elephant (Elephas maximus) immunity is poorly characterized and understood. This gap in knowledge is particularly concerning as Asian elephants are an endangered species threatened by a newly discovered herpesvirus known as elephant endotheliotropic herpesvirus (EEHV), which is the leading cause of death for captive Asian elephants born after 1980 in North America. While reliable diagnostic assays have been developed to detect EEHV DNA, serological assays to evaluate elephant anti-EEHV antibody responses are lacking and will be needed for surveillance and epidemiological studies and also for evaluating potential treatments or vaccines against lethal EEHV infection. Previous studies have shown that Asian elephants produce IgG in serum, but they failed to detect IgM and IgA, further hampering development of informative serological assays for this species. To begin to address this issue, we determined the constant region genomic sequence of Asian elephant IgM and obtained some limited protein sequence information for putative serum IgA. The information was used to generate or identify specific commercial antisera reactive against IgM and IgA isotypes. In addition, we generated a monoclonal antibody against Asian elephant IgG. These three reagents were used to demonstrate that all three immunoglobulin isotypes are found in Asian elephant serum and milk and to detect antibody responses following tetanus toxoid booster vaccination or antibodies against a putative EEHV structural protein. The results indicate that these new reagents will be useful for developing sensitive and specific assays to detect and characterize elephant antibody responses for any pathogen or vaccine, including EEHV.     

Nurse-Led Medicines' Monitoring for Patients with Dementia in Care Homes: A Pragmatic Cohort Stepped Wedge Cluster Randomised Trial

Background

People with dementia are susceptible to adverse drug reactions (ADRs). However, they are not always closely monitored for potential problems relating to their medicines: structured nurse-led ADR Profiles have the potential to address this care gap. We aimed to assess the number and nature of clinical problems identified and addressed and changes in prescribing following introduction of nurse-led medicines’ monitoring.

Design

Pragmatic cohort stepped-wedge cluster Randomised Controlled Trial (RCT) of structured nurse-led medicines’ monitoring versus usual care.

Setting

Five UK private sector care homes

Participants

41 service users, taking at least one antipsychotic, antidepressant or anti-epileptic medicine.

Intervention

Nurses completed the West Wales ADR (WWADR) Profile for Mental Health Medicines with each participant according to trial step.

Outcomes

Problems addressed and changes in medicines prescribed.

Data Collection and Analysis

Information was collected from participants’ notes before randomisation and after each of five monthly trial steps. The impact of the Profile on problems found, actions taken and reduction in mental health medicines was explored in multivariate analyses, accounting for data collection step and site.

Results

Five of 10 sites and 43 of 49 service users approached participated. Profile administration increased the number of problems addressed from a mean of 6.02 [SD 2.92] to 9.86 [4.48], effect size 3.84, 95% CI 2.57–4.11, P <0.001. For example, pain was more likely to be treated (adjusted Odds Ratio [aOR] 3.84, 1.78–8.30), and more patients attended dentists and opticians (aOR 52.76 [11.80–235.90] and 5.12 [1.45–18.03] respectively). Profile use was associated with reduction in mental health medicines (aOR 4.45, 1.15–17.22).

Conclusion

The WWADR Profile for Mental Health Medicines can improve the quality and safety of care, and warrants further investigation as a strategy to mitigate the known adverse effects of prescribed medicines.

Trial Registration

ISRCTN 48133332     

Genome sequencing reveals fine scale diversification and reticulation history during speciation in Sus

Background

Elucidating the process of speciation requires an in-depth understanding of the evolutionary history of the species in question. Studies that rely upon a limited number of genetic loci do not always reveal actual evolutionary history, and often confuse inferences related to phylogeny and speciation. Whole-genome data, however, can overcome this issue by providing a nearly unbiased window into the patterns and processes of speciation. In order to reveal the complexity of the speciation process, we sequenced and analyzed the genomes of 10 wild pigs, representing morphologically or geographically well-defined species and subspecies of the genus Sus from insular and mainland Southeast Asia, and one African common warthog.

Results

Our data highlight the importance of past cyclical climatic fluctuations in facilitating the dispersal and isolation of populations, thus leading to the diversification of suids in one of the most species-rich regions of the world. Moreover, admixture analyses revealed extensive, intra- and inter-specific gene-flow that explains previous conflicting results obtained from a limited number of loci. We show that these multiple episodes of gene-flow resulted from both natural and human-mediated dispersal.

Conclusions

Our results demonstrate the importance of past climatic fluctuations and human mediated translocations in driving and complicating the process of speciation in island Southeast Asia. This case study demonstrates that genomics is a powerful tool to decipher the evolutionary history of a genus, and reveals the complexity of the process of speciation.