Identification and partial characterization of cAMP-phosphodiesterases in the ciliate Euplotes raikovi.

In the ciliate Euplotes raikovi, two specific isoforms of cAMP- dependent phosphodiesterases were identified, one in the soluble and the other in the particulate fraction of the cell. Their activity was shown to be stimulated by Mg2+, insensitive to Ca2+ and cGMP, and scarcely inhibited by theophylline and 3-isobutyl-1-methyl-xanthine. They appear to be related to some phosphodiesterases of class II of other unicellular organisms in their biochemical features, and their enzymatic activity is up-regulated by elevation of intracellular cAMP level similarly to PDE-4 isoforms of mammals

Cross-talk between the autocrine (mitogenic) pheromone loop of the ciliate Euplotes raikovi and the intracellular cyclic AMP concentration

Cell type-specific protein signals, called pheromones, are constitutively secreted by Euplotes raikovi and bound back in autocrine fashion, with a positive effect on the vegetative (mitotic) cell growth. In cells growing suspended with their secreted pheromone, it was found that any interruption of this autocrine signaling loop was immediately followed by an effective enhancement of the basal intracellular cyclic AMP (cAMP) level. To establish a cause-effect relationship between these pheromone-induced variations in the cytoplasmic cAMP level and cell growth, cells ready to pass from a resting stage to a new growth cycle were conditioned either to incorporate a cAMP analog resistant to phosphodiesterase degradation, or to utilize cAMP released (following cell irradiation) from incorporated “caged” cAMP. Cells responded at every induced increase in their basal cAMP level by markedly decreasing their commitment to start a new growth cycle. It was deduced that the autocrine signaling of E. raikovi pheromones involves cAMP as inhibitor of its mitogenic activity

Exploiting Residue Curve Maps to Assess Thermodynamic Feasibility Boundaries under Uncertain Operating Conditions

The very first step of almost any separation process design procedure is the thermodynamic feasibility analysis. In the case of distillation, residue curve maps (RCMs) represent an essential tool to assess whether the separation is feasible or not. However, the analysis is generally carried out by referring to nominal operating conditions and product purities as specification. This means that, when process parameters are likely to undergo fluctuations, the prediction of the system response is not that obvious. An ABE/W (acetone−butanol−ethanol/water) mixture was then selected as a case study since it allows us to discuss several non-ideal thermodynamic behaviors and because of the renewed interest in biorefinery and sustainable processes during recent years. Residue curve mapping was then exploited to determine the thermodynamic feasibility range for multicomponent distillation processes as well as for distillation trains and process-intensified solutions taking into account both product purity and product recovery specifications. The final product of this study is a thorough procedure to determine the flexibility boundaries of feed and product compositions as well as an immediate and intuitive graphical representation from a binary standard distillation column to a complex multicomponent dividing wall column applicatio

Self/non-self recognition in the ciliate Euplotes raikovi: characterization of Er-MAPK1, a downstream component of the autocrine signal transduction pathway. REPORT of the IXth scientific meeting of the Italian Association of Developmental and Comparative Immunobiology (IADCI), Varese

In the ciliate Euplotes raikovi, cell type-specific, water-borne signal proteins (pheromones) control self/non-self recognition phenomena by binding their target cell-surface receptors and activating downstream signal transduction pathways. Immunorecognition analyses of E. raikovi cell extracts revealed that at least three distinct protein kinases are activated (phosphorylated) in functional association with the autocrine pheromone-receptor loop that promotes the vegetative (mitogenic) cell growth. One of these kinases, designated as Er-MAPK1 (from Mitogen-Activated Protein Kinases), was structurally characterized by molecular cloning of the relevant gene. The Er-MAPK1 N-terminal half of 300 amino acids bears unmistakable structural homology with the “intestinal cell kinase” and the “male-germ cell associated kinase”, that are involved in the regulation of proliferation and differentiation of specialized animal cells. It contains all the basic structural features that are required for a MAPK catalytic activity, in particular the dual phosphorylation site represented by the Thr-Asp-Tyr motif in the activation loop. In contrast, the Er-MAPK1 C-terminal half of 331 amino acids appears to be structurally unique. It is particularly rich in glycine residues and potential sites of regulatory activities, and shows sequence motifs that clearly predicts a nuclear localization of Er-MAPK1

A Novel Protein Kinase from the Ciliate Euplotes raikovi with Close Structural Identity to the Mammalian Intestinal and Male-Germ Cell Kinases: Characterization and Functional Implications in the Autocrine Pheromone Signaling Loop

In the free-living ciliate Euplotes raikovi, we identified (and designated as Er-MAPK1) a protein kinase of 631 amino acids, that appears to be constantly phosphorylated in cells which are in growth stage and interact in autocrine fashion with their water-soluble signal pheromones. Er-MAPK1 is specified by a gene that requires aþ1 translational frame-shift to be expressed. Its amino-terminal region represents a canonical catalytic domain and carries an activation loop distinctive of the mitogenactivated protein kinases, with the Thr-Asp-Tyr motif deputed to be site of double phosphorylation. In contrast, the carboxy-terminal region appears to be structurally unique. It shows a strongly basic amino acid composition, is very rich in glycine repetitions, and contains a bipartite signal for translocation of Er-MAPK1 into the nucleus