Altered cytoskeletal regulation impacts numerous physiological phenomena: cell motility, apoptosis, oncogenic transformation and parasitic infection. The protein tyrosine phosphatase (PTP)-PEST contains multiple motifs mediating its recruitment to signalling components, and is required for actin filament organization. However, little is known regarding either the importance of PTP-PEST subcellular localization, or the role of PTP-PEST in either parasitic infection or apoptosis. My doctoral research was therefore focussed on elucidating the effect of subcellular distribution on PTP-PEST activity, specifically with respect to regulation of p130Cas (a PTP-PEST substrate), as well as on the involvement of PTP-PEST in both host-pathogen relations and apoptosis. First, PTP-PEST was found both within the cytosol and at the plasma membrane. Using PTP-PEST -/- rescued cell lines, I observed that tyrosine phosphorylation-dependent p130Cas interactions were controlled primarily by cytosolic PTP-PEST. Secondly, infection of fibroblasts with Leishmania major was observed to induce dramatic actin rearrangements, and to alter the phosphorylation state of numerous proteins. Importantly, both PTP-PEST and p130Cas were processed by the parasitic protease GP63 during infection. GP63 was also required for the cleavage of additional host proteins: cortactin, TC-PTP and caspase-3. Of note, Leishmania parasites mediated p38 inactivation, correlating with the proteolysis of its upstream activator TAB1, in a GP63dependent manner. These results indicate that GP63 plays a key role in a number of biochemical events, potentially contributing to Leishmania infectivity. Finally, PTP-PEST was found to relocalize to the edges of retracting membrane ruffles of apoptotic cells. Surprisingly, PTP-PEST was specifically cleaved by caspase-3 at the 549DSPD motif during apoptosis; leading to modification of catalytic activity and scaffolding properties, and sensitizing cells to Fas-mediated detachment. As this data demonstrated a potential role for caspase cleavage in PTP regulation, I also investigated the presence of conserved putative caspase-c1eavage sites in other family members. In summary, the data presented herein links PTP-PEST with various biological processes: oncogenic signalling, host-pathogen interactions, and apoptosis. In addition to demonstrating the involvement of PTP-PEST in diverse signalling pathways, these studies underscore the importance of subcellular localization and proteolysis in the regulation of this PTP.

The short analysis of vibromotors (VM) and rolamite type mechanisms is submitted, their advantages are revealed. Dynamic processes, occurring in vibromotor, are investigated experimentally in which the slanting impact of the converter of longitudinal fluctuations (CLF) to a rotor is used, and rotates it. It is established optimum angle of elastic pressing of the converter of longitudinal fluctuations to a rotor makes 125º- are getting the greatest revolutions of a rotor. The size elastic pressing of the converter of longitudinal fluctuations to a rotor unsignificantly influences the form of the mechanical characteristics. It is recommending for precision systems working in a wide temperature range and at linear and vibrating overloads to use the circuits with included in a contour of the generator by converters or to construct auto ascillatory generators working on resonant frequency of the converter. The value of non synchronicity of rotation of rotors most grows in elementary vibromotor with two rotors, least - in rolamite vibromotor, in which the converter of longitudinal fluctuations rotates both rotors. The variant of rolamite vibromotor, in which the converter of longitudinal fluctuations rotates both rotors, is optimum. Four problems inherent vibromotors are established and ways of their decision are offered. Expediently CLF fix in its middle. Thus it is provided the least damping of vibrations. Expediently to use waveguides in the construction of VM. Thus it is... [to full text]

Calcium ions (Ca2+) are admitted into presynaptic nerve terminals through voltage gated calcium channels and diffuse to bind and activate the secretory vesicle discharge mechanism. Current research favors a highly ‘modal’ organization where the release sites are activated by one or a few closely apposed channels (Stanley, 1997). To fully understand the nanophysiology of transmitter release site activation, it is necessary to determine the rate of Ca2+ flux through individual channels at normal physiological external concentrations. OBJECTIVE: To explore the relationship between CaV2.2 channel conductance and external Ca2+ across the physiological range. CONCLUSION: The conductance of the CaV2.2 channel was determined across the range of 1-100 mM [Ca2+]EXT . With 2 mM [Ca2+]EXT, the conductance was determined to be 2.76 ± 0.24 pS.

Calcium ions (Ca2+) are admitted into presynaptic nerve terminals through voltage gated calcium channels and diffuse to bind and activate the secretory vesicle discharge mechanism. Current research favors a highly ‘modal’ organization where the release sites are activated by one or a few closely apposed channels (Stanley, 1997). To fully understand the nanophysiology of transmitter release site activation, it is necessary to determine the rate of Ca2+ flux through individual channels at normal physiological external concentrations. OBJECTIVE: To explore the relationship between CaV2.2 channel conductance and external Ca2+ across the physiological range. CONCLUSION: The conductance of the CaV2.2 channel was determined across the range of 1-100 mM [Ca2+]EXT . With 2 mM [Ca2+]EXT, the conductance was determined to be 2.76 ± 0.24 pS.

A study was undertaken to characterize two dynamic fuel types not included in the Canadian Forest Fire Behaviour Prediction System: forested bogs and blowdown. Fuel load and structure were measured at ten forested bog sites in central Alberta along a 108 year post-fire chronosequence. Canopy bulk density increased following a sigmoidal curve between 0.00 and 0.54 kg•m-3. Crown fire potential was modeled using a general crown fire behaviour model and found to follow a similar sigmoidal pattern increasing with time-since-fire. Blowdown fuel loads were measured at six sites in northwestern Ontario and ranged from 13.4 to 18.9 kg•m-2. Elevated fine blowdown fuels were found to have faster reaction times and dry more rapidly than predicted by the Fine Fuel Moisture Code. Detailed observations were also made of fire behaviour in blowdown fuels

Various real processes, occurring in the nature, technology, etc., are usually described by differential equations. Due to the development of computer software, computers have become the main tool for solving problems of different fields. They enable not only to solve complex differential equations or their systems, but also to analyze the dependence of differential equations solutions on various parameters and initial values. Up to the present many methods for the solution of differential equations have been developed, therefore, the user can solve differential equation, using several different methods. Different methods of solution enable to avoid various mistakes and to reduce errors. Differential equations can be solved not only using numerical methods, but also by applying methods of algebraic operator equations. When the latter method is being used, solutions are expressed in power series, the convergence of which has to be analyzed separately. This paper includes the analysis of Mathieu-type differential equations solutions dependence on initial conditions and parameters, as well as the establishment of solutions attractor zones and curves, which separate different attractor zones. It is very important to indicate the most exact crossing limits among different attractor zones. In order to avoid huge errors, we carried out the research by using two methods: operator and Runge-Kutta.

Cardiovascular disease is the most common cause of morbidity and mortality in type 2 diabetes. Short-term studies of glucagon-like peptide-1 (GLP-1)-targeted therapies suggest potential beneficial effects on cardiovascular outcomes. The mechanism behind this unexpectedly rapid effect is not known. In this study, full-length human GLP-1 receptor (GLP-1R) mRNA was cloned and sequenced from a human megakaryocyte cell line. Quantitative RT-PCR results showed that expression levels were comparable to other GLP-1R expressing tissues. Furthermore, incubation with GLP-1 and the GLP-1R agonist exenatide elicited a cAMP response in these cells. As megakaryocytes are the cellular precursors of platelets, the effect of GLP-1 and exenatide were studied in gel-filtered human platelet aggregation, where they were both shown to have an inhibitory effect on thrombin-stimulated platelet aggregation. Platelet inhibition by GLP-1 and GLP-1R agonists presents a potential mechanism for the reduced incidence of atherothrombotic events thought to be associated with GLP-1-targeted therapies.

Natriuretic peptides are a group of hormones, including atrial-, brain-, and C-type- natriuretic peptides (ANP, BNP, CNP). BNP can bind to two NP receptors (NPRs) denoted NPR-A (activates guanylyl cyclase) and NPR-C (activates inhibitory G- proteins). This study investigated the electrophysiological effects of BNP on isolated mouse atrial myocytes. Current-clamp experiments show that BNP had no effect on action potential (AP) parameters in basal conditions; however, when pre-stimulated with the ?-adrenergic receptor agonist isoproterenol (ISO), BNP prolonged AP duration. Voltage-clamp experiments demonstrate that BNP increased L-type calcium current (ICa,L) in the presence of ISO without altering cardiac potassium currents. The BNP effect on ICa,L was blocked by A71915 (a selective NPR-A antagonist), maintained in myocytes lacking NPR-C receptors and blocked by the phosphodiesterase-3 (PDE-3) inhibitor milrinone. These data demonstrate that BNP prolongs AP duration and increases ICa,L in atrial myocytes by activating NPR-A, increasing intracellular cGMP, and inhibiting PDE- 3.

Enteropathogenic Escherichia coli (EPEC) is an enteric pathogen that causes potentially fatal infantile diarrhea. A type III secretion system is employed by EPEC to inject bacterial effector proteins directly into host intestinal epithelial cells. The multivalent chaperone, CesT, interacts with nine effectors and is a significant contributor to EPEC pathogenesis. A putative transcriptional promoter region was identified directly upstream of cesT. In silico analyses identified conserved elements that suggest the cesT promoter is recognized by ?70. Using transcriptional fusions to lux reporter genes I showed that the cesT promoter region is active under conditions known to induce virulence-gene expression. I conclude that the cesT promoter is active early during an in vitro assay, and regulated by different mechanisms than those affecting the Ptir operon promoter.

Shigellae are pathogenic bacteria that cause the disease shigellosis. Two methods for studying secreted effectors encoded by this pathogen’s virulence plasmid are described. First, protein microarrays were used to identify substrates of an E3 ubiquitin ligase called IpaH7.8. Second, a deletion collection containing mutants for every gene on the virulence plasmid was used in two screens: one to identify mutants that elicit atypical levels of Interleukin-8 (IL-8) from U937 cells, and one to identify mutants that bind the dye Congo red abnormally. Although protein microarrays were an ineffective tool, the deletion collection proved valuable. Most mutants were less effective at sequestering Congo red than wild-type S. flexneri, although this ability was enhanced in several mutants. Four mutants, ?ospB, ?orf186, ?mxiH and ?mxiK, elicited higher levels of IL-8 from U937 cells than wild type S. flexneri. These results validate the use of the deletion collection as a tool for studying bacterial pathogenesis.