Characterization of the chloroquine resistance transporter homologue in Toxoplasma gondii

Mutations in the Plasmodium falciparum chloroquine resistance transporter (PfCRT) protein confer resistance to the antima-larial drug chloroquine. PfCRT localizes to the parasite digestive vacuole, the site of chloroquine action, where it mediates resistance by transporting chloroquine out of the digestive vacuole. PfCRT belongs to a family of transporter proteins called the chlo-roquine resistance transporter family. CRT family proteins are found throughout the Apicomplexa, in some protists, and in plants. Despite the importance of PfCRT in drug resistance, little is known about the evolution or native function of CRT proteins. The apicomplexan parasite Toxoplasma gondii contains one CRT family protein. We demonstrate that T. gondii CRT (TgCRT) colocalizes with markers for the vacuolar (VAC) compartment in these parasites. The TgCRT-containing VAC is a highly dynamic organelle, changing its morphology and protein composition between intracellular and extracellular forms of the parasite. Regulated knockdown of TgCRT expression resulted in modest reduction in parasite fitness and swelling of the VAC, indicating that TgCRT contributes to parasite growth and VAC physiology. Together, our findings provide new information on the role of CRT family proteins in apicomplexan parasites

Spatial Localisation of Actin Filaments across Developmental Stages of the Malaria Parasite

Actin dynamics have been implicated in a variety of developmental processes during the malaria parasite lifecycle. Parasite motility, in particular, is thought to critically depend on an actomyosin motor located in the outer pellicle of the parasite cell. Efforts to understand the diverse roles actin plays have, however, been hampered by an inability to detect microfilaments under native conditions. To visualise the spatial dynamics of actin we generated a parasite-specific actin antibody that shows preferential recognition of filamentous actin and applied this tool to different lifecycle stages (merozoites, sporozoites and ookinetes) of the human and mouse malaria parasite species Plasmodium falciparum and P. berghei along with tachyzoites from the related apicomplexan parasite Toxoplasma gondii. Actin filament distribution was found associated with three core compartments: the nuclear periphery, pellicular membranes of motile or invasive parasite forms and in a ring-like distribution at the tight junction during merozoite invasion of erythrocytes in both human and mouse malaria parasites. Localisation at the nuclear periphery is consistent with an emerging role of actin in facilitating parasite gene regulation. During invasion, we show that the actin ring at the parasite-host cell tight junction is dependent on dynamic filament turnover. Super-resolution imaging places this ring posterior to, and not concentric with, the junction marker rhoptry neck protein 4. This implies motor force relies on the engagement of dynamic microfilaments at zones of traction, though not necessarily directly through receptor-ligand interactions at sites of adhesion during invasion. Combined, these observations extend current understanding of the diverse roles actin plays in malaria parasite development and apicomplexan cell motility, in particular refining understanding on the linkage of the internal parasite gliding motor with the extra-cellular milieu

Predicted Functions of MdmX in Fine-Tuning the Response of p53 to DNA Damage

Tumor suppressor protein p53 is regulated by two structurally homologous proteins, Mdm2 and MdmX. In contrast to Mdm2, MdmX lacks ubiquitin ligase activity. Although the essential interactions of MdmX are known, it is not clear how they function to regulate p53. The regulation of tumor suppressor p53 by Mdm2 and MdmX in response to DNA damage was investigated by mathematical modeling of a simplified network. The simplified network model was derived from a detailed molecular interaction map (MIM) that exhibited four coherent DNA damage response pathways. The results suggest that MdmX may amplify or stabilize DNA damage-induced p53 responses via non-enzymatic interactions. Transient effects of MdmX are mediated by reservoirs of p53∶MdmX and Mdm2∶MdmX heterodimers, with MdmX buffering the concentrations of p53 and/or Mdm2. A survey of kinetic parameter space disclosed regions of switch-like behavior stemming from such reservoir-based transients. During an early response to DNA damage, MdmX positively or negatively regulated p53 activity, depending on the level of Mdm2; this led to amplification of p53 activity and switch-like response. During a late response to DNA damage, MdmX could dampen oscillations of p53 activity. A possible role of MdmX may be to dampen such oscillations that otherwise could produce erratic cell behavior. Our study suggests how MdmX may participate in the response of p53 to DNA damage either by increasing dependency of p53 on Mdm2 or by dampening oscillations of p53 activity and presents a model for experimental investigation

Aryl amino acetamides prevent Plasmodium falciparum ring development via targeting the lipid-transfer protein PfSTART1.

With resistance to most antimalarials increasing, it is imperative that new drugs are developed. We previously identified an aryl acetamide compound, MMV006833 (M-833), that inhibited the ring-stage development of newly invaded merozoites. Here, we select parasites resistant to M-833 and identify mutations in the START lipid transfer protein (PF3D7_0104200, PfSTART1). Introducing PfSTART1 mutations into wildtype parasites reproduces resistance to M-833 as well as to more potent analogues. PfSTART1 binding to the analogues is validated using organic solvent-based Proteome Integral Solubility Alteration (Solvent PISA) assays. Imaging of invading merozoites shows the inhibitors prevent the development of ring-stage parasites potentially by inhibiting the expansion of the encasing parasitophorous vacuole membrane. The PfSTART1-targeting compounds also block transmission to mosquitoes and with multiple stages of the parasite's lifecycle being affected, PfSTART1 represents a drug target with a new mechanism of action

Proceedings of the Thirteenth International Society of Sports Nutrition (ISSN) Conference and Expo

Meeting Abstracts: Proceedings of the Thirteenth International Society of Sports Nutrition (ISSN) Conference and Expo Clearwater Beach, FL, USA. 9-11 June 201

General dynamical sharp-interface conditions for phase transformations in viscous heat-conducting fluids

We develop a complete set of equations governing the evolution of a sharp interface separating two fluid phases undergoing transformation. In addition to the conventional balances for mass, linear momentum, and energy these equations include also a counterpart of the Gibbs???Thomson equation familiar from theories for crystal growth. This additional equation arises from a consideration of configurational forces within a thermodynamical framework. While the notion of configurational forces is well-developed and understood for the description of materials, like crystalline solids, that possess natural reference con.gurations, very little has been done regarding their role in materials, such as viscous .uids, that do not possess preferred reference states. We therefore provide a comprehensive discussion of configurational forces, the balance of configurational momentum, and configurational thermodynamics that does not require a choice of reference configuration. The general evolution equations arising from our theory account for the thermodynamic structure of the bulk phases and the interface and for viscous and thermal dissipation in the bulk phases and for viscous dissipation on the interface. Due to the complexity of these equations, we provide a reduced system of equations based on simplified constitutive assumptions and approximations common in the literature on phase transformations.published or submitted for publicationis peer reviewe

Final technical report: Third SIAM Conference on Mathematical Aspects of Materials Science [Program and abstracts]

The meeting gathered the interdisciplinary community of scientists working on mathematical and computational aspects of materials science. One goal was to examine recent methodological advances in areas such as analysis of moving interfaces, macroscopic consequences of microstructure, and defects in materials. A second goal was to highlight recent accomplishments in areas such as materials design, synthesis, and processing; simulation of phase transformation dynamics; growth and morphology of thin films; and electromagnetic materials. A third goal was to identify promising directions for new developments of a mathematical or computational nature in areas such as multiscale analysis of materials--from atomic to continuum; nanoscale structures; liquid crystals, glasses, and polymers; and soft materials and bio-materials. There were 270 registered participants. The meeting included both plenary lectures and minisymposia

A1-L10 phase boundaries and anisotropy via multiple-order-parameter theory for an fcc alloy

The dependence of thermodynamic properties of planar interphase boundaries (IPBs) and antiphase boundaries (APBs) in a binary alloy on an fcc lattice is studied as a function of their orientation. Using a recently developed diffuse interface model based on three non-conserved order parameters and the concentration, and a free energy density that gives a realistic phase diagram with one disordered phase (A1) and two ordered phases (L12 and L10) such as occur in the Cu-Au system, we are able to find IPBs and APBs between any pair of phases and domains, and for all orientations. The model includes bulk and gradient terms in a free energy functional, and assumes that there is no mismatch in the lattice parameters for the disordered and ordered phases.We catalog the appropriate boundary conditions for all IPBs and APBs. We then focus on the IPB between the disordered A1 phase and the L10 ordered phase. For this IPB we compute the numerical solution of the boundary value problem to find its interfacial energy, γ as a function of orientation, temperature, and chemical potential (or composition). We determine the equilibrium shape for a precipitate of one phase within the other using the Cahn-Hoffman "-vector" formalism. We find that the profile of the interface is determined only by one conserved and one non-conserved order parameter, which leads to a surface energy which, as a function of orientation, is "transversely isotropic" with respect to the tetragonal axis of the L10 phase. We verify the model's consistency with the Gibbs adsorption equation.NASA; National Science Foundation (DMS-9722854-DMS-9631287