Two essential light chains regulate the MyoA lever arm to promote Toxoplasma gliding motility

Key to the virulence of apicomplexan parasites is their ability to move through tissue and to invade and egress from host cells. Apicomplexan motility requires the activity of the glideosome, a multicomponent molecular motor composed of a type XIV myosin, MyoA. Here we identify a novel glideosome component, essential light chain 2 (ELC2), and functionally characterize the two essential light chains (ELC1 and ELC2) of MyoA in Toxoplasma. We show that these proteins are functionally redundant but are important for invasion, egress, and motility. Molecular simulations of the MyoA lever arm identify a role for Ca2+ in promoting intermolecular contacts between the ELCs and the adjacent MLC1 light chain to stabilize this domain. Using point mutations predicted to ablate either the interaction with Ca2+ or the interface between the two light chains, we demonstrate their contribution to the quality, displacement, and speed of gliding Toxoplasma parasites. Our work therefore delineates the importance of the MyoA lever arm and highlights a mechanism by which this domain could be stabilized in order to promote invasion, egress, and gliding motility in apicomplexan parasites

Brain Communications

Brain charts for the human lifespan have been recently proposed to build dynamic models of brain anatomy in normal aging and various neurological conditions. They offer new possibilities to quantify neuroanatomical changes from preclinical stages to death, where longitudinal MRI data are not available. In this study, we used brain charts to model the progression of brain atrophy in progressive supranuclear palsy – Richardson syndrome (PSPRS). We combined multiple datasets (n=8170 quality controlled MRI of healthy subjects from 22 cohorts covering the entire lifespan, and n=62 MRI of PSP-RS patients from the 4 Repeat Tauopathy Neuroimaging Initiative) to extrapolate lifetime volumetric models of healthy and PSP-RS brain structures. We then mapped in time and space the sequential divergence between healthy and PSP-RS charts. We found six major consecutive stages of atrophy progression: (i) ventral diencephalon (including subthalamic nuclei, substantia nigra, and red nuclei), (ii) pallidum, (iii) brainstem, striatum and amygdala, (iv) thalamus, (v) frontal lobe and (vi) occipital lobe. The three structures with most severe atrophy over time were the thalamus, followed by the pallidum and the brainstem. These results match the neuropathological staging of tauopathy progression in PSP-RS, where the pathology is supposed to start in the pallido-nigro-luysian system and spreads rostrally via the striatum and the amygdala to the cerebral cortex, and caudally to the brainstem. This study supports the use of brain charts for the human lifespan to study the progression of neurodegenerative diseases, especially in the absence of specific biomarkers as in PSP.Apprentissage profond pour la volumétrie cérébrale : vers le BigData en neuroscienceInitiative d'excellence de l'Université de Bordeau

Apicoplast isoprenoid precursor synthesis and the molecular basis of fosmidomycin resistance in Toxoplasma gondii

Expression of a bacterial transporter protein in Toxoplasma gondii results in parasite susceptibility to Formidomycin, a drug targeting isoprenoid precursor synthesis

A Systematic Screen to Discover and Analyze Apicoplast Proteins Identifies a Conserved and Essential Protein Import Factor

Parasites of the phylum Apicomplexa cause diseases that impact global health and economy. These unicellular eukaryotes possess a relict plastid, the apicoplast, which is an essential organelle and a validated drug target. However, much of its biology remains poorly understood, in particular its elaborate compartmentalization: four membranes defining four different spaces. Only a small number of organellar proteins have been identified in particular few proteins are known for non-luminal apicoplast compartments. We hypothesized that enlarging the catalogue of apicoplast proteins will contribute toward identifying new organellar functions and expand the realm of targets beyond a limited set of characterized pathways. We developed a bioinformatic screen based on mRNA abundance over the cell cycle and on phyletic distribution. We experimentally assessed 57 genes, and of 30 successful epitope tagged candidates eleven novel apicoplast proteins were identified. Of those, seven appear to target to the lumen of the organelle, and four localize to peripheral compartments. To address their function we then developed a robust system for the construction of conditional mutants via a promoter replacement strategy. We confirm the feasibility of this system by establishing conditional mutants for two selected genes – a luminal and a peripheral apicoplast protein. The latter is particularly intriguing as it encodes a hypothetical protein that is conserved in and unique to Apicomplexan parasites and other related organisms that maintain a red algal endosymbiont. Our studies suggest that this peripheral plastid protein, PPP1, is likely localized to the periplastid compartment. Conditional disruption of PPP1 demonstrated that it is essential for parasite survival. Phenotypic analysis of this mutant is consistent with a role of the PPP1 protein in apicoplast biogenesis, specifically in import of nuclear-encoded proteins into the organelle

Determination of pion-baryon coupling constants from QCD sum rules

We evaluate the pi-N-N, pi-Sigma-Sigma and pi-Sigma-Lambda coupling constants using QCD sum rules based on pion-to-vacuum matrix elements of correlators of two interpolating baryon fields. The parts of the correlators with Dirac structure k-slash gamma_5 are used, keeping all terms up to dimension 5 in the OPE and including continuum contributions on the phenomenological side. The ratios of these sum rules to baryon mass sum rules yield stable results with values for the couplings of g_{pi NN}=12+/-5, g_{pi Sigma Sigma}=7+/-4 and g_{pi Sigma Lambda}=6+/-3. The sources of uncertainty are discussed.Comment: 20 pages (RevTeX), 1 figure (attached). Revised version with corrected form for Y2 piece of hyperon correlators; numerical results differ from previous versio

13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides

We present here a set of 13C-direct detected NMR experiments to facilitate the resonance assignment of RNA oligonucleotides. Three experiments have been developed: (1) the (H)CC-TOCSY-experiment utilizing a virtual decoupling scheme to assign the intraresidual ribose 13C-spins, (2) the (H)CPC-experiment that correlates each phosphorus with the C4′ nuclei of adjacent nucleotides via J(C,P) couplings and (3) the (H)CPC-CCH-TOCSY-experiment that correlates the phosphorus nuclei with the respective C1′,H1′ ribose signals. The experiments were applied to two RNA hairpin structures. The current set of 13C-direct detected experiments allows direct and unambiguous assignment of the majority of the hetero nuclei and the identification of the individual ribose moieties following their sequential assignment. Thus, 13C-direct detected NMR methods constitute useful complements to the conventional 1H-detected approach for the resonance assignment of oligonucleotides that is often hindered by the limited chemical shift dispersion. The developed methods can also be applied to large deuterated RNAs

Identification of Rhoptry Trafficking Determinants and Evidence for a Novel Sorting Mechanism in the Malaria Parasite Plasmodium falciparum

The rhoptry of the malaria parasite Plasmodium falciparum is an unusual secretory organelle that is thought to be related to secretory lysosomes in higher eukaryotes. Rhoptries contain an extensive collection of proteins that participate in host cell invasion and in the formation of the parasitophorous vacuole, but little is known about sorting signals required for rhoptry protein targeting. Using green fluorescent protein chimeras and in vitro pull-down assays, we performed an analysis of the signals required for trafficking of the rhoptry protein RAP1. We provide evidence that RAP1 is escorted to the rhoptry via an interaction with the glycosylphosphatidyl inositol-anchored rhoptry protein RAMA. Once within the rhoptry, RAP1 contains distinct signals for localisation within a sub-compartment of the organelle and subsequent transfer to the parasitophorous vacuole after invasion. This is the first detailed description of rhoptry trafficking signals in Plasmodium

Autophagy Protein Atg3 is Essential for Maintaining Mitochondrial Integrity and for Normal Intracellular Development of Toxoplasma gondii Tachyzoites

Autophagy is a cellular process that is highly conserved among eukaryotes and permits the degradation of cellular material. Autophagy is involved in multiple survival-promoting processes. It not only facilitates the maintenance of cell homeostasis by degrading long-lived proteins and damaged organelles, but it also plays a role in cell differentiation and cell development. Equally important is its function for survival in stress-related conditions such as recycling of proteins and organelles during nutrient starvation. Protozoan parasites have complex life cycles and face dramatically changing environmental conditions; whether autophagy represents a critical coping mechanism throughout these changes remains poorly documented. To investigate this in Toxoplasma gondii, we have used TgAtg8 as an autophagosome marker and showed that autophagy and the associated cellular machinery are present and functional in the parasite. In extracellular T. gondii tachyzoites, autophagosomes were induced in response to amino acid starvation, but they could also be observed in culture during the normal intracellular development of the parasites. Moreover, we generated a conditional T. gondii mutant lacking the orthologue of Atg3, a key autophagy protein. TgAtg3-depleted parasites were unable to regulate the conjugation of TgAtg8 to the autophagosomal membrane. The mutant parasites also exhibited a pronounced fragmentation of their mitochondrion and a drastic growth phenotype. Overall, our results show that TgAtg3-dependent autophagy might be regulating mitochondrial homeostasis during cell division and is essential for the normal development of T. gondii tachyzoites