Institute Of Recherches En Technologies Et Science Pour Le Vivant

Grenoble, France

Institute Of Recherches En Technologies Et Science Pour Le Vivant

Grenoble, France
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Jezegou A.,University of Paris Descartes | Jezegou A.,University Paris - Sud | Llinares E.,Free University of Colombia | Anne C.,University of Paris Descartes | And 15 more authors.
Proceedings of the National Academy of Sciences of the United States of America | Year: 2012

Cystinosin, the lysosomal cystine exporter defective in cystinosis, is the foundingmember of a family of heptahelicalmembrane proteins related to bacteriorhodopsin and characterized by a duplicated motif termed the PQ loop. PQ-loop proteins are more frequent in eukaryotes than in prokaryotes; except for cystinosin, their molecular function remains elusive. In this study,we report that three yeast PQ-loop proteins of unknown function,Ypq1,Ypq2, andYpq3, localize to the vacuolar membrane and are involved in homeostasis of cationic amino acids (CAAs).We also showthat PQLC2, amammalian PQ-loop protein closely related to yeast Ypq proteins, localizes to lysosomes and catalyzes a robust, electrogenic transport that is selective for CAAs and strongly activated at low extracytosolic pH. Heterologous expression of PQLC2 at the yeast vacuole rescues the resistance phenotype of an ypq2 mutant to canavanine, a toxic analog of arginine efficiently transported by PQLC2. Finally, PQLC2 transports a lysine-like mixed disulfide that serves as a chemical intermediate in cysteamine therapy of cystinosis, and PQLC2 gene silencing trapped this intermediate in cystinotic cells. We conclude that PQLC2 and Ypq1-3 proteins are lysosomal/vacuolar exporters of CAAs and suggest that small-molecule transport is a conserved feature of the PQ-loop protein family, in agreementwith its distant similarity to SWEET sugar transporters and to the mitochondrial pyruvate carrier. The elucidation of PQLC2 functionmay help improve cysteamine therapy. It may also clarify the origin of CAA abnormalities in Batten disease.


Jouhet J.,French National Center for Scientific Research | Jouhet J.,University Grenoble Alpes | Jouhet J.,Institute Of Recherches En Technologies Et Science Pour Le Vivant | Jouhet J.,French National Institute for Agricultural Research
Frontiers in Plant Science | Year: 2013

Domains are present in every natural membrane. They are characterized by a distinctive protein and/or lipid composition. Their size is highly variable from the nano- to the micrometer scale. The domains confer specific properties to the membrane leading to original structure and function. The determinants leading to domain organization are therefore important but remain obscure. This review presents how the ability of lipids to organize into hexagonal II or lamellar phases can promote particular local structures within membranes. Since biological membranes are composed of a mixture of lipids, each with distinctive biophysical properties, lateral and transversal sorting of lipids can promote creation of domains inside the membrane through local modulation of the lipid phase. Lipid biophysical properties have been characterized for long based on in vitro analyses using non-natural lipid molecules; their re-examinations using natural lipids might open interesting perspectives on membrane architecture occurring in vivo in various cellular and physiological contexts.


Delaroche D.,University Pierre and Marie Curie | Delaroche D.,CNRS Natural Product Chemistry Institute | Cantrelle F.-X.,CNRS Natural Product Chemistry Institute | Subra F.,Ecole Normale Superieure de Cachan | And 8 more authors.
Journal of Biological Chemistry | Year: 2010

Cell-penetrating peptides can cross cell membranes and are commonly seen as biologically inert molecules. However, we found that some cell-penetrating peptides could remodel actin cytoskeleton in oncogene-transformed NIH3T3/EWS-Fli cells. These cells have profound actin disorganization related to their tumoral transformation. These arginine- and/or tryptophanrich peptides could cross cell membrane and induce stress fiber formation in these malignant cells, whereas they had no perceptible effect in non-tumoral fibroblasts. In addition, motility (migration speed, random motility coefficient, wound healing) of the tumor cells could be decreased by the cell-permeant peptides. Although the peptides differently influenced actin polymerization in vitro, they could directly bind monomeric actin as determined by NMR and calorimetry studies. Therefore, cell-penetrating peptides might interact with intracellular protein partners, such as actin. In addition, the fact that they could reverse the tumoral phenotype is of interest for therapeutic purposes. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.


Galland R.,Institute Of Recherches En Technologies Et Science Pour Le Vivant | Leduc P.,Laboratoire Delectronique Des Technologies Of Linformation | Guerin C.,Institute Of Recherches En Technologies Et Science Pour Le Vivant | Peyrade D.,CNRS Microelectronics Technology Laboratory | And 2 more authors.
Nature Materials | Year: 2013

A promising approach to improve the performance of microelectronic devices is to build three-dimensional (3D) chips made of stacked circuits. However, a major hurdle lies in the fabrication of dense arrays of electrical interconnections between these layers, where accessibility is limited. Here we show that the directed growth and self-organization of actin filaments can offer a solution to this problem. We defined the shape and orientation of 3D actin networks through both micropatterning of actin nucleation factors and biochemical control of actin filament polymerization. Networks growing from two opposing layers were able to interpenetrate and form mechanically stable connections, which were then coated with gold using a selective metallization process. The electrical conductivity, robustness and modularity of the metallized self-organized connections make this approach potentially attractive for 3D chip manufacturing. © 2013 Macmillan Publishers Limited. All rights reserved.


Chicanne G.,University Paul Sabatier | Severin S.,University Paul Sabatier | Boscheron C.,Joseph Fourier University | Boscheron C.,Institute Of Recherches En Technologies Et Science Pour Le Vivant | And 6 more authors.
Biochemical Journal | Year: 2012

PtdIns3P is recognized as an important player in the control of the endocytotic pathway and in autophagy. Recent data also suggest that PtdIns3P contributes to molecular mechanisms taking place at the plasma membrane and at the midbody during cytokinesis. This lipid is present in low amounts in mammalian cells and remains difficult to quantify either by traditional techniques based on radiolabelling followed by HPLC to separate the different phosphatidylinositol monophosphates, or by high-sensitive liquid chromatography coupled to MS,which is still under development. In the present study,we describe amass assay to quantify this lipid from various biological samples using the recombinant PtdIns3P 5-kinase, PIKfyve. Using this assay, we show an increase in the mass level of PtdIns3P in mouse and human platelets following stimulation, loss of this lipid in Vps34-deficient yeasts and its relative enrichment in early endosomes isolated from BHK cells. © 2012 The Author(s).


Adam V.,Catholic University of Leuven | Adam V.,CNRS Institute of Pharmacology and Structural Biology | Adam V.,Institute Of Recherches En Technologies Et Science Pour Le Vivant | Moeyaert B.,Catholic University of Leuven | And 9 more authors.
Chemistry and Biology | Year: 2011

Advanced fluorescence imaging, including subdiffraction microscopy, relies on fluorophores with controllable emission properties. Chief among these fluorophores are the photoactivatable fluorescent proteins capable of reversible on/off photoswitching or irreversible green-to-red photoconversion. IrisFP was recently reported as the first fluorescent protein combining these two types of phototransformations. The introduction of this protein resulted in new applications such as super-resolution pulse-chase imaging. However, the spectroscopic properties of IrisFP are far from being optimal and its tetrameric organization complicates its use as a fusion tag. Here, we demonstrate how four-state optical highlighting can be rationally introduced into photoconvertible fluorescent proteins and develop and characterize a new set of such enhanced optical highlighters derived from mEosFP and Dendra2. We present in particular NijiFP, a promising new fluorescent protein with photoconvertible and biphotochromic properties that make it ideal for advanced fluorescence-based imaging applications. © 2011 Elsevier Ltd All rights reserved.


Fron E.,Catholic University of Leuven | Van Der Auweraer M.,Catholic University of Leuven | Moeyaert B.,Catholic University of Leuven | Michiels J.,Catholic University of Leuven | And 5 more authors.
Journal of Physical Chemistry B | Year: 2013

Green-to-red photoconversion is a reaction that occurs in a limited number of fluorescent proteins and that is currently mechanistically debated. In this contribution, we report on our investigation of the photoconvertible fluorescent protein Dendra2 by employing a combination of pump-probe, up-conversion and single photon timing spectroscopic techniques. Our findings indicate that upon excitation of the neutral green state an excited state proton transfer proceeds with a time constant of 3.4 ps between the neutral green and the anionic green states. In concentrated solution we detected resonance energy transfer (25 ps time constant) between green and red monomers. The time-resolved emission spectra suggest also the formation of a super-red species, first observed for DsRed (a red fluorescent protein from the corallimorph species Discosoma) and consistent with peculiar structural details present in both proteins. © 2013 American Chemical Society.


Caudron F.,ETH Zurich | Denarier E.,Joseph Fourier University | Denarier E.,Institute Of Recherches En Technologies Et Science Pour Le Vivant | Thibout-Quintana J.-C.,Center Hospitalier Lyon Sud | And 5 more authors.
PLoS ONE | Year: 2010

Ser172 of β tubulin is an important residue that is mutated in a human brain disease and phosphorylated by the cyclin-dependent kinase Cdk1 in mammalian cells. To examine the role of this residue, we used the yeast S. cerevisiae as a model and produced two different mutations (S172A and S172E) of the conserved Ser172 in the yeast b tubulin Tub2p. The two mutants showed impaired cell growth on benomyl-containing medium and at cold temperatures, altered microtubule (MT) dynamics, and altered nucleus positioning and segregation. When cytoplasmic MT effectors Dyn1p or Kar9p were deleted in S172A and S172E mutants, cells were viable but presented increased ploidy. Furthermore, the two b tubulin mutations exhibited synthetic lethal interactions with Bik1p, Bim1p or Kar3p, which are effectors of cytoplasmic and spindle MTs. In the absence of Mad2p-dependent spindle checkpoint, both mutations are deleterious. These findings show the importance of Ser172 for the correct function of both cytoplasmic and spindle MTs and for normal cell division. © 2010 Caudron et al.


Cherradi N.,French Institute of Health and Medical Research | Cherradi N.,Institute Of Recherches En Technologies Et Science Pour Le Vivant | Cherradi N.,University Grenoble Alpes
Frontiers in Endocrinology | Year: 2016

Adrenocortical carcinoma (ACC) is a rare malignancy with poor prognosis and limited therapeutic options. Over the last decade, pan-genomic analyses of genetic and epigenetic alterations and genome-wide expression profile studies allowed major advances in the understanding of the molecular genetics of ACC. Besides the well-known dysfunctional molecular pathways in adrenocortical tumors, such as the IGF2 pathway, the Wnt pathway, and TP53, high-throughput technologies enabled a more comprehensive genomic characterization of adrenocortical cancer. Integration of expression profile data with exome sequencing, SNP array analysis, methylation, and microRNA (miRNA) profiling led to the identification of subgroups of malignant tumors with distinct molecular alterations and clinical outcomes. miRNAs post-transcriptionally silence their target gene expression either by degrading mRNA or by inhibiting translation. Although our knowledge of the contribution of deregulated miRNAs to the pathogenesis of ACC is still in its infancy, recent studies support their relevance in gene expression alterations in these tumors. Some miRNAs have been shown to carry potential diagnostic and prognostic values, while others may be good candidates for therapeutic interventions. With the emergence of disease-specific blood-borne miRNAs signatures, analyses of small cohorts of patients with ACC suggest that circulating miRNAs represent promising non-invasive biomarkers of malignancy or recurrence. However, some technical challenges still remain, and most of the miRNAs reported in the literature have not yet been validated in sufficiently powered and longitudinal studies. In this review, we discuss the current knowledge regarding the deregulation of tumor-associated and circulating miRNAs in ACC patients, while emphasizing their potential significance in pathogenic pathways in light of recent insights into the role of miRNAs in shaping the tumor microenvironment. © 2016 Cherradi.


Connorton J.M.,University of Manchester | Connorton J.M.,Institute Of Recherches En Technologies Et Science Pour Le Vivant | Webster R.E.,University of Manchester | Cheng N.,Baylor College of Medicine | Pittman J.K.,University of Manchester
PLoS ONE | Year: 2012

Cation/H+ exchangers encoded by CAX genes play an important role in the vacuolar accumulation of metals including Ca2+ and Mn2+. Arabidopsis thaliana CAX1 and CAX3 have been previously shown to differ phylogenetically from CAX2 but the physiological roles of these different transporters are still unclear. To examine the functions and the potential of redundancy between these three cation transporters, cax1/cax2 and cax2/cax3 double knockout mutants were generated and compared with wild type and cax single knockouts. These double mutants had equivalent metal stress responses to single cax mutants. Both cax1 and cax1/cax2 had increased tolerance to Mg stress, while cax2 and cax2/cax3 both had increased sensitivity to Mn stress. The cax1/cax2 and cax2/cax3 mutants did not exhibit the deleterious developmental phenotypes previously seen with the cax1/cax3 mutant. However, these new double mutants did show alterations in seed germination, specifically a delay in germination time. These alterations correlated with changes in nutrient content within the seeds of the mutants, particularly the cax1/cax2 mutant which had significantly higher seed content of Ca and Mn. This study indicates that the presence of these Arabidopsis CAX transporters is important for normal germination and infers a role for CAX proteins in metal homeostasis within the seed. © 2012 Connorton et al.

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