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See detailFunctional amphiphilic and biodegradable copolymers for intravenous vectorization
Van Butsele, Kathy ULg; Jérôme, Robert ULg; Jérôme, Christine ULg

in Polymer (2007), 48

This paper aims at reporting on the design of polymeric drug nanocarriers used in cancer therapy, with a special emphasis on the control of their biodistribution. First, the prominent role of poly ... [more ▼]

This paper aims at reporting on the design of polymeric drug nanocarriers used in cancer therapy, with a special emphasis on the control of their biodistribution. First, the prominent role of poly(ethylene oxide) in the lifetime of nanocarriers circulating in the blood stream is highlighted, and the origin of a passive targeting based on a difference in the anatomy of tumors and normal tissues is discussed. The main body of the review is devoted to the targeting of nanocarriers towards tumors and the underlying concepts. As a rule, either the constitutive polymer is stimuli-responsive and the locus of drug release is where the stimulation occurs, or a ligand endowed with specific recognition is grafted onto the nanocarrier. Finally, the fate of the nanocarrier after drug delivery and the bioelimination of the polymer(s) involved are briefly considered. [less ▲]

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See detailFunctional amphiphilic and degradable copolymers for drug delivery systems
Freichels, Hélène ULg; Pourcelle, Vincent; Plapied, Laurence et al

Poster (2008, December 18)

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See detailFunctional Analysis and the Finite Element Method
Stainier, Laurent; Tossings, Patricia ULg

Learning material (2008)

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See detailFunctional analysis of an FLC-LIKE gene in root chicory
Périlleux, Claire ULg; Pieltain, Alexandra ULg; D'Aloia, Maria ULg et al

in Comparative Biochemistry and Physiology. Part A, Physiology (2009), 153A(2/Suppl.), 198-199

Vernalization is known to promote flowering in Arabidopsis thaliana by inhibiting the expression of a strong repressor: FLOWERING LOCUS C (FLC). The recent cloning of an FLC-LIKE gene in sugar beet (Beta ... [more ▼]

Vernalization is known to promote flowering in Arabidopsis thaliana by inhibiting the expression of a strong repressor: FLOWERING LOCUS C (FLC). The recent cloning of an FLC-LIKE gene in sugar beet (Beta vulgaris; BvFL1) and – here – in root chicory (Cichorium intybus; CiFL1) suggests the conservation of FLC biological function during evolution of eudicots. Hence physiological questions that remain difficult to address in Arabidopsis can be studied in other species. We investigated the correlation between CiFL1 expression and plant-age dependent responsiveness to vernalization. We also studied the effect of post-vernalization growing temperature, which can stabilize or erase the vernalized state. [less ▲]

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See detailFunctional analysis of dual-specificity protein phosphatases in angiogenesis
Amand, Mathieu; ERPICUM, Charlotte ULg; Gilles, Christine ULg et al

in Pulido, Rafael (Ed.) Protein Tyrosine Phosphatases: Methods and Protocols (in press)

Therapeutic perspectives targeting angiogenesis in cancer stimulated an intense investigation of the mechanisms triggering and governing angiogenic processes. Several publications have highlighted the ... [more ▼]

Therapeutic perspectives targeting angiogenesis in cancer stimulated an intense investigation of the mechanisms triggering and governing angiogenic processes. Several publications have highlighted the importance of typical dual-specificity phosphatases (DSPs) or MKPs in endothelial cells and their role in controlling different biological functions implicated in angiogenesis such as migration, proliferation, apoptosis, tubulogenesis and cell adhesion. However, among atypical DSPs, the only one investigated in angiogenesis was DUSP3. We recently identified this DSP as new key player in endothelial cells and angiogenesis. In this chapter we provide with detailed protocols and models used to investigate the role of DUSP3 in endothelial cells and angiogenesis. We start the chapter with an overview of the role of several DSPs in angiogenesis. We continue with providing a full description of a highly efficient transfection protocol to deplete DUSP3 using small interfering RNA (siRNA) in the primary Human Umbilical Vein Endothelial Cells (HUVEC). We next describe the major assays used to investigate different processes involved in angiogenesis such as tube formation assay, proliferation assay and spheroids sprouting assay. We finish the chapter by validating our results in DUSP3-knockout mice using in vivo angiogenesis assays such as Matrigel plug and Lewis lung carcinoma cell subcutaneous xenograft model followed by anti-CD31 immunofluorescence and ex vivo aortic ring assay. All methods described can be adapted to other phosphatases and signaling molecules. [less ▲]

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See detailFunctional analysis of FRD3 in Arabidopsis
Scheepers, Maxime ULg; Charlier, Jean-Benoit ULg; Spielmann, Julien ULg et al

Poster (2015, June)

Zinc and iron are two essential micronutrients for plants. The homeostasis networks of the two metals are intertwined. The FRD3 (FERRIC REDUCTASE DEFECTIVE 3) protein, a member of the MATE family of ... [more ▼]

Zinc and iron are two essential micronutrients for plants. The homeostasis networks of the two metals are intertwined. The FRD3 (FERRIC REDUCTASE DEFECTIVE 3) protein, a member of the MATE family of membrane transporters, is a citrate transporter involved in iron homeostasis and playing a role in zinc tolerance in Arabidopsis. The FRD3 gene displays a complex regulation. Alternative transcript initiation for FRD3 determines two transcripts, which differ in their 5'UTRs and have differential translation efficiency. The two transcripts are selectively regulated under stress conditions: iron and zinc depletion, zinc excess or cadmium presence. We are aiming to determine the FRD3 function in zinc and iron homeostasis in Arabidopsis. We will present data (i) on the functional characterization of the alternative transcripts and their role in metal homeostasis in Arabidopsis and (ii) on the zinc phenotypes of the frd3 mutant. [less ▲]

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See detailFunctional analysis of FRD3 in Arabidopsis relatives
Scheepers, Maxime ULg; Charlier, Jean-Benoit ULg; Motte, Patrick ULg et al

Poster (2014, November 28)

Transcriptomic studies identified genes which are constitutively over-expressed in A. halleri compared to A. thaliana and which may have a role in metal tolerance or accumulation (1-3). A candidate gene ... [more ▼]

Transcriptomic studies identified genes which are constitutively over-expressed in A. halleri compared to A. thaliana and which may have a role in metal tolerance or accumulation (1-3). A candidate gene encodes FRD3, a member of the MATE family of membrane transporters (56 members in A. thaliana). It is a citrate transporter involved in iron homeostasis (4-6) and playing a role in zinc tolerance in A. thaliana (7). We are aiming to analyse the FRD3 high expression in A. halleri and the FRD3 function in zinc and iron homeostasis in A. thaliana. [less ▲]

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See detailFunctional analysis of hydrogen photoproduction in respiratory-deficient mutants of Chlamydomonas reinhardtii
Lecler, Renaud ULg; Godaux, Damien ULg; Vigeolas, Hélène ULg et al

in International Journal of Hydrogen Energy (2011), 36

In this paper, mitochondrial mutants of Chlamydomonas reinhardtii defective for respiratory complex I (NADH:ubiquinone oxidoreductase), complex III (ubiquinol cytochrome c oxidoreductase) and both ... [more ▼]

In this paper, mitochondrial mutants of Chlamydomonas reinhardtii defective for respiratory complex I (NADH:ubiquinone oxidoreductase), complex III (ubiquinol cytochrome c oxidoreductase) and both complexes I and III were analyzed for H2 photoproduction. Several parameters were followed during the S-deficiency stage and the anaerobic stage leading to H2 photoproduction. At the early aerobic S-deficiency stage, starch and neutral lipids accumulated in all strains but their amount was significantly decreased in mutants compared to wild type. During the H2 photoproduction process, whereas starch content strongly decreased in all strains, neutral lipid amount remained nearly unchanged, suggesting that starch degraded by glycolysis is the preferential substrate for energy production during anaerobiosis. The mutants displayed a decrease in H2 photoproduction correlating to the number of active mitochondrial proton-pumping sites lost in the strains. Our results thus highlight the critical role of oxidative phosphorylation during the first (aerobic) stage of S-starvation when carbon resources are accumulated. [less ▲]

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See detailFunctional analysis of protein tyrosine phosphatases in thrombosis and haemostasis
Rahmouni, Souad ULg; Hego, Alexandre ULg; Delierneux, Céline ULg et al

in Protein Tyrosine Phosphatases: Methods and Protocols (in press)

Platelets are small blood cells derived from cytoplasmic fragments of megakaryocytes and play an essential role in thrombosis and haemostasis. Platelet activation depends on the rapid phosphorylation and ... [more ▼]

Platelets are small blood cells derived from cytoplasmic fragments of megakaryocytes and play an essential role in thrombosis and haemostasis. Platelet activation depends on the rapid phosphorylation and dephosphorylation of key signaling molecules, and a number of kinases and phosphatases have been identified as major regulators of platelet function. However, the investigation of novel signaling proteins has suffered from technical limitations due to the anucleate nature of platelets and their very limited levels of mRNA and de novo protein synthesis. In the past, experimental methods were restricted to the generation of genetically modified mice and the development of specific antibodies. More recently, novel (phospho)proteomic technologies and pharmacological approaches using specific small-molecule inhibitors have added additional capabilities to investigate specific platelet proteins. In this chapter, we report methods for using genetic and pharmacological approaches to investigate the function of platelet signaling proteins. While the described experiments focus on the role of the dual-specificity phosphatase 3 (DUSP3) in platelet signaling, the presented methods are applicable to any signaling enzyme. Specifically, we describe a testing strategy that includes 1) aggregation and secretion experiments with mouse and human platelets, 2) immunoprecipitation and immunoblot assays to study platelet signaling events, 3) detailed protocols to use selected animal models in order to investigate thrombosis and haemostasis in vivo, and 4) strategies for utilizing pharmacological inhibitors on human platelets. [less ▲]

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See detailFunctional analysis of the Antirrhinum floral homeotic DEFICIENS gene in vivo and in vitro by using a temperature-sensitive mutant.
Zachgo, S.; Silva, E de A; Motte, Patrick ULg et al

in Development (1995), 121(9), 2861-75

Flowers of the temperature-sensitive DEFICIENS (DEF) mutant, def-101, display sepaloid petals and carpelloid stamens when grown at 26 degrees C, the non-permissive temperature. In contrast, when ... [more ▼]

Flowers of the temperature-sensitive DEFICIENS (DEF) mutant, def-101, display sepaloid petals and carpelloid stamens when grown at 26 degrees C, the non-permissive temperature. In contrast, when cultivated under permissive conditions at 15 degrees C, the morphology of def-101 flowers resembles that of the wild type. Temperature shift experiments during early and late phases of flower development revealed that second and third whorl organ development is differentially sensitive to changes in DEF expression. In addition, early DEF expression seems to control the spatially correct initiation of fourth whorl organ development. Reduction of the def-101 gene dosage differentially affects organogenesis in adjacent whorls: at the lower temperature development of petals in the second whorl and initiation of carpels in the centre of the flower is not affected while third whorl organogenesis follows the mutant (carpelloid) pattern. The possible contribution of accessory factors to organ-specific DEF functions is discussed. In situ analyses of mRNA and protein expression patterns during def-101 flower development at 15 degrees C and at 26 degrees C support previously proposed combinatorial regulatory interactions between the MADS-box proteins DEF and GLOBOSA (GLO), and provide evidence that the autoregulatory control of DEF and GLO expression by the DEF/GLO heterodimer starts after initiation of all organ primordia. Immunolocalisation revealed that both proteins are located in the nucleus. Interestingly, higher growth temperature affects the stability of both the DEF-101 and GLO proteins in vivo. In vitro DNA binding studies suggest that the temperature sensitivity of the def-101 mutant is due to an altered heterodimerisation/DNA-binding capability of the DEF-101 protein, conditioned by the deletion of one amino acid within the K-box, a protein region thought to be involved in protein-protein interaction. In addition, we introduce a mutant allele of GLO, glo-confusa, where insertion of one amino acid impairs the hydrophobic carboxy-terminal region of the MADS-box, but which confers no strong phenotypic changes to the flower. The strong mutant phenotype of flowers of def-101/glo-conf double mutants when grown in the cold represents genetic evidence for heterodimerisation between DEF and GLO in vivo. The potential to dissect structural and functional domains of MADS-box transcription factors is discussed. [less ▲]

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See detailFunctional analysis of the cell division protein FtsW of Escherichia coli
Pastoret, Soumya; Fraipont, Claudine ULg; den Blaauwen, Tanneke et al

in Journal of Bacteriology (2004), 186(24), 8370-8379

Site-directed mutagenesis experiments combined with fluorescence microscopy shed light on the role of Escherichia coli FtsW, a membrane protein belonging to the SEDS family that is involved in ... [more ▼]

Site-directed mutagenesis experiments combined with fluorescence microscopy shed light on the role of Escherichia coli FtsW, a membrane protein belonging to the SEDS family that is involved in peptidoglycan assembly during cell elongation, division, and sporulation. This essential cell division protein has 10 transmembrane segments (TMSs). It is a late recruit to the division site and is required for subsequent recruitment of penicillin-binding protein 3 (PBP3) catalyzing peptide cross-linking. The results allow identification of several domains of the protein with distinct functions. The localization of PBP3 to the septum was found to be dependent on the periplasmic loop located between TMSs 9 and 10. The E240-A249 amphiphilic peptide in the periplasmic loop between TMSs 7 and 8 appears to be a key element in the functioning of FtsW in the septal peptidoglycan assembly machineries. The intracellular loop (containing the R166-FI78 amphiphilic peptide) between TMSs 4 and 5 and Gly 311 in TMS 8 are important components of the amino acid sequence-folding information. [less ▲]

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See detailThe functional analysis of the Mousterian and Micoquian assemblages of Sesselfelsgrotte, Germany. Tool use and Hafting in the European Late Middle Paleolithic
Rots, Veerle ULg

in Quartär : Jahrbuch für Erforschung des Eiszeitalters und seiner Kulturen (2009), 56

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See detailFunctional analysis of the three HMA4 copies of the metal hyperaccumulator Arabidopsis halleri
Nouet, Cécile ULg; Charlier, Jean-Benoit; Carnol, Monique ULg et al

in Journal of Experimental Botany (2015), 66

In Arabidopsis halleri, the AhHMA4 gene has an essential function in Zn/Cd hypertolerance and hyperaccumulation by mediating root to shoot translocation of metals. Constitutive high expression of AhHMA4 ... [more ▼]

In Arabidopsis halleri, the AhHMA4 gene has an essential function in Zn/Cd hypertolerance and hyperaccumulation by mediating root to shoot translocation of metals. Constitutive high expression of AhHMA4 results from a tandem triplication and cis-activation of the promoter of all three copies. The three AhHMA4 copies possess divergent promoter sequences, but highly conserved coding sequences, and display identical expression profiles in the root and shoot vascular system. Here, we expressed an AhHMA4::GFP fusion under the control of each three A. halleri HMA4 promoters in a hma2hma4 double mutant of Arabidopsis thaliana to individually examine the function of each A. halleri AhHMA4 copy. The protein localized non-polarly at the plasma membrane of the root pericycle cells of both A. thaliana and A. halleri. The expression of each AhHMA4::GFP copy complemented the severe Zn deficiency phenotype of the hma2hma4 mutant by restoring root-to-shoot translocation of zinc. However, each copy had different impact on metal homeostasis in the A. thaliana genetic background: AhHMA4 copies 2 and 3 were more highly expressed and provided higher Zn tolerance in roots and accumulation in shoots than copy 1, whereas AhHMA4 copy 3 also increased Cd tolerance in roots. Our data suggest a certain extent of functional differentiation among the three A. halleri HMA4 copies, stemming from differences in expression levels rather than in expression profile. HMA4 is a key node of the Zn homeostasis network and small changes in expression level can have major impact on Zn allocation to root or shoot tissues. [less ▲]

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See detailFunctional analysis of yeast bcs1 mutants highlights the role of Bcs1p-specific amino acids in the AAA domain.
Nouet, Cécile ULg; Truan, Gilles; Mathieu, Lise et al

in Journal of Molecular Biology (2009), 388(2), 252-61

The mitochondrial protein Bcs1p is conserved from Saccharomyces cerevisiae to humans and its C-terminal region exhibits an AAA (ATPases associated with diverse cellular activities) domain. The absence of ... [more ▼]

The mitochondrial protein Bcs1p is conserved from Saccharomyces cerevisiae to humans and its C-terminal region exhibits an AAA (ATPases associated with diverse cellular activities) domain. The absence of the yeast Bcs1p leads to an assembly defect of the iron-sulfur protein (ISP) subunit within the mitochondrial respiratory complex III, whereas human point mutations located all along the protein cause various pathologies. We have performed a structure-function analysis of the yeast Bcs1p by randomly generating a collection of respiratory-deficient point mutants. We showed that most mutations are in the C-terminal region of Bcs1p and have localized them on a theoretical three-dimensional model based on the structure of several AAA proteins. The mutations can be grouped into classes according to their respiratory competence and their location on the three-dimensional model. We have further characterized five mutants, each substituting an amino acid conserved in yeast and mammalian Bcs1 proteins but not in other AAA proteins. The effects on respiratory complex assembly and Bcs1p accumulation were analyzed. Intragenic and extragenic compensatory mutations able to restore complex III assembly to the mutants affecting the AAA domain were isolated. Our results bring new insights into the role of specific residues in critical regions that are also conserved in the human Bcs1p. We show that (1) residues located at the junction between the Bcs1p-specific and the AAA domains are important for the activity and stability of the protein and (2) the residue F342 is important for interactions with other partners or substrate proteins. [less ▲]

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See detailFunctional analysis of ZNF85 KRAB zinc finger protein, a member of the highly homologous ZNF91 family
Poncelet, Dominique A.; Bellefroid, Eric J.; Bastiaens, P. V. et al

in DNA & Cell Biology (1998), 17(11), 931-43

We previously identified the ZNF85 (HPF4) KRAB zinc finger gene, a member of the human ZNF91 family. Here, we show that the ZNF85 gene is highly expressed in normal adult testis, in seminomas, and in the ... [more ▼]

We previously identified the ZNF85 (HPF4) KRAB zinc finger gene, a member of the human ZNF91 family. Here, we show that the ZNF85 gene is highly expressed in normal adult testis, in seminomas, and in the NT2/D1 teratocarcinoma cell line. Immunocytochemical localization of a panel of beta-Gal/ZNF85 fusion proteins revealed that ZNF85 contains at least one nuclear localization signal located in the spacer region connecting the KRAB domain with the zinc finger repeats. Bacterially expressed ZNF85 zinc finger domain bound strongly and exclusively to DNA in vitro in a zinc-dependent manner. The KRAB(A) domain of the ZNF85 protein and of several other members of the ZNF91 family exhibited repressing activity when tested in Gal4 fusion protein assays. The repression was significantly enhanced by the addition of the KRAB (B) domain, whereas further addition of other conserved regions had no effect. The ZNF85 KRAB(A) and (B) domains in vitro bound several nuclear proteins that might constitute critical cofactors for repression. [less ▲]

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See detailThe functional anatomy of inhibition processes investigated with the Hayling task
Collette, Fabienne ULg; Van der Linden, Martial ULg; Delfiore, Guy et al

in Neuroimage (2001), 14(2), 258-267

The cortical areas involved in inhibition processes were examined with positron emission tomography (PET). The tasks administered to subjects were an adaptation of the Hayling test. In the first condition ... [more ▼]

The cortical areas involved in inhibition processes were examined with positron emission tomography (PET). The tasks administered to subjects were an adaptation of the Hayling test. In the first condition (response initiation), subjects had to complete sentences with a word clearly suggested by the context, whereas in the second condition (response inhibition), subjects had to produce a word that made no sense in the context of the sentence. Results indicated that the response initiation processes were associated to increases of activity in the left inferior frontal gyrus (BA 45/47), whereas response inhibition processes led to increases in a network of left prefrontal areas, including the middle (BA 9 and BA 10) and inferior (BA 45) frontal areas. [less ▲]

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See detailFunctional anatomy of verbal and visuospatial span tasks in Alzheimer's disease
Collette, Fabienne ULg; Salmon, Eric ULg; Van der Linden, Martial ULg et al

in Human Brain Mapping (1997), 5(2), 110-118

The aim of the study was to emphasize cerebral regions which subserve the performance of short-term memory tasks in patients with Alzheimer's disease. We correlated scores obtained on span tasks with ... [more ▼]

The aim of the study was to emphasize cerebral regions which subserve the performance of short-term memory tasks in patients with Alzheimer's disease. We correlated scores obtained on span tasks with cerebral metabolism measured at rest with positron emission tomography. Scores obtained on the digit span task correlated with glucose metabolism in a brain area centered on the premotor cortex and extending to the adjacent motor and parietal gyri. There exists some evidence suggesting that this area may subserve the sequential organization of material stored in short-term memory. In a secondary analysis, we also observed significant interregional correlations between left-sided brain areas which are part of the neural network subserving verbal working memory processes in healthy controls. These data suggest that individual performance on verbal span tasks in AD patients may essentially depend on the preservation of their ordination processing capacity. The absence of correlation with prefrontal regions suggests that AD patients might not spontaneously engage central executive resources to reach their maximal span score. For simultaneous visuospatial span task, the performance of patients correlated with posterior brain regions, and not with prefrontal cortices. Hum. Brain Mapping 5:110-118, 1997. © 1997 Wiley-Liss Inc. [less ▲]

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See detailFunctional and cooperative interactions between the homeodomain PDX1, Pbx, and Prep1 factors on the somatostatin promoter
Goudet, Ghylène; Delhalle, Sylvie; Biemar, Frédéric et al

in Journal of Biological Chemistry (1999), 274(7), 4067-73

Expression of the somatostatin gene in endocrine pancreatic cells is controlled by several regulatory cis-elements located in the promoter region. Among these, the adjacent UE-A and TSEI elements, located ... [more ▼]

Expression of the somatostatin gene in endocrine pancreatic cells is controlled by several regulatory cis-elements located in the promoter region. Among these, the adjacent UE-A and TSEI elements, located from -113 to -85 relative to the transcription initiation site, function in combination and act as a pancreas-specific mini-enhancer. The TSEI element is recognized by the pancreatic homeodomain factor PDX1. In the present study, we show that the UE-A element binds a heterodimeric complex composed of a Pbx factor and the Prep1 protein, both belonging to the atypical three-amino acid loop extension homeodomain family. Recombinant Pbx1 and Prep1 proteins bind cooperatively to the UE-A site, whereas neither protein can bind this site alone. Transient transfection experiments reveal that both Pbx1 and Prep1 are required to generate a strong transcriptional activation from the UE-A element when this element is inserted close to the TATA box. In contrast, in the context of the intact somatostatin promoter or mini-enhancer, Pbx1 and Prep1 alone have no effect, but they produce a drastic activation when the pancreatic homeodomain factor PDX1 is also coexpressed. Thus, the activity of the somatostatin mini-enhancer is mediated by a cooperative interaction between the Pbx-Prep1 heterodimeric complex and the pancreatic factor PDX1. [less ▲]

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