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See detailZinc triggers a complex transcriptional and post-transcriptional regulation of the metal homeostasis gene FRD3 in Arabidopsis relatives
Charlier, Jean_Benoit; Polese, Catherine; Nouet, Cécile ULg et al

in Journal of Experimental Botany (2015)

In Arabidopsis thaliana, FRD3 (Ferric Chelate Reductase Defective 3) plays a central role in metal homeostasis. FRD3 is among a set of metal homeostasis genes that are constitutively highly expressed in ... [more ▼]

In Arabidopsis thaliana, FRD3 (Ferric Chelate Reductase Defective 3) plays a central role in metal homeostasis. FRD3 is among a set of metal homeostasis genes that are constitutively highly expressed in roots and shoots of Arabidopsis halleri, a zinc hyperaccumulating and hypertolerant species. Here, we examined the regulation of FRD3 by zinc in both species to shed light on the evolutionary processes underlying the evolution of hyperaccumulation in A. halleri. We combined gene expression studies with the use of GUS and GFP reporter constructs to compare the expression profile, transcriptional and post-transcriptional regulation of FRD3 in both species. The AtFRD3 and AhFRD3 genes display a conserved expression profile. In A. thaliana, alternative transcription initiation sites from two promoters determine transcript variants which are differentially regulated by zinc supply in roots and shoots to favour the most highly translated variant under zinc excess conditions. In A. halleri, a single transcript variant with higher transcript stability and enhanced translation has been maintained. The FRD3 gene thus undergoes complex transcriptional and post-transcriptional regulation in Arabidopsis relatives. Our study reveals that a diverse set of mechanisms underlie increased gene dosage in the A. halleri lineage and illustrates how an environmental challenge can alter gene regulation. [less ▲]

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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), in press

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 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 study of the Ser/Arg-rich splicing factor SRSF5a during zebrafish embryonic development
Joris, Marine ULg; Muller, Marc ULg; Motte, Patrick ULg

Poster (2014, June 05)

Nuclear pre-mRNA splicing is a key process regulating gene expression in eukaryotes. Splicing consists in the removal of introns and the joining of exons within a dynamic macromolecular complex called the ... [more ▼]

Nuclear pre-mRNA splicing is a key process regulating gene expression in eukaryotes. Splicing consists in the removal of introns and the joining of exons within a dynamic macromolecular complex called the spliceosome, which consist of five small nuclear ribonucleoproteins (snRNPs) and numerous non snRNPs proteins (1). Amongst these non snRNPs proteins, the SR proteins family constituted an important group of splicing factors that are involved in constitutive and alternative splicing (2,3). SR proteins are structurally related as they are characterized by one or two RNA-recognition motifs (RRMs) in N-ter and a C-terminal domain enriched in dipeptide Ser/Arg. Phylogenetic inference using the RRM domain allowed us to identify 13 encoding genes for SR proteins in the vertebrate model organism, Danio rerio. The Zebrafish is increasingly recognized as a powerful model for the study of vertebrate embryonic development in a physiological context. The roles of SR splicing factors during animal cell differentiation and development are largely unknown. The aim of the present research is to investigate SR proteins functions during zebrafish development by using molecular and genetic approaches. In this study, we investigated the role of the SR splicing factor SRSF5a. The expression profile was determined by in situ hybridization at 24, 48 and 72 hours post-fertilization and showed SRSF5a expression mainly in brain, retina and pharyngeal arches at these stages. Furthermore, SRSF5a knock-down by morpholinos microinjection strongly suggests an important role of this specific splicing factor during eyes and brain development. In order to gain insight into the molecular function of SRSF5a, we analysed control and morphant transcriptomes using high throughput RNA sequencing. Finally, we use a complementary approach to morpholinos and generate SRSF5a mutant fishes using TALENs (Tal effector nucleases)(4,5). 1. Wahl MC, Will CL, & Luhrmann R (2009) The spliceosome: design principles of a dynamic RNP machine. Cell 136(4):701-718. 2. Long JC & Caceres JF (2009) The SR protein family of splicing factors: master regulators of gene expression. Biochem J 417(1):15-27. 3. Graveley BR (2000) Sorting out the complexity of SR protein functions. RNA 6(9):1197-1211. 4. Moore et al. (2012) Improved Somatic Mutagenesis in Zebrafish Using Transcription Activator-Like Effector Nucleases (TALENs). Plos One 120(1):1-12. 5. Cade et al (2012) Highly efficient generation of heritable zebrafish gene mutations using homo- and heterodimeric TALENs. Nucleic acid research Vol 40, No 16:8001-8010. [less ▲]

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See detailComplex regulation of the FRD3 gene in Arabidopsis relatives
Charlier, Jean Benoit; Polese, Catherine ULg; Nouet, Cécile ULg et al

Poster (2014, March 31)

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See detailFunctional study of the Ser/Arg-rich splicing factor SRSF5a during zebrafish embryonic development.
Joris, Marine ULg; Larbuisson, Arnaud ULg; Muller, Marc ULg et al

Poster (2013, April 18)

To investigate the role of the splicing factor SRSF5a during zebrafish embryonic development, we performed SRSF5a knockdown by morpholino microinjection and we analysed control and morphant transcriptomes ... [more ▼]

To investigate the role of the splicing factor SRSF5a during zebrafish embryonic development, we performed SRSF5a knockdown by morpholino microinjection and we analysed control and morphant transcriptomes using RNA sequencing. [less ▲]

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See detailComplex regulation of the FRD3 gene in Arabidopsis relatives
Charlier, Jean-Benoit ULg; Polese, Catherine ULg; Nouet, Cécile ULg et al

Poster (2013, February 26)

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See detailHard Selective Sweep and Ectopic Gene Conversion in a Gene Cluster Affording Environmental Adaptation
Hanikenne, Marc ULg; Kroymann, Juergen; Trampczynska, Aleksandra et al

in PLoS Genetics (2013), 9

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See detailUse of red autofluorescence for monitoring prodiginine biosynthesis
Tenconi, Elodie ULg; Guichard, Paul; Motte, Patrick ULg et al

in Journal of Microbiological Methods (2013), 93

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See detailInhibition of Tumor Angiogenesis and Growth by a Small-Molecule Multi-FGF Receptor Blocker with Allosteric Properties.
Bono, Francoise; De Smet, Frederik; Herbert, Corentin et al

in Cancer Cell (2013), 23(4), 477-88

Receptor tyrosine kinases (RTK) are targets for anticancer drug development. To date, only RTK inhibitors that block orthosteric binding of ligands and substrates have been developed. Here, we report the ... [more ▼]

Receptor tyrosine kinases (RTK) are targets for anticancer drug development. To date, only RTK inhibitors that block orthosteric binding of ligands and substrates have been developed. Here, we report the pharmacologic characterization of the chemical SSR128129E (SSR), which inhibits fibroblast growth factor receptor (FGFR) signaling by binding to the extracellular FGFR domain without affecting orthosteric FGF binding. SSR exhibits allosteric properties, including probe dependence, signaling bias, and ceiling effects. Inhibition by SSR is highly conserved throughout the animal kingdom. Oral delivery of SSR inhibits arthritis and tumors that are relatively refractory to anti-vascular endothelial growth factor receptor-2 antibodies. Thus, orally-active extracellularly acting small-molecule modulators of RTKs with allosteric properties can be developed and may offer opportunities to improve anticancer treatment. [less ▲]

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See detailPromoter analysis of three HMA4 gene copies in the zinc hyperaccumulator Arabidopsis halleri
Nouet, Cécile ULg; Cebula, Justyna; Motte, Patrick ULg et al

Poster (2012, September 20)

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See detailZinc hyperaccumulation: a model to examine metal homeostasis in plants
Hanikenne, Marc ULg; Nouet, Cécile ULg; Charlier, Jean-Benoit et al

Conference (2012, September 20)

The plant Arabidopsis halleri exhibits naturally selected metal hypertolerance and extraordinarily high levels of leaf metal accumulation. Metal hyperaccumulator species attract interest as they represent ... [more ▼]

The plant Arabidopsis halleri exhibits naturally selected metal hypertolerance and extraordinarily high levels of leaf metal accumulation. Metal hyperaccumulator species attract interest as they represent an extreme end of natural variation of the metal homeostasis network. This might be useful to reveal the global functioning of metal homeostasis networks and uncover key nodes whose alterations can drastically modify metal accumulation and tolerance. In addition, metal hyperaccumulation is a compelling model to study adaptation. In the last few years, major progress has been achieved in our understanding of the mechanisms underlying metal hyperaccumulation in A. halleri. High rates of metal uptake by roots, root-to-shoot translocation and efficient shoot vacuolar sequestration play central roles in determining hyperaccumulation and hypertolerance. Enhanced functions of several metal transporter-encoding genes result from gene copy number amplification and/or (cis)-regulatory changes. We will describe the function of several transporters in zinc and cadmium hyperaccumulation and hypertolerance, and in the adjustment of nutrient homeostasis in A. halleri. Recent work aiming to determine if selection acted during the evolutionary history of A. halleri on a loci required for metal tolerance and accumulation will be presented. [less ▲]

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See detailFunctional study of Arabidopsis thaliana ASF/SF2-like pre-mRNA SR splicing factors
Stankovic, Nancy ULg; Tillemans, Vinciane; Leponce, Isabelle et al

Poster (2012, July 04)

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See detailEvolution of Metal Hyperaccumulation in Arabidopsis halleri
Hanikenne, Marc ULg; Kroymann, Juergen; Bernal, Maria et al

Conference (2012, February 06)

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See detailA Single Ancient Origin for Prototypical Serine/Arginine-Rich Splicing Factors1[W][OA]
Califice, Sophie; Baurain, Denis ULg; Hanikenne, Marc ULg et al

in Plant Physiology (2012), 158(2), 546-560

Eukaryotic pre-mRNA splicing is a process involving a very complex RNA-protein edifice. Serine/arginine-rich (SR) proteins play essential roles in pre-mRNA constitutive and alternative splicing, and have ... [more ▼]

Eukaryotic pre-mRNA splicing is a process involving a very complex RNA-protein edifice. Serine/arginine-rich (SR) proteins play essential roles in pre-mRNA constitutive and alternative splicing, and have been suggested to be crucial in plant-specific forms of developmental regulation and environmental adaptation. Despite their functional importance, little is known about their origin and evolutionary history. SR splicing factors have a modular organization featuring at least one RRM domain and a C-terminal region enriched in Ser/Arg dipeptides. To investigate the evolution of SR proteins, we infer phylogenies for >12,000 RRM domains representing >200 broadly sampled organisms. Our analyses reveal that the RRM domain is not restricted to eukaryotes and that all prototypical SR proteins share a single ancient origin, including the plant-specific SR45 protein. Based on these findings, we propose a scenario for their diversification into four natural families, each corresponding to a main SR architecture, and a dozen subfamilies, of which we profile both sequence conservation and composition. Finally, using operational criteria for computational discovery and classification, we catalogue SR proteins in 20 model organisms, with a focus on green algae and land plants. Altogether, our study confirms the homogeneity and antiquity of SR splicing factors, while establishing robust phylogenetic relationships between animal and plant proteins, which should enable functional analyses of lesser characterized SR family members, especially in green plants. [less ▲]

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