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See detailCharacterization of FRD3, a zinc and iron homeostasis actor in Arabidopsis relatives
Scheepers, Maxime ULg; Charlier, Jean Benoit; Spielmann, Julien ULg et al

Conference (2016, July 14)

Zinc and iron are two essential micronutrients for plants. The homeostasis networks of the two metals are intertwined. Arabidopsis halleri is a zinc- and cadmium-tolerant and zinc-hyperaccumulating ... [more ▼]

Zinc and iron are two essential micronutrients for plants. The homeostasis networks of the two metals are intertwined. Arabidopsis halleri is a zinc- and cadmium-tolerant and zinc-hyperaccumulating species, which also present adaptation of its iron homeostasis(1,4). Transcriptomic studies identified genes which are constitu-tively over-expressed in Arabidopsis halleri compared to Arabidopsis thaliana and which may have a role in metal tolerance or accumulation(2-4). Among them, a candidate gene encodes the FRD3 (FERRIC REDUCTASE DEFECTIVE 3) protein, a member of the MATE family of membrane transporters. FRD3 is a citrate transporter involved in iron homeostasis(5-7) and plays a role in zinc tolerance in A. thaliana(8). The FRD3 gene displays a complex regulation. In A. thaliana, alternative transcript initiation for FRD3 determines two transcripts, which dif-fer 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(9). In A. halleri, a single highly ex-pressed FRD3 transcript with high translation efficiency is present(9). We are further examining the FRD3 function in zinc and iron homeostasis in A. thaliana and A. halleri. We will present data (i) on the high expression of FRD3 in A. halleri, (ii) on the functional characterization of the two alternative FRD3 transcripts and their role in metal homeostasis in A. thaliana in comparison with the A. halleri FRD3 transcript and (iii) on the zinc phenotypes of the frd3 A. thaliana mutant. [less ▲]

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See detailCharacterization of FRD3, a iron and zinc homeostasis actor in Arabidopsis relatives
Scheepers, Maxime ULg; Charlier, Jean-Benoit; Spielmann, Julien ULg et al

Poster (2016, June 09)

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See detailCharacterization of FRD3, a zinc and iron homeostasis actor, in Arabidopsis
Scheepers, Maxime ULg; Charlier, Jean-Benoit; Spielmann, Julien ULg et al

Poster (2016, May 13)

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See detailDynamic Distribution and Interaction of the Arabidopsis SRSF1 Subfamily Splicing Factors
Stankovic, Nancy ULg; Schloesser, Marie ULg; Joris, Marine ULg et al

in Plant Physiology (2016), 170

Serine/Arginine-rich (SR) proteins are essential nucleus-localized splicing factors. Our prior studies showed that Arabidopsis RSZ22, a homolog of the human SRSF7 SR factor, exits the nucleus through two ... [more ▼]

Serine/Arginine-rich (SR) proteins are essential nucleus-localized splicing factors. Our prior studies showed that Arabidopsis RSZ22, a homolog of the human SRSF7 SR factor, exits the nucleus through two pathways, either dependent or independent on the XPO1 receptor. Here, we examined the expression profiles and shuttling dynamics of the Arabidopsis SRSF1 subfamily (SR30, SR34, SR34a and SR34b) under control of their endogenous promoter in Arabidopsis and in transient expression assay. Due to its rapid nucleocytoplasmic shuttling and high expression level in transient assay, we analysed the multiple determinants that regulate the localisation and shuttling dynamics of SR34. By site-directed mutagenesis of SR34 RNA-binding sequences and RS domain, we further show that functional RRM1 or RRM2 are dispensable for the exclusive protein nuclear localization and speckle-like distribution. However, mutations of both RRMs induced aggregation of the protein whereas mutation in the RS domain decreased the stability of the protein and suppressed its nuclear accumulation. Furthermore, the RNA-binding motif mutants are defective for their export through the XPO1 (CRM1/Exportin-1) receptor pathway, but retain nucleocytoplasmic mobility. We performed a yeast two hybrid screen with SR34 as bait and discovered SR45 as a new interactor. SR45 is an unusual SR splicing factor bearing two RS domains. These interactions were confirmed in planta by FLIM-FRET and BiFC and the roles of SR34 domains in protein-protein interactions were further studied. Altogether, our report extends our understanding of shuttling dynamics of Arabidopsis SR splicing factors [less ▲]

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See detailMetal binding to the N-terminal cytoplasmic domain of the PIB ATPase HMA4 is required for metal transport in Arabidopsis.
Laurent, Clémentine ULg; Lekeux, Gilles ULg; Ukuwela, Ashwinie A et al

in Plant Molecular Biology (2016), 90

PIB ATPases are metal cation pumps that transport metals across membranes. These proteins possess N- and C-terminal cytoplasmic extensions that contain Cys- and His-rich high affinity metal binding ... [more ▼]

PIB ATPases are metal cation pumps that transport metals across membranes. These proteins possess N- and C-terminal cytoplasmic extensions that contain Cys- and His-rich high affinity metal binding domains, which may be involved in metal sensing, metal ion selectivity and/or in regulation of the pump activity. The PIB ATPase HMA4 (Heavy Metal ATPase 4) plays a central role in metal homeostasis in Arabidopsis thaliana and has a key function in zinc and cadmium hypertolerance and hyperaccumulation in the extremophile plant species Arabidopsis halleri. <br />Here, we examined the function and structure of the N-terminal cytoplasmic metal-binding domain of HMA4. We mutagenized a conserved CCTSE metal-binding motif in the domain and assessed the impact of the mutations on protein function and localization in planta, on metal-binding properties in vitro and on protein structure by Nuclear Magnetic Resonance spectroscopy. <br />The two Cys residues of the motif are essential for the function, but not for localization, of HMA4 in planta, whereas the Glu residue is important but not essential. These residues also determine zinc coordination and affinity. Zinc binding to the N-terminal domain is thus crucial for HMA4 protein function, whereas it is not required to maintain the protein structure. <br />Altogether, combining in vivo and in vitro approaches in our study provides insights towards the molecular understanding of metal transport and specificity of metal P-type ATPases. [less ▲]

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See detailFunctional characterization of FRD3,  a citrate transporter, in Arabidopsis relatives
Scheepers, Maxime ULg; Charlier, Jean-Benoit; Spielmann, Julien ULg et al

Poster (2015, December 03)

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 detailTowards the identification of cis-elements responsible for the high expression of HMA4, an essential gene for Zn hyperaccumulation in Arabidopsis halleri.
Spielmann, Julien ULg; Nouet, Cécile ULg; Scheepers, Maxime ULg et al

Conference (2015, June 26)

A. halleri is a Zn and Cd hypertolerant and hyperaccumulator plant which stores metals in shoot vacuoles. One key step for hyperaccumulation is a highly efficient root-to-shoot translocation of metals ... [more ▼]

A. halleri is a Zn and Cd hypertolerant and hyperaccumulator plant which stores metals in shoot vacuoles. One key step for hyperaccumulation is a highly efficient root-to-shoot translocation of metals. The Zn/Cd plasmamembrane pump HMA4 (heavy metal ATPase 4) plays a central role in this process and is involved in xylem loading/unloading. Moreover, HMA4 is overexpressed in A. halleri compare to its non-tolerant and non-hyperaccumulator sister, Arabidopsis thaliana. The HMA4 overexpression is determined by a combination of gene triplication and cis-regulatory changes increasing transcription levels of each three HMA4 copies. In this study, we are focusing on the two most active HMA4 promoters and we aim to identify the cis-elements underlying high expression of HMA4 in A. halleri using promoter-reporter constructs. Through deletion analysis, two important regions for a high expression have been identified. Ongoing analysis of more detailed deletions will be presented. [less ▲]

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See detailTowards the identification of mechanisms underlying the high expression of HMA4, an essential gene for zinc hyperaccumulation in Arabidopsis halleri.
Spielmann, Julien ULg; Nouet, Cécile ULg; Scheepers, Maxime ULg et al

Poster (2015, June 08)

To maintain metal homeostasis within physiological limits independently of metal concentrations present in soil, plants developed a complex network including metal uptake, chelation, trafficking, and ... [more ▼]

To maintain metal homeostasis within physiological limits independently of metal concentrations present in soil, plants developed a complex network including metal uptake, chelation, trafficking, and storage processes. In this network, transporters have a central role. To study the molecular basis of zinc homeostasis in plants, we are using A. halleri, a zinc- and cadmium-tolerant and zinc-hyperaccumulating species representing the extreme end of natural variation in terms of metal homeostasis. Indeed, A. halleri stores high amount of metals in vacuoles of above-ground tissues (>1% Zn or 0.01% Cd in shoot dry weight). Cross-species transcriptomic studies identified about thirty genes which are constitutively over-expressed in A. halleri compared to its non-tolerant and non-hyperaccumulator sister species, Arabidopsis thaliana (1-3). HMA4 is one of these genes. It encodes a Zn/Cd plasmamembrane pump involved in xylem loading/unloading which is a key step in the hyperaccumulation process (4-5). The HMA4 overexpression is determined by a combination of gene triplication and cis-regulatory changes increasing transcription levels of each three HMA4 copies (5). We aim to identify the cis-elements underlying high expression of HMA4 in A. halleri using promoter-reporter constructs. [less ▲]

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See detailExpression study of FRD3 in Arabidopsis relatives
Scheepers, Maxime ULg; Charlier, Jean-Benoit ULg; Spielmann, Julien ULg et al

Poster (2015, June 08)

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 ▲]

Detailed reference viewed: 60 (11 ULg)
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 ▲]

Detailed reference viewed: 88 (12 ULg)
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See detailTowards the identification of cis-elements responsible for the high expression of HMA4, an essential gene for zinc hyperaccumulation in Arabidopsis halleri.
Spielmann, Julien ULg; Nouet, Cécile ULg; Scheepers, Maxime ULg et al

Poster (2015, May 13)

A. halleri is a Zn and Cd hypertolerant and hyperaccumulator plant which stores metals in shoot vacuoles. One key step for hyperaccumulation is a highly efficient root-to-shoot translocation of metals ... [more ▼]

A. halleri is a Zn and Cd hypertolerant and hyperaccumulator plant which stores metals in shoot vacuoles. One key step for hyperaccumulation is a highly efficient root-to-shoot translocation of metals. The Zn/Cd plasmamembrane pump HMA4 (heavy metal ATPase 4) plays a central role in this process and is involved in xylem loading/unloading. Moreover, HMA4 is overexpressed in A. halleri compare to its non-tolerant and non-hyperaccumulator sister, Arabidopsis thaliana. The HMA4 overexpression is determined by a combination of gene triplication and cis-regulatory changes increasing transcription levels of each three HMA4 copies. In this study, we are focusing on the two most active HMA4 promoters and we aim to identify the cis-elements underlying high expression of HMA4 in A. halleri using promoter-reporter constructs. Through deletion analysis, two important regions for a high expression have been identified. Ongoing analysis of more detailed deletions will be presented. [less ▲]

Detailed reference viewed: 99 (6 ULg)