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J F Briat

Publications and source records attributed to J F Briat.

At least 37 records · Page 2Linked to original sources
PubMed1996

Post-transcriptional regulation of plant ferritin accumulation in response to iron as observed in the maize mutant ys1.

The maize mutant ys1 accumulates iron in leaves to a lower extent than a Fe-efficient genotype. In this mutant, ferritin mRNA accumulates in response to iron to a similar level as in other genotypes. However, ferritin protein and mRNA abundance does not correlate in ys1 leaves, demonstrating that iron also controls plant ferritin protein accumulation at the post-transcriptional level.

Ferric Compounds↗
PubMed1996

Characterization of a ferritin mRNA from Arabidopsis thaliana accumulated in response to iron through an oxidative pathway independent of abscisic acid.

A ferritin cDNA, AtFer1, from seedlings of Arabidopsis thaliana has been characterized. The deduced amino acid sequence of the AtFer1 protein indicates that A. thaliana ferritin shares the same characteristics as the plant ferritin already characterized from the Leguminosae and Graminacea families: (i) it contains an additional sequence in its N-terminal part composed of two domains: a transit peptide responsible for plastid targeting and an extension peptide; (ii) amino acids that form the ferroxidase centre of H-type animal ferritin, as well as Glu residues characteristic of L-type animal ferritin, are conserved in AtFer1; (iii) the C-terminal part of the A. thaliana ferritin subunit defining the E-helix is divergent from its animal counterpart, and confirms that 4-fold-symmetry axis channels are hydrophilic in plant ferritin. Southern blot experiments indicate that AtFer1 is likely to be encoded by a unique gene in the A. thaliana genome, although a search in the NCBI dbEST database indicates that other ferritin genes, divergent from AtFer1, may exist. Iron loading of A. thaliana plantlets increased ferritin mRNA and protein abundance. In contrast to maize, the transcript abundance of a gene responding to abscisic acid (RAB18) did not increase in response to iron loading treatment, and A. thaliana ferritin mRNA abundance is not accumulated in response to a treatment with exogenous abscisic acid, at least in the culture system used in this study. In addition, iron-induced increases in ferritin mRNA abundance were the same as wild-type plants in abi1 and abi2 mutants of A. thaliana, both affected in the abscisic acid response in vegetative tissues. Increased AtFer1 transcript abundance in response to iron is inhibited by the antioxidant N-acetylcysteine. These results indicate that an oxidative pathway, independent of abscisic acid, could be responsible for the iron induction of ferritin synthesis in A. thaliana.

Abscisic Acid↗
PubMed1995

Structure and differential expression of two maize ferritin genes in response to iron and abscisic acid.

In plants, synthesis of the iron-storage protein ferritin in response to iron is not regulated at the translational level; this is in contrast to ferritin synthesis in animals. Part of the response is mediated through a transduction pathway which involves the plant hormone abscisic acid. In this work, we report the cloning and sequencing of two maize ferritin genes (ZmFer1 and ZmFer2) coding for members of the two ferritin mRNA subclasses, FM1 and FM2, respectively. Although plant and animal ferritins are closely related proteins, a major difference is observed between the organisation of the genes. Both maize ferritin genes are organised as eight exons and seven introns, the positions of which are identical within the two genes, while animal ferritin genes are interrupted by three introns, at positions different from those found in maize genes. Sequence divergence between the 3' untranslated regions of these genes has allowed the use of specific probes to study the accumulation of FM1 and FM2 transcripts in response to various environmental cues. Such probes have shown that FM1 and FM2 transcripts accumulate with differential kinetics in response to iron; FM1 mRNA accumulate earlier than FM2 mRNA and only FM2 transcripts accumulate in response to exogenous abscisic acid or water stress. Mapping of the transcriptional initiation region of these two genes defined their 5' upstream regions and allowed a sequence comparison of their promoters, which appeared highly divergent. This raises the possibility that the differential accumulation of FM1 and FM2 mRNAs in response to iron, abscisic acid and drought could be due to differential transcription of ZmFer1 and ZmFer2.

Abscisic Acid↗
PubMed1995

Conformational changes and in vitro core-formation modifications induced by site-directed mutagenesis of the specific N-terminus of pea seed ferritin.

Plant ferritin has a three-dimensional structure predicted to be very similar to that of animal ferritin. It has, however, an additional specific sequence of 24 amino acids at its N-terminus named extension peptide (EP). In order to determine precisely the interactions between EP and other domains of the pea seed ferritin subunit, three point mutations were performed. The mutated residues were chosen by three-dimensional computer modelling of the pea seed ferritin subunit structure [Lobréaux, Yewdall, Briat and Harrison (1992) Biochem. J. 228, 931-939]. The mutant recombinant proteins were expressed in Escherichia coli and purified to homogeneity; all the mutants were found to be assembled as 24-mers. When Ala-13 was replaced by His, as in mammalian ferritins, ferroxidase activity was significantly reduced. Moreover, in vitro iron-core formation in Pro-X-->Ala, Lys-R-->Glu and Ala-13-->His mutants was increased after denaturation by urea followed by renaturation; this was also observed with the EP deletion mutant (r delta TP/EP). The recombinant ferritins were also analysed using tryptophan fluorescence spectra. The r delta TP/EP, Pro-X-->Ala and Lys-R-->Glu mutants were found to be more susceptible to denaturation by urea than the native r delta TP pea seed ferritin.

Amino Acid Sequence↗
PubMed1995

Purification and characterization of recombinant pea-seed ferritins expressed in Escherichia coli: influence of N-terminus deletions on protein solubility and core formation in vitro.

Plant ferritin subunits are synthesized as precursor molecules; the transit peptide (TP) in their NH2 extremity, responsible for plastid targeting, is cleaved during translocation to this compartment. In addition, the N-terminus of the mature subunit contains a plant-specific sequence named extension peptide (EP) [Ragland, Briat, Gagnon, Laulhère, Massenet, and Theil, E.C. (1990) J. Biol. Chem. 265, 18339-18344], the function of which is unknown. A novel pea-seed ferritin cDNA, with a consensus ferroxidase centre conserved within H-type animal ferritins has been characterized. This pea-seed ferritin cDNA has been engineered using oligonucleotide-directed mutagenesis to produce DNA fragments (1) corresponding to the wild-type (WT) ferritin precursor, (2) with the TP deleted, (3) with both the TP and the plant specific EP sequences deleted and (4) containing the TP but with the EP deleted. These four DNA fragments have been cloned in an Escherichia coli expression vector to produce the corresponding recombinant pea-seed ferritins. Expression at 37 degrees C led to the accumulation of recombinant pea-seed ferritins in inclusion bodies, whatever the construct introduced in E. coli. Expression at 25 degrees C in the presence of sorbitol and betaine allowed soluble proteins to accumulate when constructs with the TP deleted were used; under this condition, E. coli cells transformed with constructs containing the TP were unable to accumulate recombinant protein. Recombinant ferritins purified from inclusion bodies were found to be assembled only when the TP was deleted; however assembled ferritin under this condition had a ferroxidase activity undetectable at acid pH. On the other hand, soluble recombinant ferritins with the TP deleted and expressed at 25 degrees C were purified as 24-mers containing an average of 40-50 iron atoms per molecule. Despite the conservation in the plant ferritin subunit of a consensus ferroxidase centre, the iron uptake activity in vitro at pH 6.8 was found to be lower than that of the recombinant human H-ferritin, though it was much more active than the recombinant human L-ferritin. The recombinant ferritin with both the TP and the EP deleted (r delta TP/EP) assembled correctly as a 24-mer; it has slightly higher ferroxidase activity and decreased solubility compared with the wild-type protein with the TP deleted (r delta TP). In addition, on denaturation by urea followed by renaturation by dialysis the r delta TP/EP protein showed a 25% increase in core-formation in vitro compared with the r delta TP protein.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1995

Formation of the ferritin iron mineral occurs in plastids.

Ferritin in plants is a nuclear-encoded, multisubunit protein found in plastids; an N-terminal transit peptide targets the protein to the plastid, but the site for formation of the ferritin Fe mineral is unknown. In biology, ferritin is required to concentrate Fe to levels needed by cells (approximately 10(-7) M), far above the solubility of the free ion (10(-18) M); the protein directs the reversible phase transition of the hydrated metal ion in solution to hydrated Fe-oxo mineral. Low phosphate characterizes the solid-phase Fe mineral in the center of ferritin of the cytosolic animal ferritin, but high phosphate is the hallmark of Fe mineral in prokaryotic ferritin and plant (pea [Pisum sativum L.] seed) ferritin. Earlier studies using x-ray absorption spectroscopy showed that high concentrations of phosphate present during ferritin mineralization in vivo altered the local structure of Fe in the ferritin mineral so that it mimicked the prokaryotic type, whether the protein was from animals or bacteria. The use of x-ray absorption spectroscopy to analyze the Fe environment in pea-seed ferritin now shows that the natural ferritin mineral in plants has an Fe-P interaction at 3.26A, similar to that of bacterial ferritin; phosphate also prevented formation of the longer Fe-Fe interactions at 3.5A found in animal ferritins or in pea-seed ferritin reconstituted without phosphate. Such results indicate that ferritin mineralization occurs in the plastid, where the phosphate content is higher; a corollary is the existence of a plastid Fe uptake system to allow the concentration of Fe in the ferritin mineral.

Biological Transport↗
PubMed1993

Iron release and uptake by plant ferritin: effects of pH, reduction and chelation.

Ferritins are iron-storage proteins that accumulate in plastids during seed formation, and also in leaves during senescence or iron overload. Iron release from ferritins occurs during growth of seedlings and greening of plastids. Depending on the concentration of the reducing agent ascorbate, either an overall iron release or uptake by ferritins from iron(III) citrate may occur. We have designed methods to measure these simultaneous and independent uptake and release fluxes. Each individual step of the exchange was studied using different iron chelates and an excess of ligand. It is shown that: (i) the chelated form of iron, and not ionic Fe3+, is the substrate for iron reduction, which controls the subsequent uptake by ferritin; (ii) iron uptake by ferritins is faster at pH 8.4 than at pH 7 or 6 and is inhibited by an excess of strongly binding free ligands; and (iii) strongly binding free ligands are inhibitory during iron release by ascorbate. When reactions are allowed to proceed simultaneously, the iron chelating power is shown to be a key factor in the overall exchange. The interactions of iron chelating power, reducing capacity and pH are discussed with regard to their influence on the biochemical mobilization of iron.

Ascorbic Acid↗
PubMed1993

Structure and composition of ferritin cores from pea seed (Pisum sativum).

Iron cores from native pea seed (Pisum sativum) ferritin have been analysed by electron microscopy and Mössbauer spectroscopy and shown to be amorphous. This correlates with their relatively high phosphate content (Fe: P = 2.83; 1800 Fe, 640 P atoms/molecule). Reconstituted cores obtained by adding iron (2000 Fe atoms/molecule) in the absence of phosphate to pea seed apoferritin were crystalline ferrihydrite. In vitro rates of formation of pea-seed ferritin iron cores were intermediate between those of recombinant human H-chain and horse spleen apoferritin and this may reflect the amino-acid residues of its ferroxidase and putative nucleation centres. The high phosphate content of pea-seed ferritin suggests that this molecule could be involved in both phosphorus and iron storage. The high phosphate concentration found within plastids, from which the molecules were isolated, is a possible source of the ferritin phosphate.

Fabaceae↗
PubMed1993

Abscisic acid is involved in the iron-induced synthesis of maize ferritin.

The ubiquitous iron storage protein ferritin has a highly conserved structure in plants and animals, but a distinct cytological location and a different level of control in response to iron excess. Plant ferritins are plastid-localized and transcriptionally regulated in response to iron, while animal ferritins are found in the cytoplasm and have their expression mainly controlled at the translational level. In order to understand the basis of these differences, we developed hydroponic cultures of maize plantlets which allowed an increase in the intracellular iron concentration, leading to a transient accumulation of ferritin mRNA and protein (Lobréaux,S., Massenet,O. and Briat,J.F., 1992, Plant Mol. Biol., 19, 563-575). Here, it is shown that iron induces ferritin and RAB (Responsive to Abscisic Acid) mRNA accumulation relatively with abscisic acid (ABA) accumulation. Ferritin mRNA also accumulates in response to exogenous ABA. Synergistic experiments demonstrate that the ABA and iron responses are linked, although full expression of the ferritin genes cannot be entirely explained by an increase in ABA concentration. Inducibility of ferritin mRNA accumulation by iron is dramatically decreased in the maize ABA-deficient mutant vp2 and can be rescued by addition of exogenous ABA, confirming the involvement of ABA in the iron response in plants. Therefore, it is concluded that a major part of the iron-induced biosynthesis of ferritin is achieved through a pathway involving an increase in the level of the plant hormone ABA. The general conclusion of this work is that the synthesis of the same protein in response to the same environmental signal can be controlled by separate and distinct mechanisms in plants and animals.

Abscisic Acid↗
PubMed1993

Uptake of iron from ferric-citrate in the cyanobacteria Synechocystis PCC6803.

Iron is an essential element for bacterial growth. Its intracellular concentration is mainly controlled at the uptake level, which is mediated by iron-dicitrate transport in various prokaryotes. We report here that the cyanobacterium Synechocystis PCC6803 takes up iron from ferric citrate. This process is dependent of the iron:citrate ratio, with a better efficiency for the highest concentration of citrate. The iron-citrate complex is the substrate of this active uptake, inhibited in part by the uncoupler FCCP. Iron starvation activates this uptake. These data suggest the existence of a specific ferric citrate receptor on the cyanobacterial membrane.

Biological Transport, Active↗
PubMed1992

Amino-acid sequence and predicted three-dimensional structure of pea seed (Pisum sativum) ferritin.

The iron storage protein, ferritin, is widely distributed in the living kingdom. Here the complete cDNA and derived amino-acid sequence of pea seed ferritin are described, together with its predicted secondary structure, namely a four-helix-bundle fold similar to those of mammalian ferritins, with a fifth short helix at the C-terminus. An N-terminal extension of 71 residues contains a transit peptide (first 47 residues) responsible for plastid targetting as in other plant ferritins, and this is cleaved before assembly. The second part of the extension (24 residues) belongs to the mature subunit; it is cleaved during germination. The amino-acid sequence of pea seed ferritin is aligned with those of other ferritins (49% amino-acid identity with H-chains and 40% with L-chains of human liver ferritin in the aligned region). A three-dimensional model has been constructed by fitting the aligned sequence to the coordinates of human H-chains, with appropriate modifications. A folded conformation with an 11-residue helix is predicted for the N-terminal extension. As in mammalian ferritins, 24 subunits assemble into a hollow shell. In pea seed ferritin, its N-terminal extension is exposed on the outside surface of the shell. Within each pea subunit is a ferroxidase centre resembling those of human ferritin H-chains except for a replacement of Glu-62 by His. The channel at the 4-fold-symmetry axes defined by E-helices, is predicted to be hydrophilic in plant ferritins, whereas it is hydrophobic in mammalian ferritins.

Amino Acid Sequence↗
PubMed1992

Purification, characterization and function of bacterioferritin from the cyanobacterium Synechocystis P.C.C. 6803.

Storage and buffering of iron is achieved by a class of proteins, the ferritins, widely distributed throughout the living kingdoms. All ferritins have in common their three-dimensional structure and their ability to store large amounts of iron in their central cavity. However, eukaryotic ferritins from plants and animals and bacterioferritins have no sequence similarity, and besides non-haem iron bacterioferritins contain haem residues whereas eukaryotic ferritins do not. In this paper we report the first purification and characterization of a bacterioferritin from a cyanobacterium. It has a molecular mass of 400 kDa and is built up from 19 kDa subunits. Its N-terminal sequence shows 73% identity with that of the Escherichia coli bacterioferritin subunit. It contains 2300 atoms of iron and 1500 molecules of phosphate per ferritin molecule and 0.25 haem residue per subunit; the alpha-peak of the cytochrome has its maximum at 559 nm. In contrast with what is known for eukaryotic ferritins, we found that bacterioferritin from Synechocystis is not inducible by iron under the conditions that we have tested and that it has a constant concentration whatever the iron status of the cells, even at very low iron concentration. Bacterioferritin from Synechocystis P.C.C. 6803 is fully assembled in vivo and it is shown by labelling with 59Fe that it is able to load iron in vitro as well as in vivo. Bacterioferritin from Synechocystis is shown to have an iron-buffering function while the bulk of cellular iron is found associated with a pool of low-molecular-mass electronegative molecules. The role of Synechocystis bacterioferritin in iron metabolism is discussed.

Amino Acid Sequence↗
PubMed1992

Iron induces ferritin synthesis in maize plantlets.

The iron-storage protein ferritin has been purified to homogeneity from maize seeds, allowing to determine the sequence of the first 29 NH2-terminal amino acids of its subunit and to raise specific rabbit polyclonal antibodies. Addition of 500 microM Fe-EDTA/75 microM Fe-citrate to hydroponic culture solutions of maize plantlets, previously starved for iron, led to a significant increase of the iron concentration of roots and leaves, albeit root iron was mainly found associated with the apoplast. Immunodetection of ferritin by western blots indicated that this iron treatment induced ferritin protein accumulation in roots and leaves over a period of 3 days. In order to investigate this induction at the ferritin mRNA level, various ferritin cDNA clones were isolated from a cDNA library prepared from poly(A)+ mRNA isolated from roots 48 h after iron treatment. These cDNAs were classified into two groups called FM1 and FM2. Upstream of the sequence encoding the mature ferritin subunit, both of these cDNAs contained an in-frame coding sequence with the characteristics of a transit peptide for plastid targeting. Two members of the FM1 subfamily, both partial at their 5' extremity, were characterized. They are identical, except in their 3' untranslated region: FM1A extends 162 nucleotides beyond the 3' terminus of FM1B. These two mRNAs could arise from the use of two different polyadenylation signals. FM2 is 96% identical to FM1 and contains 45 nucleotides of 5' untranslated region. Northern analyses of root and leaf RNAs, at different times after iron treatment, revealed ferritin mRNA accumulation in response to iron. Ferritin mRNA accumulation was transient and particularly abundant in leaves, reaching a maximum at 24 h. The level of ferritin mRNA in roots was affected to a lesser extent than in leaves.

Amino Acid Sequence↗
PubMed1991

Ferritin gene transcription is regulated by iron in soybean cell cultures.

Iron-regulated ferritin synthesis in animals is dominated by translational control of stored mRNA; iron-induced transcription of ferritin genes, when it occurs, changes the subunit composition of ferritin mRNA and protein and is coupled to translational control. Ferritins in plants and animals have evolved from a common progenitor, based on the similarity of protein sequence; however, sequence divergence occurs in the C termini; structure prediction suggests that plant ferritin has the E-helix, which, in horse ferritin, forms a large channel at the tetrameric interface. In contemporary plants, a transit peptide is encoded by ferritin mRNA to target the protein to plastids. Iron-regulated synthesis of ferritin in plants and animals appears to be very different since the 50- to 60-fold increases of ferritin protein, previously observed to be induced by iron in cultured soybean cells, is accompanied by an equivalent accumulation of hybridizable ferritin mRNA and by increased transcription of ferritin genes. Ferritin mRNA from iron-induced cells and the constitutive ferritin mRNA from soybean hypocotyls are identical. The iron-induced protein is translocated normally to plastids. Differences in animal ferritin structure coincide with the various iron storage functions (reserve for iron proteins and detoxification). In contrast, the constancy of structure of soybean ferritin, iron-induced and constitutive, coupled with the potential for vacuolar storage of excess iron in plants suggest that rapid synthesis of ferritin from a stored ferritin mRNA may not be needed in plants for detoxification of iron.

Amino Acid Sequence↗
PubMed1991

Ferritin accumulation and degradation in different organs of pea (Pisum sativum) during development.

Iron concentration and ferritin distribution have been determined in different organs of pea (Pisum sativum) during development under conditions of continuous iron supply from hydroponic cultures. No ferritin was detected in total protein extracts from roots or leaves. However, a transient iron accumulation in the roots, which corresponds to an increase in iron uptake, was observed when young fruits started to develop. Ferritin was detectable in total protein extracts of flowers and pods, and it accumulated in seeds. In seeds, the same relative amount of ferritin was detected in cotyledons and in the embryo axis. In cotyledons, ferritin and iron concentration decrease progressively during the first week of germination. Ferritin in the embryo axis was processed, and disappeared, during germination, within the first 4 days of radicle and epicotyl growth. This degradation of ferritin in vivo was marked by a shortening of a 28 kDa subunit, giving 26.5 and 25 kDa polypeptides, reminiscent of the radical damage occurring in pea seed ferritin during iron exchange in vitro [Laulhere, Laboure & Briat (1989) J. Biol. Chem. 264, 3629-3635]. Developmental control of iron concentration and ferritin distribution in different organs of pea is discussed.

Fabaceae↗
PubMed1990

Purification and characterization of an iron-induced ferritin from soybean (Glycine max) cell suspensions.

Ferric citrate induces ferritin synthesis and accumulation in soybean (Glycine max) cell suspension cultures [Proudhon, Briat & Lescure (1989) Plant Physiol. 90, 586-590]. This iron-induced ferritin has been purified from cells grown for 72 h in the presence of either 100 microM- or 500 microM-ferric citrate. It has a molecular mass of about 600 kDa and is built up from a 28 kDa subunit which is recognized by antibodies raised against pea (Pisum sativum) seed ferritin and it has the same N-terminal sequence as this latter, except for residue number 3, which is alanine in pea seed ferritin instead of valine in iron-induced soybean cell ferritin. It contains an average of 1800 atoms of iron per molecule whatever the ferric citrate concentration used to induce its synthesis. It is shown that the presence of 100 microM- or 500 microM-ferric citrate in the culture medium leads respectively to an 11- and 28-fold increase in the total intracellular iron concentration and to a 30- and 60-fold increase in the ferritin concentration. However, the percentage of iron stored in the mineral core of ferritin remains constant whatever the ferric citrate concentration used and represents only 5-6% of cellular iron.

Amino Acid Sequence↗
PubMed1990

Evidence for conservation of ferritin sequences among plants and animals and for a transit peptide in soybean.

Ferritin is a large multisubunit protein that stores iron in plants, animals, and bacteria. In animals, the protein is mainly cytoplasmic and is highly conserved, while in plants ferritin is found in chloroplasts and other plastids. Ferritin is synthesized in plants as a larger precursor of the mature subunit. There is no sequence information for ferritin from plants, except an NH2-terminal peptide of 35 residues which shows little similarity to any known ferritin sequences or transit peptides (Laulhere, J. P., Laboure, A. M., and Briat, J. F. (1989) J. Biol. Chem. 264, 3629-3635). To understand the genetic origin and the location of ferritin synthesis in plant cells, as well as the structure of ferritin from plants, we have sequenced both CNBr peptides from pea seed ferritin and nucleotides of a soybean hypocotyl ferritin cDNA, identified using a frog ferritin cDNA as a probe. Comparison of pea and soybean sequences showed an identity of 89%. Alignment of the plant ferritin sequences with animal ferritins showed 55-65% sequence identity in the common regions. However, a peptide of 28 amino acids extended the NH2 terminus of the plant ferritins. Furthermore, the cDNA encoded additional amino acids which appear to be a transit peptide. None of the sequences in soybean ferritin were found in the tobacco chloroplast genome, suggesting, as does the transit peptide, a nuclear location of ferritin gene(s) in plants. Plant ferritin mRNA is 400-500 nucleotides longer than animal ferritin mRNAs, a difference accounted for in part by the extra peptides encoded. The size of soybean ferritin mRNA was constant in different tissues but expression varied in different tissues (leaf greater than hypocotyl). Thus, higher plants and animal ferritins display sequence homology and differential tissue expression. An ancient, common progenitor apparently gave rise to contemporary eukaryotic ferritins after specific modifications, e.g. transport to plasmids.

Amino Acid Sequence↗