HMA6 and HMA8 are two chloroplast Cu+-ATPases with different enzymatic properties

Sautron, Emeline; Mayerhofer, Hubert; Giustini, Cécile; Pro, Danièle; Crouzy, Serge; Ravaud, Stéphanie; Pebay‐Peyroula, Eva; Rolland, Norbert; Catty, Patrice; Seigneurin-Berny, Daphné

doi:10.1042/bsr20150065

Cited by 23 publications

(26 citation statements)

References 61 publications

(93 reference statements)

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“…As shown in Fig. 5A, all mutants are expressed in intracellular structures similar to the ones described for HMA6 (9). Critically, the addition of GFP has no impact on the phenotype because copper MICs of strains expressing either the mutant or mutant-GFP fusion are identical (Fig.…”

Section: Resultsmentioning

confidence: 73%

“…form of these ATPases induces growth arrest provided that the transported metal is present in the culture medium at the adequate concentration. We have shown that, for HMA6 and HMA8, the phenotype is linked to the localization of the transporters at the ER membrane and toxic accumulation of metal in this compartment (9). The case of HMA6 is particularly illustrative: expression of the active form of the transporter induces yeast growth arrest at 0.5 mM copper, whereas a strain expressing HMA6-AKT (an inactive form of HMA6 mutated in the phosphorylation site) tolerates up to 1.4 mM copper (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Indeed, HMA6 is thought to interact with different chaperones to provide copper to the copper/zinc superoxide dismutase and to HMA8 (5,26). The latter could interact directly with plastocyanin (9). In both case, the interaction between the ATPase and its recipient protein has not been experimentally confirmed.…”

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confidence: 99%

“…So far, the mechanism of copper transfer from HMA8 to plastocyanin is not known. However, combining enzymatic and structural approaches, we proposed that the two proteins could have favorable electrostatic interactions allowing a direct copper transfer without requirement of a metallochaperone (9). As for HMA6 and CCS, the interaction between HMA8 and plastocyanin needs to be experimentally validated.…”

mentioning

confidence: 99%

“…Copper delivery to apoplastocyanin involves two transporters of the P-ATPases family, HMA6 and HMA8, located at the chloroplast envelope (3,4) and the thylakoid membranes (5,6), respectively. Both transporters are high affinity transporters for monovalent copper (7)(8)(9). HMA6 and HMA8 loss of function mutations strongly reduce chloroplast copper content, in the stroma for the hma6 mutant and in thylakoids for the hma8 mutant.…”

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confidence: 99%

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Identification of Two Conserved Residues Involved in Copper Release from Chloroplast PIB-1-ATPases

Sautron

Giustini

Dang

et al. 2016

Journal of Biological Chemistry

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Copper is an essential transition metal for living organisms. In the plant model Arabidopsis thaliana, half of the copper content is localized in the chloroplast, and as a cofactor of plastocyanin, copper is essential for photosynthesis. Within the chloroplast, copper delivery to plastocyanin involves two transporters of the P IB-1 -ATPases subfamily: HMA6 at the chloroplast envelope and HMA8 in the thylakoid membranes. Both proteins are high affinity copper transporters but share distinct enzymatic properties. In the present work, the comparison of 140 sequences of P IB-1 -ATPases revealed a conserved region unusually rich in histidine and cysteine residues in the TMA-L1 region of eukaryotic chloroplast copper ATPases. To evaluate the role of these residues, we mutated them in HMA6 and HMA8. Mutants of interest were selected from phenotypic tests in yeast and produced in Lactococcus lactis for further biochemical characterizations using phosphorylation assays from ATP and P i . Combining functional and structural data, we highlight the importance of the cysteine and the first histidine of the CX 3 HX 2 H motif in the process of copper release from HMA6 and HMA8 and propose a copper pathway through the membrane domain of these transporters. Finally, our work suggests a more general role of the histidine residue in the transport of copper by P IB-1 -ATPases.

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Section: Resultsmentioning

confidence: 73%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Identification of Two Conserved Residues Involved in Copper Release from Chloroplast PIB-1-ATPases

Sautron

Giustini

Dang

et al. 2016

Journal of Biological Chemistry

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Copper‐transporting ATPases: The evolutionarily conserved machineries for balancing copper in living systems

Migocka

2015

IUBMB Life

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Copper ATPases (Cu-ATPases) are ubiquitous transmembrane proteins using energy from ATP to transport copper across different biological membranes of prokaryotic and eukaryotic cells. As they belong to the P-ATPase family, Cu-ATPases contain a characteristic catalytic domain with an evolutionarily conserved aspartate residue phosphorylated by ATP to form a phosphoenzyme intermediate, as well as transmembrane helices containing a cation-binding cysteine-proline-cysteine/histidine/serine (CPx) motif for catalytic activation and cation translocation. In addition, most Cu-ATPases possess the N-terminal Cu-binding CxxC motif required for regulation of enzyme activity. In cells, the Cu-ATPases receive copper from soluble chaperones and maintain intracellular copper homeostasis by efflux of copper from the cell or transport of the metal into the intracellular compartments. In addition, copper pumps play an essential role in cuproprotein biosynthesis by the uptake of copper into the cell or delivery of the metal into the chloroplasts and thylakoid lumen or into the lumen of the secretory pathway, where the metal ion is incorporated into copper-dependent enzymes. In the recent years, significant progress has been made toward understanding the function and regulation of Cu-transporting ATPases in archaea, bacteria, yeast, humans, and plants, providing new insights into the specific physiological roles of these essential proteins in various organisms and revealing some conservative regulatory mechanisms of Cu-ATPase activity. In this review, the structural, biochemical, and functional properties of Cu-ATPases from phylogenetically different organisms are summarized and discussed, with particular attention given to the recent insights into the molecular biology of copper pumps in plants. V C 2015 IUBMB Life, 67(10):737-745, 2015

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The Main Functions of Plastids

Kuntz,

Dimnet,

Pullara

et al. 2024

Methods in Molecular Biology

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HMA6 and HMA8 are two chloroplast Cu+-ATPases with different enzymatic properties

Abstract: In Arabidopsis chloroplasts, HMA8, the thylakoid Cu+-ATPase and HMA6, the envelope Cu+-ATPase, have different enzymatic properties. These differences might be related to the electrostatic properties of the Cu+ release cavities of the transporters or the nature of their Cu acceptor.

Cited by 23 publications

References 61 publications

Identification of Two Conserved Residues Involved in Copper Release from Chloroplast PIB-1-ATPases

Identification of Two Conserved Residues Involved in Copper Release from Chloroplast PIB-1-ATPases

Copper‐transporting ATPases: The evolutionarily conserved machineries for balancing copper in living systems

The Main Functions of Plastids

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