Abstract
Key message
We identified key residues of TaHMA2, and the N- and C-terminal regions of the protein have different roles in its transport function when heterologously expressed in yeast.
Abstract
TaHMA2, a P1B-type ATPase from wheat (Triticum aestivum L.), plays an important role in heavy metal homeostasis in plants. A previous study showed that overexpressing TaHMA2 in rice (Oryza sativa L.), Arabidopsis thaliana, or tobacco (Nicotiana tabacum L.) resulted in various responses to heavy metals. Here, we report the heterologous expression of TaHMA2 in the yeast Saccharomyces cerevisiae. TaHMA2 expression increased the yeast’s sensitivity to Cd, but not to Zn, Pb or Co, and increased Cd accumulation was concurrently observed. The eGFP-TaHMA2 fusion protein was localized to the plasma membrane and showed a discontinuous pattern. Mutagenesis of the cysteine and glutamate residues in the N-terminal metal-binding domain (N-MBD) impaired the function of TaHMA2. Deletion of most of the C terminus (TaHMA2ΔC, 712–1003) partially abolished the protein’s function, whereas deletion of the N terminus (TaHMA2ΔN, 2–699) completely abolished Cd sensitivity. These data suggest that cysteine and glutamate residues are important for the metal-binding/translocation function of TaHMA2. Additional studies are needed to further understand the selectivity of TaHMA2 in planta.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Andres-Colas N, Sancenon V, Rodriguez-Navarro S, Mayo S, Thiele DJ, Ecker JR, Puig S, Penarrubia L (2006) The Arabidopsis heavy metal P-type ATPase HMA5 interacts with metallochaperones and functions in copper detoxification of roots. Plant J 45:225–236. doi:10.1111/j.1365-313X.2005.02601.x
Arguello JM (2003) Identification of ion-selectivity determinants in heavy-metal transport P1B-type ATPases. J Membr Biol 195:93–108
Arguello JM, Eren E, Gonzalez-Guerrero M (2007) The structure and function of heavy metal transport P1B-ATPases. Biometals 20:233–248. doi:10.1007/s10534-006-9055-6
Arguello JM, Gonzalez-Guerrero M, Raimunda D (2011) Bacterial transition metal P1B-ATPases: transport mechanism and roles in virulence. Biochemistry 50:9940–9949. doi:10.1021/bi201418k
Axelsen KB, Palmgren MG (1998) Evolution of substrate specificities in the P-type ATPase superfamily. J Mol Evol 46:84–101
Baekgaard L, Mikkelsen MD, Sorensen DM, Hegelund JN, Persson DP, Mills RF, Yang Z, Husted S, Andersen JP, Buch-Pedersen MJ, Schjoerring JK, Williams LE, Palmgren MG (2010) A combined zinc/cadmium sensor and zinc/cadmium export regulator in a heavy metal pump. J Biol Chem 285:31243–31252. doi:10.1074/jbc.M110.111260
Bal N, Wu CC, Catty P, Guillain F, Mintz E (2003) Cd2+ and the N-terminal metal-binding domain protect the putative membranous CPC motif of the Cd2+-ATPase of Listeria monocytogenes. Biochem J 369:681–685. doi:10.1042/bj20021416
Banci L, Bertini I, Ciofi-Baffoni S, Su XC, Miras R, Bal N, Mintz E, Catty P, Shokes JE, Scott RA (2006) Structural basis for metal binding specificity: the N-terminal cadmium binding domain of the P1-type ATPase CadA. J Mol Biol 356:638–650. doi:10.1016/j.jmb.2005.11.055
Barabasz A, Wilkowska A, Tracz K, Ruszczynska A, Bulska E, Mills RF, Williams LE, Antosiewicz DM (2013) Expression of HvHMA2 in tobacco modifies Zn-Fe-Cd homeostasis. J Plant Physiol 170:1176–1186. doi:10.1016/j.jplph.2013.03.018
Bull PC, Cox DW (1994) Wilson disease and Menkes disease: new handles on heavy-metal transport. Trends Genet 10:246–252
Clemens S, Aarts MG, Thomine S, Verbruggen N (2013) Plant science: the key to preventing slow cadmium poisoning. Trends Plant Sci 18:92–99. doi:10.1016/j.tplants.2012.08.003
Courbot M, Willems G, Motte P, Arvidsson S, Roosens N, Saumitou-Laprade P, Verbruggen N (2007) A major quantitative trait locus for cadmium tolerance in Arabidopsis halleri colocalizes with HMA4, a gene encoding a heavy metal ATPase. Plant Physiol 144:1052–1065. doi:10.1104/pp.106.095133
Eren E, Arguello JM (2004) Arabidopsis HMA2, a divalent heavy metal-transporting PIB-type ATPase, is involved in cytoplasmic Zn2+ homeostasis. Plant Physiol 136:3712–3723. doi:10.1104/pp.104.046292
Eren E, Kennedy DC, Maroney MJ, Argüello JM (2006) A novel regulatory metal binding domain is present in the C terminus of Arabidopsis Zn2+-ATPase HMA2. J Biol Chem 281:33881–33891
Eren E, Gonzalez-Guerrero M, Kaufman BM, Arguello JM (2007) Novel Zn2+ coordination by the regulatory N-terminus metal binding domain of Arabidopsis thaliana Zn2+-ATPase HMA2. Biochemistry 46:7754–7764. doi:10.1021/bi7001345
Gravot A, Lieutaud A, Verret F, Auroy P, Vavasseur A, Richaud P (2004) AtHMA3, a plant P1B-ATPase, functions as a Cd/Pb transporter in yeast. FEBS Lett 561:22–28. doi:10.1016/s0014-5793(04)00072-9
Hussain D, Haydon MJ, Wang Y, Wong E, Sherson SM, Young J, Camakaris J, Harper JF, Cobbett CS (2004) P-type ATPase heavy metal transporters with roles in essential zinc homeostasis in Arabidopsis. Plant Cell 16:1327–1339. doi:10.1105/tpc.020487
Kaplan JH (2002) Biochemistry of Na, K-ATPase. Annu Rev Biochem 71:511–535. doi:10.1146/annurev.biochem.71.102201.141218
LeShane ES, Shinde U, Walker JM, Barry AN, Blackburn NJ, Ralle M, Lutsenko S (2010) Interactions between copper-binding sites determine the redox status and conformation of the regulatory N-terminal domain of ATP7B. J Biol Chem 285:6327–6336. doi:10.1074/jbc.M109.074633
Liu H, Zhang Y, Chai T, Tan J, Wang J, Feng S, Liu G (2013a) Manganese-mitigation of cadmium toxicity to seedling growth of Phytolacca acinosa Roxb. is controlled by the manganese/cadmium molar ratio under hydroponic conditions. Plant Physiol Biochem 73:144–153. doi:10.1016/j.plaphy.2013.09.010
Liu J, Zhang H, Zhang Y, Chai T (2013b) Silicon attenuates cadmium toxicity in Solanum nigrum L. by reducing cadmium uptake and oxidative stress. Plant Physiol Biochem 68:1–7. doi:10.1016/j.plaphy.2013.03.018
Lutsenko S, Barnes NL, Bartee MY, Dmitriev OY (2007) Function and regulation of human copper-transporting ATPases. Physiol Rev 87:1011–1046. doi:10.1152/physrev.00004.2006
Mills RF, Krijger GC, Baccarini PJ, Hall JL, Williams LE (2003) Functional expression of AtHMA4, a P1B-type ATPase of the Zn/Co/Cd/Pb subclass. Plant J 35:164–176
Mills RF, Francini A, Ferreira da Rocha PS, Baccarini PJ, Aylett M, Krijger GC, Williams LE (2005) The plant P1B-type ATPase AtHMA4 transports Zn and Cd and plays a role in detoxification of transition metals supplied at elevated levels. FEBS Lett 579:783–791. doi:10.1016/j.febslet.2004.12.040
Mills RF, Valdes B, Duke M, Peaston KA, Lahner B, Salt DE, Williams LE (2010) Functional significance of AtHMA4 C-terminal domain in planta. PLoS One 5:e13388. doi:10.1371/journal.pone.0013388
Mills RF, Peaston KA, Runions J, Williams LE (2012) HvHMA2, a P1B-ATPase from barley, is highly conserved among cereals and functions in Zn and Cd transport. PLoS One 7:e42640. doi:10.1371/journal.pone.0042640
Moller JV, Juul B, le Maire M (1996) Structural organization, ion transport, and energy transduction of P-type ATPases. Biochim Biophys Acta 1286:1–51
Nocito FF, Lancilli C, Dendena B, Lucchini G, Sacchi GA (2011) Cadmium retention in rice roots is influenced by cadmium availability, chelation and translocation. Plant Cell Environ 34:994–1008. doi:10.1111/j.1365-3040.2011.02299.x
Olesen C, Picard M, Winther AM, Gyrup C, Morth JP, Oxvig C, Moller JV, Nissen P (2007) The structural basis of calcium transport by the calcium pump. Nature 450:1036–1042. doi:10.1038/nature06418
Papoyan A, Kochian LV (2004) Identification of Thlaspi caerulescens genes that may be involved in heavy metal hyperaccumulation and tolerance. Characterization of a novel heavy metal transporting ATPase. Plant Physiol 136:3814–3823. doi:10.1104/pp.104.044503
Raimunda D, Gonzalez-Guerrero M, Leeber BW 3rd, Arguello JM (2011) The transport mechanism of bacterial Cu+-ATPases: distinct efflux rates adapted to different function. Biometals 24:467–475. doi:10.1007/s10534-010-9404-3
Rensing C, Ghosh M, Rosen BP (1999) Families of soft-metal-ion-transporting ATPases. J Bacteriol 181:5891–5897
Rosenzweig AC, Huffman DL, Hou MY, Wernimont AK, Pufahl RA, O’Halloran TV (1999) Crystal structure of the Atx1 metallochaperone protein at 1.02 A resolution. Structure 7:605–617
Satoh-Nagasawa N, Mori M, Nakazawa N, Kawamoto T, Nagato Y, Sakurai K, Takahashi H, Watanabe A, Akagi H (2012) Mutations in rice (Oryza sativa) heavy metal ATPase 2 (OsHMA2) restrict the translocation of zinc and cadmium. Plant Cell Physiol 53:213–224. doi:10.1093/pcp/pcr166
Smith AT, Smith KP, Rosenzweig AC (2014) Diversity of the metal-transporting P1B-type ATPases. J Biol Inorg Chem 19:947–960. doi:10.1007/s00775-014-1129-2
Takahashi R, Bashir K, Ishimaru Y, Nishizawa NK, Nakanishi H (2012) The role of heavy-metal ATPases, HMAs, in zinc and cadmium transport in rice. Plant Signal Behav 7:1605–1607. doi:10.4161/psb.22454
Tan J, Wang J, Chai T, Zhang Y, Feng S, Li Y, Zhao H, Liu H, Chai X (2013) Functional analyses of TaHMA2, a P1B-type ATPase in wheat. Plant Biotechnol J 11:420–431. doi:10.1111/pbi.12027
Ueno D, Yamaji N, Kono I, Huang CF, Ando T, Yano M, Ma JF (2010) Gene limiting cadmium accumulation in rice. Proc Natl Acad Sci USA 107:16500–16505. doi:10.1073/pnas.1005396107
Verbruggen N, Juraniec M, Baliardini C, Meyer CL (2013) Tolerance to cadmium in plants: the special case of hyperaccumulators. Biometals 26:633–638. doi:10.1007/s10534-013-9659-6
Verret F, Gravot A, Auroy P, Preveral S, Forestier C, Vavasseur A, Richaud P (2005) Heavy metal transport by AtHMA4 involves the N-terminal degenerated metal binding domain and the C-terminal His11 stretch. FEBS Lett 579:1515–1522
Vida TA, Emr SD (1995) A new vital stain for visualizing vacuolar membrane dynamics and endocytosis in yeast. J Cell Biol 128:779–792
Wang K, Sitsel O, Meloni G, Autzen HE, Andersson M, Klymchuk T, Nielsen AM, Rees DC, Nissen P, Gourdon P (2014) Structure and mechanism of Zn2+-transporting P-type ATPases. Nature 514:518–522. doi:10.1038/nature13618
Williams LE, Mills RF (2005) P1B-ATPases–an ancient family of transition metal pumps with diverse functions in plants. Trends Plant Sci 10:491–502. doi:10.1016/j.tplants.2005.08.008
Wong CK, Jarvis RS, Sherson SM, Cobbett CS (2009) Functional analysis of the heavy metal binding domains of the Zn/Cd-transporting ATPase, HMA2, in Arabidopsis thaliana. New Phytol 181:79–88. doi:10.1111/j.1469-8137.2008.02637.x
Yamaji N, Xia J, Mitani-Ueno N, Yokosho K, Feng Ma J (2013) Preferential delivery of zinc to developing tissues in rice is mediated by P-type heavy metal ATPase OsHMA2. Plant Physiol 162:927–939. doi:10.1104/pp.113.216564
Zhao FJ, Moore KL, Lombi E, Zhu YG (2014) Imaging element distribution and speciation in plant cells. Trends Plant Sci 19:183–192. doi:10.1016/j.tplants.2013.12.001
Zimmermann M, Clarke O, Gulbis JM, Keizer DW, Jarvis RS, Cobbett CS, Hinds MG, Xiao Z, Wedd AG (2009) Metal binding affinities of Arabidopsis zinc and copper transporters: selectivities match the relative, but not the absolute, affinities of their amino-terminal domains. Biochemistry 48:11640–11654. doi:10.1021/bi901573b
Acknowledgments
We thank Prof. Sophie Filleur (Université Paris 7-Denis Diderot, UFR Sciences du Vivant) for providing us with the plasmids pUG35 and pUG36 and Euroscarf (Frankfurt, Germany) for providing the yeast strains ycf1, zrc1, cot1, and end3. This work was supported by the National Natural Science Foundation of China (Grant no: C31370281).
Conflict of interest
There are no conflicts of interest.
Author information
Authors and Affiliations
Corresponding author
Additional information
Communicated by L. Tripathi.
Rights and permissions
About this article
Cite this article
Xiang, S., Feng, S., Zhang, Y. et al. The N-terminal degenerated metal-binding domain is involved in the heavy metal transport activity of TaHMA2. Plant Cell Rep 34, 1615–1628 (2015). https://doi.org/10.1007/s00299-015-1813-x
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00299-015-1813-x