Abstract
In response to heavy metal stress, plants and certain fungi, such as the fission yeast Schizosaccharomyces pombe, synthesize small metal-binding peptides known as phytochelatins. We have identified a cadmium sensitive S. pombe mutant deficient in the accumulation of a sulfide-containing phytochelatin-cadmium complex, and have isolated the gene, designated hmt1, that complements this mutant. The deduced protein sequence of the hmt1 gene product shares sequence identity with the family of ABC (ATP-binding cassette)-type transport proteins which includes the mammalian P-glycoproteins and CFTR, suggesting that the encoded product is an integral membrane protein. Analysis of fractionated fission yeast cell components indicates that the HMT1 polypeptide is associated with the vacuolar membrane. Additionally, fission yeast strains harboring an hmt1-expressing multicopy plasmid exhibit enhanced metal tolerance along with a higher intracellular level of cadmium, implying a relationship between HMT1 mediated transport and compartmentalization of heavy metals. This suggests that tissue-specific overproduction of a functional hmt1 product in transgenic plants might be a means to alter the tissue localization of these elements, such as for sequestering heavy metals away from consumable parts of crop plants.
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- Bowman E. J., Bowman B. J. Purification of vacuolar membranes, mitochondria, and plasma membranes from Neurospora crassa and modes of discriminating among the different H+-ATPases. Methods Enzymol. 1988;157:562–573. doi: 10.1016/0076-6879(88)57104-5. [DOI] [PubMed] [Google Scholar]
- Chen C. M., Misra T. K., Silver S., Rosen B. P. Nucleotide sequence of the structural genes for an anion pump. The plasmid-encoded arsenical resistance operon. J Biol Chem. 1986 Nov 15;261(32):15030–15038. [PubMed] [Google Scholar]
- Cowman A. F., Karcz S., Galatis D., Culvenor J. G. A P-glycoprotein homologue of Plasmodium falciparum is localized on the digestive vacuole. J Cell Biol. 1991 Jun;113(5):1033–1042. doi: 10.1083/jcb.113.5.1033. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Davis R. H. Compartmental and regulatory mechanisms in the arginine pathways of Neurospora crassa and Saccharomyces cerevisiae. Microbiol Rev. 1986 Sep;50(3):280–313. doi: 10.1128/mr.50.3.280-313.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Eisenberg D., Schwarz E., Komaromy M., Wall R. Analysis of membrane and surface protein sequences with the hydrophobic moment plot. J Mol Biol. 1984 Oct 15;179(1):125–142. doi: 10.1016/0022-2836(84)90309-7. [DOI] [PubMed] [Google Scholar]
- Germann U. A., Gottesman M. M., Pastan I. Expression of a multidrug resistance-adenosine deaminase fusion gene. J Biol Chem. 1989 May 5;264(13):7418–7424. [PubMed] [Google Scholar]
- Grill E., Löffler S., Winnacker E. L., Zenk M. H. Phytochelatins, the heavy-metal-binding peptides of plants, are synthesized from glutathione by a specific gamma-glutamylcysteine dipeptidyl transpeptidase (phytochelatin synthase). Proc Natl Acad Sci U S A. 1989 Sep;86(18):6838–6842. doi: 10.1073/pnas.86.18.6838. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hamer D. H. Metallothionein. Annu Rev Biochem. 1986;55:913–951. doi: 10.1146/annurev.bi.55.070186.004405. [DOI] [PubMed] [Google Scholar]
- Hayashi Y., Nakagawa C. W., Mutoh N., Isobe M., Goto T. Two pathways in the biosynthesis of cadystins (gamma EC)nG in the cell-free system of the fission yeast. Biochem Cell Biol. 1991 Feb-Mar;69(2-3):115–121. doi: 10.1139/o91-018. [DOI] [PubMed] [Google Scholar]
- Jones E. W. Tackling the protease problem in Saccharomyces cerevisiae. Methods Enzymol. 1991;194:428–453. doi: 10.1016/0076-6879(91)94034-a. [DOI] [PubMed] [Google Scholar]
- Juranka P. F., Zastawny R. L., Ling V. P-glycoprotein: multidrug-resistance and a superfamily of membrane-associated transport proteins. FASEB J. 1989 Dec;3(14):2583–2592. doi: 10.1096/fasebj.3.14.2574119. [DOI] [PubMed] [Google Scholar]
- Kamijo K., Taketani S., Yokota S., Osumi T., Hashimoto T. The 70-kDa peroxisomal membrane protein is a member of the Mdr (P-glycoprotein)-related ATP-binding protein superfamily. J Biol Chem. 1990 Mar 15;265(8):4534–4540. [PubMed] [Google Scholar]
- Klein P., Kanehisa M., DeLisi C. The detection and classification of membrane-spanning proteins. Biochim Biophys Acta. 1985 May 28;815(3):468–476. doi: 10.1016/0005-2736(85)90375-x. [DOI] [PubMed] [Google Scholar]
- Klionsky D. J., Emr S. D. A new class of lysosomal/vacuolar protein sorting signals. J Biol Chem. 1990 Apr 5;265(10):5349–5352. [PubMed] [Google Scholar]
- Klionsky D. J., Herman P. K., Emr S. D. The fungal vacuole: composition, function, and biogenesis. Microbiol Rev. 1990 Sep;54(3):266–292. doi: 10.1128/mr.54.3.266-292.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kozak M. Point mutations define a sequence flanking the AUG initiator codon that modulates translation by eukaryotic ribosomes. Cell. 1986 Jan 31;44(2):283–292. doi: 10.1016/0092-8674(86)90762-2. [DOI] [PubMed] [Google Scholar]
- Kuchler K., Sterne R. E., Thorner J. Saccharomyces cerevisiae STE6 gene product: a novel pathway for protein export in eukaryotic cells. EMBO J. 1989 Dec 20;8(13):3973–3984. doi: 10.1002/j.1460-2075.1989.tb08580.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Logemann J., Schell J., Willmitzer L. Improved method for the isolation of RNA from plant tissues. Anal Biochem. 1987 May 15;163(1):16–20. doi: 10.1016/0003-2697(87)90086-8. [DOI] [PubMed] [Google Scholar]
- McGrath J. P., Varshavsky A. The yeast STE6 gene encodes a homologue of the mammalian multidrug resistance P-glycoprotein. Nature. 1989 Aug 3;340(6232):400–404. doi: 10.1038/340400a0. [DOI] [PubMed] [Google Scholar]
- Murasugi A., Wada C., Hayashi Y. Occurrence of acid-labile sulfide in cadmium-binding peptide 1 from fission yeast. J Biochem. 1983 Feb;93(2):661–664. doi: 10.1093/oxfordjournals.jbchem.a134222. [DOI] [PubMed] [Google Scholar]
- Murasugi A., Wada C., Hayashi Y. Purification and unique properties in UV and CD spectra of Cd-binding peptide 1 from Schizosaccharomyces pombe. Biochem Biophys Res Commun. 1981 Dec 15;103(3):1021–1028. doi: 10.1016/0006-291x(81)90911-6. [DOI] [PubMed] [Google Scholar]
- Mutoh N., Hayashi Y. Isolation of mutants of Schizosaccharomyces pombe unable to synthesize cadystin, small cadmium-binding peptides. Biochem Biophys Res Commun. 1988 Feb 29;151(1):32–39. doi: 10.1016/0006-291x(88)90555-4. [DOI] [PubMed] [Google Scholar]
- Nriagu J. O., Pacyna J. M. Quantitative assessment of worldwide contamination of air, water and soils by trace metals. Nature. 1988 May 12;333(6169):134–139. doi: 10.1038/333134a0. [DOI] [PubMed] [Google Scholar]
- Pickett C. B., Lu A. Y. Glutathione S-transferases: gene structure, regulation, and biological function. Annu Rev Biochem. 1989;58:743–764. doi: 10.1146/annurev.bi.58.070189.003523. [DOI] [PubMed] [Google Scholar]
- Rauser W. E. Phytochelatins. Annu Rev Biochem. 1990;59:61–86. doi: 10.1146/annurev.bi.59.070190.000425. [DOI] [PubMed] [Google Scholar]
- Reese R. N., White C. A., Winge D. R. Cadmium-Sulfide Crystallites in Cd-(gammaEC)(n)G Peptide Complexes from Tomato. Plant Physiol. 1992 Jan;98(1):225–229. doi: 10.1104/pp.98.1.225. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Reese R. N., Winge D. R. Sulfide stabilization of the cadmium-gamma-glutamyl peptide complex of Schizosaccharomyces pombe. J Biol Chem. 1988 Sep 15;263(26):12832–12835. [PubMed] [Google Scholar]
- Riordan J. R., Rommens J. M., Kerem B., Alon N., Rozmahel R., Grzelczak Z., Zielenski J., Lok S., Plavsic N., Chou J. L. Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA. Science. 1989 Sep 8;245(4922):1066–1073. doi: 10.1126/science.2475911. [DOI] [PubMed] [Google Scholar]
- Roberts C. J., Raymond C. K., Yamashiro C. T., Stevens T. H. Methods for studying the yeast vacuole. Methods Enzymol. 1991;194:644–661. doi: 10.1016/0076-6879(91)94047-g. [DOI] [PubMed] [Google Scholar]
- Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Smith C. L., Matsumoto T., Niwa O., Klco S., Fan J. B., Yanagida M., Cantor C. R. An electrophoretic karyotype for Schizosaccharomyces pombe by pulsed field gel electrophoresis. Nucleic Acids Res. 1987 Jun 11;15(11):4481–4489. doi: 10.1093/nar/15.11.4481. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Speiser D. M., Abrahamson S. L., Banuelos G., Ow D. W. Brassica juncea Produces a Phytochelatin-Cadmium-Sulfide Complex. Plant Physiol. 1992 Jul;99(3):817–821. doi: 10.1104/pp.99.3.817. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Steffens J. C., Hunt D. F., Williams B. G. Accumulation of non-protein metal-binding polypeptides (gamma-glutamyl-cysteinyl)n-glycine in selected cadmium-resistant tomato cells. J Biol Chem. 1986 Oct 25;261(30):13879–13882. [PubMed] [Google Scholar]
- Thumann J., Grill E., Winnacker E. L., Zenk M. H. Reactivation of metal-requiring apoenzymes by phytochelatin-metal complexes. FEBS Lett. 1991 Jun 17;284(1):66–69. doi: 10.1016/0014-5793(91)80763-s. [DOI] [PubMed] [Google Scholar]
- Trowsdale J., Hanson I., Mockridge I., Beck S., Townsend A., Kelly A. Sequences encoded in the class II region of the MHC related to the 'ABC' superfamily of transporters. Nature. 1990 Dec 20;348(6303):741–744. doi: 10.1038/348741a0. [DOI] [PubMed] [Google Scholar]
- Vögeli-Lange R., Wagner G. J. Subcellular localization of cadmium and cadmium-binding peptides in tobacco leaves : implication of a transport function for cadmium-binding peptides. Plant Physiol. 1990 Apr;92(4):1086–1093. doi: 10.1104/pp.92.4.1086. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Walker J. E., Saraste M., Runswick M. J., Gay N. J. Distantly related sequences in the alpha- and beta-subunits of ATP synthase, myosin, kinases and other ATP-requiring enzymes and a common nucleotide binding fold. EMBO J. 1982;1(8):945–951. doi: 10.1002/j.1460-2075.1982.tb01276.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weisman L. S., Bacallao R., Wickner W. Multiple methods of visualizing the yeast vacuole permit evaluation of its morphology and inheritance during the cell cycle. J Cell Biol. 1987 Oct;105(4):1539–1547. doi: 10.1083/jcb.105.4.1539. [DOI] [PMC free article] [PubMed] [Google Scholar]
- West I. C. What determines the substrate specificity of the multi-drug-resistance pump? Trends Biochem Sci. 1990 Feb;15(2):42–46. doi: 10.1016/0968-0004(90)90171-7. [DOI] [PubMed] [Google Scholar]
- von Heijne G. Signal sequences. The limits of variation. J Mol Biol. 1985 Jul 5;184(1):99–105. doi: 10.1016/0022-2836(85)90046-4. [DOI] [PubMed] [Google Scholar]