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2012 | 61 | 4 | 587-596
Article title

Telomery i telomeraza w komórkach roślinnych

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EN
Telomeres and telomerase in plant cells.
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PL EN
Abstracts
PL
W budowie i funkcji kompleksu telomerowego występującego w komórkach roślin i zwierząt, obserwuje się szereg podobieństw. W jego skład wchodzą telomerowe DNA, białka telomerowe oraz enzym telomeraza. Telomery to struktury zbudowane z kompleksu białek i tandemowo powtórzonych sekwencji DNA, zlokalizowane na końcach chromosomów eukariotycznych. Pełnią szereg istotnych funkcji w komórkach organizmów żywych. Najważniejszą ich rolą jest ochrona genomu przed potencjalną niestabilnością. O ile sekwencja telomerowego DNA jest stosunkowo silnie zakonserwowana nawet u organizmów odległych ewolucyjnie, o tyle kompleks białek telomerowych charakteryzuje się dużym zróżnicowaniem. Z telomerami współdziała telomeraza - enzym o aktywności odwrotnej transkryptazy, który na matrycy własnego RNA dobudowuje na końcach chromosomów sekwencje telomerowe. Istniej wiele przesłanek wskazujących na udział kompleksu telomerowego w starzeniu się komórek roślinnych. Jednak dotychczas nie udało się w pełni zweryfikować tej hipotezy. Wiedza z zakresu budowy i funkcji telomerów i telomerazy w komórkach roślinnych ciągle pozostaje daleko w tyle za tą uzyskaną dla komórek ssaków.
EN
Both in plants and animals cells several similarities in structure and function of telomere complex are observed. Telomere complex consists of DNA, proteins and telomerase. Telomeres are the special structures composed of proteins and tandem repeated DNA sequences, localized at the physical end of eukariotic chromosomes. They carry out many important functions in the cells. The most important of their role is to protect the genome from potential instability. While the telomeric DNA sequence is relatively highly conserved even among evolutionarily distant organisms, telomeric protein complex has a great diversity. Telomerase interacts with telomeres. This enzyme has a reverse transcriptase activity and based on its own RNA template adds telomeric sequences at the ends of chromosomes. There are lots of indications that telomere complex participates in the plant cells aging process. But so far this hypothesis has not been fully verified. Knowledge in the field of structure and function of telomeres and telomerase in plant cells still remains far behind that achieved in mammalian cells.
Keywords
Journal
Year
Volume
61
Issue
4
Pages
587-596
Physical description
Dates
published
2012
Contributors
  • Polska Akademia Nauk Ogród Botaniczny Centrum Zachowania Różnorodności Biologicznej w Powsinie Prawdziwka 2, 02-973 Warszawa, Polska
  • Polska Akademia Nauk Ogród Botaniczny Centrum Zachowania Różnorodności Biologicznej w Powsinie Prawdziwka 2, 02-973 Warszawa, Polska
References
  • Adams S. P., Hartman T. P., Lim K. Y., Chase M. W., Bennett M. D., Leitch I. J., Leitch A. R., 2001. Loss and recovery of Arabidopsis-type telomere repeat sequences 5'-(TTTAGGG)(n)-3' in the evolution of a major radiation of flowering plants. Proc. Biol. Sci. 268, 1541-1546.
  • Aparicio O. M., Billington B. L., Gottschling D. E., 1991. Modifiers of position effect are shared between telomeric and sileni miting type loci in S cerevisiae. Cell 66, 1279-1287.
  • Baumann P., Cech T. R., 2001. Pot1, the putative telomere end-binding protein in fission yeast and humans. Science 292, 1171-1175.
  • Bianchi A., Shore D., 2008. How telomerase reaches its end: mechanism of telomerase regulation by the telomeric complex. Mol. Cell 31, 153-165.
  • Bilaud T., Koering C. E., Binet-Brasselet E., Ancelin K., Pollice A., Gasser S. M., Gilson E., 1996. The telobox, a Myb-related telomeric DNA binding motif found in proteins from yeast, plants and human. Nucl. Acids Res. 24, 1294-1303.
  • Bilaud T., Brun C., Ancelin K., Koering C. E., Laroche T., Gilson E., 1997. Telomeric localization of TRF2, a novel human telobox protein. Nat. Genet. 17, 236-239.
  • Blackburn E. H., Gall J. G., 1978. A tandemly repeated sequence at the termini of the extrachromosomal ribosomal RNA genes in Tetrahymena. J. Mol. Biol. 120, 33-53.
  • Blackburn E. H., 2001. Switching and signaling at the telomere. Cell 106, 661-673.
  • Blackburn E. H., 2005.Telomeres and telomerase: their mechanisms of action and their effects of altering their functions. FEBS Lett. 579, 859-862.
  • Blanc G ., Duncan G ., Agarkova I ., Borodovsky M ., Gurnon J ., Kuo A ., Lindquist E ., Lucas S ., Pangilinan J ., Polle J ., Salamov A ., Terry A ., Yamada T ., Dunigan D. D ., Grigoriev I. V ., Claverie J. M ., Van Etten J. L ., 2010. The Chlorella variabilis NC64A genome reveals adaptation to photosymbiosis, coevolution with viruses, and cryptic sex. Plant Cell 22, 2943-2955.
  • Blasco M. A., 2002. Telomerase beyond telomeres. Nat. Rev. Cancer 2, 627-632.
  • Blasco M., 2007 . The epigenetic regulation of mammalian telomeres . Nat. Rev. Genet. 8, 299-309.
  • Boubriak I., Polischuk V., Grodzinsky A., Osborne D. J., 2007. Telomeres and seed banks. Cytol. Genet. 41, 18-24.
  • Bucholc M., Buchowicz J., 1992. Synthesis of extra chromosomal DNA and telomere-releted sequences in germinating wheat embryos. Seed Sci. Res. 2, 141-146.
  • Bucholc M., Buchowicz J., 1995. An extrachromosomal fragment of telomeric DNA in wheat. Plant Mol. Biol. 27, 435-439.
  • Cesare A. J., Quinney N., Willcox S., Subramanian D., Griffith J. D., 2003. Telomere looping in P. sativum (common garden pea). Plant J. 36, 271-279.
  • Chakhparonian M., Wellinger R. J., 2003. Telomere maintance and DNA replication: how closely are these two connected? Trends Genet. 19, 439-446.
  • Chan S. W. L., Blackburn E. H., 2002. New ways not to make ends meet: telomerase, DNA damage proteins and heterohromatin. Oncogene 21, 553-563.
  • Chen J. L., Blasco M. A., Greider C. W., 2000. Secondary structure of vertebrate telomerase RNA. Cell 100, 503-514.
  • Choi J., Southworth L. K., Sarin K. Y., Venteicher A. S., Ma W., Chang W., Cheung P., Jun S., Artandi M. K., Shah N., Kim s. K., Artandi S. E., 2008. TERT promotes epithelial proliferation through transcriptional control of a Myc- and Wnt related developmental program. PLoS Genet. 4, e10.
  • Chong L., van Steensel B., Broccoli D., Erdjument-Bromage H., Hanish J., Tempst P., de Lange T., 1995. A human telomeric protein. Science 270, 1663-1667.
  • Cohen S. B., Graham M. E., Loverecz G. O., Bache N., Robinson P. J., Reddel R. R., 2007. Protein composition of catalytically active human telomerase from immortal cells. Science 315, 1850-1853.
  • Cifuentes-Rojas C., Kannan K., Tseng L., Shiooen D. E., 2011. Two RNA subunits and POT1a are components of Arabidopsis telomerase. Proc. Natl. Acad. Sci. USA 108, 73-78.
  • Colgin L. M., Reddel R. R., 1999. Telomere maintance mechanisms and cellular immortalization. Curr. Opin. Genet. Develop. 9, 97-103.
  • Counter C. M., Avilion A. A., LeFeuvrel C. E., Stewart N. G., Greider C. W., Harley C. B., Bacchettil S., 1992. Telomere shortening associated with chromosome instability is arrested in immortal cells which express telomerase activity. EMBO J. 11, 1921-1929.
  • de Lange T., Shiue L., Myers R. M., Cox D. R., Naylor S. L., Killery A. M., Varmus H. E., 1990. Structure and variability of human chromosome ends. Mol. Cell. Biol. 10, 518-527.
  • de Lange T., 2005. Shelterin: The protein complex that shapes and safeguards human telomeres. Genes Develop. 19, 2100-2110.
  • Dubrana K., Perrod S., Gasser S. M., 2001. Turning telomeres off and on. Curr. Opin. Cell Biol. 13, 281-289.
  • Fajkus J., Kovarík A., Královics R., Bezděk M., 1995. Organization of telomeric and subtelomeric chromatin in the higher plant Nicotiana tabacum. Mol. Gen. Genet. 247, 633-638.
  • Fajkus J., Sýkrová E., Leitch A. R., 2005. Telomeres in evolution and evolution of telomeres. Chromosome Res. 13, 469-479.
  • Fizgerald M. S., McKnight T. D., Shippen D. E. 1996. Characterization and developmental patterns of telomerase expression in plants. Proc. Natl. Acad. Sci. USA 93, 14422-14427.
  • Fizgerald M. S., Riha K., Gao F., Ren S., McKnight T. D., Shippen D. E., 1999. Disruption of the telomerase catalytic subunit gene from Arabidopsis inactivates telomerase anl leads to sa slow loss of telomeric DNA. Proc. Natl. Acad. Sci. USA 96, 14813-14818.
  • Flanary B. E., Kletetschka G., 2005. Analysis of telomere length and telomerase activity in tree species of various life-spans, and with age in the bristlecone pine Pinus longaeva. Biogerontology 6, 101-111.
  • Fu W., Killen M. W., Culmsee C., Dhar S., Pandita T., Mattson M. P., 2000. The catalytic subunit of telomerase is expressed in developing brain neurons and serves a cell survival-promoting function. J. Mol. Neurosci. 14, 3-15.
  • Fuchs I., Brandes A., Schubert I., 1995. Telomere sequence localization and karyotype evolution in higher plants. Plant Systemat. Evol. 196, 227-241.
  • Gnal M. W., Lapitan N. L. V., Tanksley S. D., 1991. Macrostructure of the tomato telomeres. Plant Cell 3, 87-94.
  • Greider C. W., 1995. Telomerase Biochemistry and Regulation [W:] Telomeres. Blackburn E. H., Greider C. W. (red.). Cold Spring Harbor Labolatory Press, NY, 35-68.
  • Greider C. W., 1996. Telomere length regulation. Ann. Rev. Biochem. 65, 337-365.
  • Greider C. W ., 1999. Telomeres Do D-Loop-T-Loop. Cell 97, 419-422.
  • Greider C. W., Blackburn E. H., 1985. Identification of a specific telomere terminal transferase activity in Tetrahymena extracts. Cell 43, 405-413.
  • Greider C. W., Blackburn E. H., 1987. The telomere terminal transferase of Tetrahymena is a ribonucleoprotein enzyme with two kinds of primer specificity. Cell 51, 887-898.
  • Greider C. W., Blackburn E. H., 1989. A telomeric sequence in the RNA of Tetrahymena telomerase required for telomere repeat synthesis. Nature 337, 331-337.
  • Griffith J. D., Comeau L., Rosenfield S., Stansel R. M., Bianchi A., Moss H., de Lange T., 1999. Mammalian telomeres end in a large duplex loop. Cell 97, 503-514.
  • Haendeler J., Hoffmann J., DiehlJ. F., Vasa M., Spyridopoulos I., Zeiher A.M., Dimmelers., 2004. Antioxidants inhibit nuclear export of telomerase reverse transcriptase and delay replicative senescence of endothelial cells. Circulation Res. 94, 768-775.
  • Heller K., Kilian A., Piatyszek M. A., Kleinhofs A., 1996. Telomerase activity in plant extracts. Mol. Gen. Genet. 252, 342-345.
  • Henderson E. R., Blackburn E. H., 1989. An overhanging 3' terminus is a conserved feature of telomeres. Mol. Cell. Biol. 9, 345-348.
  • Higashiyama T., Maki S., Yamada T., 1995. Molecular organisation of Chlorella vulgaris chromosome I: presence of telomeric repeats that are conserved in higher plants. Mol. Gen. Genet. 246, 29-36.
  • Hirata Y., Suzuki C., Sakai S., 2004. Characterization and gene cloning of telomere-binding protein from tobacco BY-2 cells. Plant Physiol. Biochem. 42, 7-14.
  • Hwang M. G., Chung I. K., Kang B. G., Cho M. H., 2001. Sequence-specific binding property of Arabidopsis thaliana telomeric DNA binding protein 1 (AtTBP1). FEBS Lett. 503, 35-40.
  • Karamysheva Z. N., Surovtseva Y. V., Vespa L., Shakirov E. V., Shippen D. E., 2004. A C-terminal Myb extension domain defines a novel family of double-strand telomeric DNA-binding proteins in Arabidopsis. J. Biol. Chem. 279, 47799-47807.
  • Kelleher C., Teixeira M. T., Forstemann K., Lingner J., 2002. Telomerase: biochemical considerations for enzyme and substrate. Trends Biochem. Sci. 27, 572-579.
  • Kilian A., Stiff C., Kleinhofs A., 1995. Barley telomeres shorten during differentioation but grow in callus culture. Proc. Natl. Acad. Sci. USA 92, 9555-9559.
  • Kilian A., Bowtell D. D., Abud H. E., Hime G.R., Venter D. J., Keese P.K., Duncan E. L., Reddel R. R., Jefferson R. A., 1997.Isolation of a candidate human telomerase catalytic subunit gene which reveals complex splicing patterns in different cell types. Human Mol. Genet. 6, 2011-2019.
  • Kilian A., Heller K., Kleinhofs A., 1998. Developmental patterns of telomerase activity in barley and maize. Plant Mol. Biol. 37, 621-628.
  • Kwon C., Chung I. K., 2004. Interaction of an Arabidopsis RNA-binding Protein with Plant Single-stranded Telomeric DNA Modulates Telomerase Activity. J. Biol. Chem. 279, 12812-12818.
  • Lee J. H., Kim J. H., Kim W. T., KangB. G., Chung I. K., 2000. Characterization and developmental expression of single-stranded telomeric DNA-binding proteins from mung bean (Vigna radiata). Plant Mol. Biol. 42, 547-557.
  • Lee Y. W., Kim W. T., 2011. Roles of NtGTBP1 in telomere stability. Plant Signal. Behav. 6, 523-525.
  • Leonardi J., Box J. A., Bunch J. T., Baumann P., 2008. TER1, the RNA subunit of fission yeast telomerase. Nature Struct. Mol. Biol. 15, 26-33.
  • Li B. B., Lustig A. J., 1996. A novel mechanism for telomere size control in Saccharomyces cerevisiae. Genes Develop. 10, 1310-1326.
  • Lingner J., Hendrick L. L., Cech T. R,. 1994. Telomerase RNAs of different cilitates have a common secondary structure and permuted template. Genes Develop. 8, 1984-1998.
  • Lingner J., Hughes T. R., Shevchenko A., Mann M., Lundblad V, Cech T. R., 1997. Reverse transcriptase motifs in the catalytic subunit of telomerase. Science 276, 561-567.
  • Liu D., o Connor M. S., Qin J., Songyang Z., 2004. Telosome a mammalian telomere-associated complex formed by multiple telomeric proteins. J. Biol. Chem. 279, 51338-51342.
  • Lundblad V., 1998. Telomerase catalysis: a phylogenetically conserved reverse transcriptase. Proc. Natl. Acad. Sci. USA 95, 8415-8416.
  • Maddar H., Ratzkovsky N., Krauskopf A., 2001. Role for Telomere Cap Structure in Meiosis. Mol. Biol. Cell 12, 3191-3203.
  • Maida Y., Yasukawa M., Furuuchi M., Lassmann T., Possemato R., Okamoto N., Kasim V., Hayashizaki Y., HahnW. C., MasutomiK., 2009. An RNA-dependent RNA polymerase formed by TERT and the RMRP RNA. Nature 461, 230-235.
  • Marian C. O., Bordoli S. J., Goltz M., Santarella R. A., Jackson L. P., Danilevskaya O., Beckstette M., Meeley R., Bass H. W., 2003. The maize Single myb histone 1 gene, Smh1, belongs to a novel gene family and encodes a protein that binds telomere DNA repeats in vitro. Plant Physiol. 133, 1336-1350.
  • Martinez P., Thanasoula M., CarlosA. R., Gómez-López G., Tejera A. M., Schoeftner S., Dominguez O., PisanoD. G., Tarsounas M., Blasco M. A., 2010. Mammalian Rap1 controls telomere function and gene expression through binding to telomeric and extratelomeric sites. Nature Cell Biol. 12, 768-780.
  • McClintock B., 1941. The stability of broken ends of chromosomes in Zea mays. Genetics 26, 234-282.
  • Mechelli R., Anselmi C., Cacchione S., De Santis P., Saviano M., 2004. Organisation of telomeric nucleosomes: atomic force microscopy imaging and theoretical modelling. FEBS Lett. 566, 131-135.
  • Meyerson M., Counter C. M., Eaton E. N., Ellisen L. W., Steiner P., Caddle S. D., Ziaugra L., Beijersbergen R. L., Davidoff M. J., Liu Q., Bacchetti S., Haber D. A., Weinberg R. A., 1997. hEST2, the putative human telomerase catalytic subunit gene, is up-regulated in tumor cells and during immortalization. Cell 90, 785-795.
  • Muller H. J., 1938. The remaking of chromosome. Collecting Net 8, 182-198.
  • Oguchi K., Liu H., Tamura K., Takahasi H., 1999. Molecular cloning and characterization of AtTERT, a telomerase reverse transcriptase homolog in Arabidopsis thaliana. FEBS Lett. 457, 465-469.
  • Palm W., de Lange T ., 2008. How shelterin protects Mammalian telomeres . Ann. Rev. Genet. 42, 301-334.
  • Petracek M. E., Lefebvre P. A., Silflow C. D., Berman J., 1990. Chlamydomonas telomere sequences are A+T-rich but contain three consecutive G-Cbase pairs. Proc. Natl. Acad. Sci. USA 87, 8222-8226.
  • Pisano S., Pascucci E. C., Cacchione S., De Santis P., Saviano M., 2006. AFM imaging and theoretical modelling studies of sequence-dependent nucleosome positioning. Biophys. Chem. 124, 81-89.
  • Pisano S., Marchioni E., Galati A., Mechelli R., Saviano M., Cacchione S., 2007. Telomeric nucleosomes are intrnscally mobile . J. Mol. Biol. 369, 1153-1162.
  • Riha K., Fajkus J., Siroky J., Vyskot B., 1998. Developmental control of telomere lengths and telomerase activity in plants. Plant Cell 10, 1691-1698.
  • Riha K., McKnight t. D., Fajkus j., Vyskot B., Shippend. E., 2000. Analysis of the G-overhang structures on plant telomeres: evidence for two distinct telomere architectures. Plant J. 23, 633-641.
  • Richards E. J., Ausbel F. M., 1988. Isolation of a higher eucariotic telomere from Arabidopsis thaliana. Cell 53, 345-348.
  • Richards E. J., Vongs A., Walsh M., Yang J., Chao S., 1993. Substructure of telomere repeat arrays. [W:] The chromosome. Heslop-Harrison J. S., Flavell R. B. (red.). Bios Scientific Publisher, Oxford, 103-114.
  • Schrumpfova P., Kuchar M., Mikova G., Skrisovska L, Kubicarova T, Fajkus J., 2004. Characterization of two Arabidopsis thaliana myb-like proteins showing affinity to telomeric DNA sequence. Genome 47, 316-324.
  • Schrumpfova P., Kuchar M., Palecek J., Fajkus J ., 2008. Mapping of interaction domains of putative telomere-binding proteins AtTRB1 and AtPOT1b from Arabidopsis thaliana. FEBS Lett. 582, 1400-1406.
  • Schwarzacher T., Heslop-Harrison J. S., 1991. In situ hybridization to plant telomeres using synthetic oligomers. Genome 34, 317-323.
  • Shakirov E. V., Surovtseva Y. V., Osbun N, Shippen D. E., 2005. The Arabidopsis Pot1 and Pot2 proteins function in telomere length homeostasis and chromosome end protection. Mol. Cell. Biol. 25, 7725-7733.
  • Shakirov E. V., Perroud P. F., Nelson A. D., Cannell M. E., Quatrano R. S., Shippen D. E., 2010 . Protection of Telomeres 1 is required for telomere integrity in the moss Physcomitrella patens. Plant Cell 22, 1838-1848.
  • Stewart S. A., Ben-Porath I., Carey V. J., O'Connor B. F., Hahn W. C., Weinberg R. A., 2003 . Erosion of the telomeric single-strand overhang at replicative senescence. Nat. Genet. 33, 492-496.
  • Surovtseva Y. V., Shakirov E. V., Vespa L., Osbun N., Song X., Shippen D. E., 2007. Arabidopsis POT1 associates with the telomerase RNP and is required for telomere maintenance. EMBO J. 26, 3653-3661.
  • Syrkowa E., Fajkus J., 2009. Structure-function relationships in telomerase genes. Biol. Cell 101, 375-392.
  • Szostak J. W., Blackburn E. H., 1982. Cloning yeast telomeres on linear plasmids. Cell 29, 245-255.
  • Tani A., Murata M., 2005. Alternative splicing of Pot1 (Protection of telomere)-like genes in Arabidopsis thaliana. Genes Genet. Syst. 80, 41-48.
  • Tremousaygue D., Manevski A., Bardet C., Lescure N., Lescure B.,1999. Plant interstitial telomere motifs participate in the control of gene expression in root meristems. Plant J. 20, 552-561.
  • Tsujimoto H., Yamada T., Sasakuma, T., 1997. Molecular structure of a wheat chromosome end healed after gametocidal gene-induced breakage. Proc. Natl. Acad. Sci. USA 94, 3140-3144.
  • Tsujimoto H., Usami N., Hasegawa K., Yamada T., Sasakuma T., 1999. De novo synthesis of telomere sequences at the healed breakpoints of wheat deletionchromosomes. Mol. Gen. Genet. 262, 851-856.
  • Vaquero-Sedas M., Gamez-Arjona F. M., Vega-Palas M. A., 2010. Arabidopsis thaliana telomeres exhibit euchromatic features. Nucl. Acids Res. 39, 2007-2017.
  • Wang F., Podell E. R., Zaug A. J., Yang Y., Baciu P., Cech T. R., Lei M., 2007. POT1-TPP1 telomere complex is a telomerase processivity factor. Nature 445, 506-510.
  • Watson J. M., Riha K., 2010. Comperative biology of telomeres: where plants stand. FEBS Lett. 584, 3752-3759.
  • Webb C. J., Zakian V. A., 2008. Identification and characterisation of the Schizosaccharomyces pombe TER1 telomerase RNA. Nat. Struct. Mol. Biol. 15, 34-42.
  • Xin H., Liu D., Wan M., Safari A., Kim H., Sun W., O'Connor M. S., Songyang Z., 2007. TPP1 is a homologue of ciliate TEBP-beta and interacts with POT1 to recruit telomerase. Nature 455, 559-562.
  • Yang S. W., Kim D. H., Lee J. J., Chun Y. J., Lee J. H., Kim Y. J., Chung I. K., Kim W. T., 2003. Expression of the telomeric repeat binding factor gene NgTRF1 is closely coordinated with the cell division program in tobacco BY-2 suspension culture cells. J. Biol. Chem. 278, 21395-21407.
  • Yu E. Y., Kim S. E., Kim J. H., Ko J. H., Cho M. H., Chung I. K., 2000. Sequence-specific DNA recognition by the Myb-like domain of plant telomeric protein RTBP1. J. Biol. Chem. 275, 24208-24214.
  • Yu G. L., Bradley J. D., Attardi L. D., Blackburn E. H., 1990. In vivo alteration of telomere sequences and senescence caused by mutated Tetrahymena telomerase RNAs. Nature 344, 126-132.
  • Zentgraf U., Hinderhofer K., Kolb D., 2000. Specific association of a small protein with the telomeric DNA-protein complex during the onset of leaf senescence in Arabidopsis thaliana. Plant Mol. Biol. 42, 429-438.
  • Zhao Y., Sfeir A .J, Zou Y., Buseman C.M., Chow T.T., Shay J.W., Wright W.E., 2009. Telomere extension occurs at most chromosome ends and is uncoupled from fill-in in human cancer cells. Cell 138, 463-475.
  • Zhu H., Fu W., Mattson M. P., 2000. The catalytic subunit of telomerase protects neurons against amyloid beta-peptide-induced apoptosis. J. Neurochem. 75, 117-124.
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bwmeta1.element.bwnjournal-article-ksv61p587kz
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