Publikationsserver der Universitätsbibliothek Marburg

Titel:Histon-Acetylierung ist essentiell für die Spermatidenentwicklung von histonbasiertem zu protaminbasiertem Chromatin in Drosophila melanogaster
Autor:Awe, Stephan
Weitere Beteiligte: Renkawitz-Pohl, Renate (Prof. Dr.)
Veröffentlicht:2010
URI:https://archiv.ub.uni-marburg.de/diss/z2010/0091
URN: urn:nbn:de:hebis:04-z2010-00918
DOI: https://doi.org/10.17192/z2010.0091
DDC: Biowissenschaften, Biologie
Titel (trans.):Histone acetylation is essential for spermatid development from a histone-based to a protamine-based chromatin in Drosophila melanogaster
Publikationsdatum:2010-03-09
Lizenz:https://rightsstatements.org/vocab/InC-NC/1.0/

Dokument

Schlagwörter:
spermatogenesis, inhibitor-treatment, Spermatogenese, histone-protamine switch, Histon-Protamin Wechsel, chromatin, Zysten-Zellkultur, Chromatin, Trichostatine, Organkultur, Anacardsäure, organ culture

Zusammenfassung:
In der Spermatogenese vieler Tiere wird die histonbasierte Chromatinstruktur des paternalen Genoms aufgelöst, um durch eine Struktur auf Grundlage von Protaminen ersetzt zu werden. Der korrekte Ablauf dieses Histon-Protamin Wechsels (H-P Wechsel) ist essentiell für die männliche Fertilität. Der H-P Wechsel findet auch in den Spermatiden von Drosophila statt. Bisher war nicht bekannt, in welchem Zeitrahmen der H-P Wechsel innerhalb der postmeiotischen Entwicklung abläuft. In dieser Arbeit wurde dazu ein Kultursystem für isolierte, pupale Testes und einzelne Spermatidenverbände (Zysten) etabliert. Mit Hilfe von Histon-RFP- und Protamin-eGFP-exprimierenden Fliegen wurde in fluoreszenzmikroskopischen Zeitraffer-Aufnahmen die Entwicklung isolierter Zysten durch die Meiose und den H-P Wechsel in vivo beobachtet. Der H-P Wechsel findet 50-60 h nach der Meiose statt und ist innerhalb von 5 h abgeschlossen. Sowohl bei Vertebraten als auch bei Drosophila geht dem H-P Wechsel eine Hyperacetylierung des Histons H4 voraus und er wird begleitet von DNA-Strangbrüchen. Im Säuger-Modell werden die Histon H4-Hyperacetylierung als mögliches Startsignal des H-P Wechsels und Topoisomerasen als verantwortlich für die DNA-Strangbrüche angesehen. In der vorliegenden Arbeit wurde überprüft, ob diese Annahmen auch für die Spermatogenese von Drosophila zutreffen. Durch Inhibitorbehandlung von pupalen Testes und isolierten Zysten in Kultur wurde die Aktivität von Histon-Acetyltransferasen (HATs) und Histon-Deacetylasen (HDACs) manipuliert. Histon-Acetylierung zeigte sich als essentiell für die korrekte Entwicklung postmeiotischer Keimzellen. Allerdings induzierte eine HDAC-Inhibitor-vermittelte, vorzeitige Histon-Acetylierung kein vorzeitiges Auftreten von Merkmalen des H-P Wechsels. Histon H4-Acetylierung ist somit nicht hinreichend als das alleinige Startsignal des H-P Wechsels in Drosophila. Die Inhibition von Topoisomerasen des Typs I und II zeigte keinen Effekt auf den H-P Wechsel und die Entwicklung postmeiotischer Spermatiden. Ein Zusammenhang zwischen Topoisomeraseaktivität und DNA-Brüchen in Spermatiden von Drosophila scheint daher nicht zu bestehen. Bisher war nicht bekannt, welche HATs an der Histon H4-Acetylierung des H-P Wechsels beteiligt sind. Nach manueller, stadienspezifischer Isolation von Einzelzysten konnte hier mittels RT-PCR die Expression einiger HATs während des H-P Wechsels geprüft werden und ein Kandidat identifiziert werden. Die in dieser Arbeit gewonnenen Ergebnisse wurden genutzt, um ein Arbeitsmodell zum molekularen Ablauf des H-P Wechsels aufzustellen.

Bibliographie / References

  1. Thomas, T., Loveland, K.L. und Voss, A.K. (2007). The genes coding for the MYST family histone acetyltransferases, Tip60 and Mof, are expressed at high levels during sperm development. Gene Expr. Patt. 7, 657–665.
  2. Cross, D.P. und Shellenbarger, D.L. (1979). The dynamics of Drosophila melanogaster spermatogenesis in in vitro cultures. J. Embryol. exp. Morph. 53, 345–351.
  3. Meistrich, M.L., Mohapatra, B., Shirley, C.R. und Zhao, M. (2003). Roles of transition nuclear proteins in spermiogenesis. Chromosoma 111, 483–488.
  4. Cho, C., Willis, W.D., Goulding, E.H., Jung-Ha, H., Choi, Y.C., Hecht, N.B. und Eddy, E.M. (2001). Haploinsufficiency of protamine-1 or -2 causes infertility in mice. Nat. Genet. 28, 82–86.
  5. Sonnack, V., Failing, K., Bergmann, M. und Steger, K. (2002). Expression of hy- peracetylated histone H4 during normal and impaired human spermatogenesis. Andrologia 34, 384–390.
  6. Lindsley, D. und Tokuyasu, K.T. (1980). Spermatogenesis. In: Ashburner, M., Wright, T.R.F. (Eds.), Genetics and Biology of Drosophila. Vol. 2d. Academic Press, New York USA, 225–294.
  7. The common marmoset (Callithrix jacchus) as a model for histone and protamine expression during human spermatogenesis. Hum. Reprod. 24, 536–545.
  8. Steger, K., Fink, L., Failing, K., Bohle, R.M., Kliesch, S., Weidner, W. und Berg- mann, M. (2003). Decreased protamine-1 transcript levels in testes from infertile men. Mol. Hum. Reprod. 9, 331–336.
  9. Tweedie, S., Ashburner, M., Falls, K., Leyland, P., McQuilton, P., Marygold, S., Mill- burn, G., Osumi-Sutherland, D., Schroeder, A., Seal, R., Zhang, H. und The Flyba- se Consortium (2009). FlyBase: enhancing Drosophila Gene Ontology annotations. Nuc. Acids Res. 37, D555–D559.
  10. Yu, J., Pacifico, S., Liu, G. und Finley, R.L. Jr. (2008). DroID: the Drosophila Interacti- ons Database, a comprehensive resource for annotated gene and protein interactions. BMC Genomics 9, 461.
  11. Zhao, M., Shirley, C.R., Mounsey, S. und Meistrich, M.L. (2004). Nucleoprotein transi- tions during spermiogenesis in mice with transition nuclear protein Tnp1 and Tnp2 mutations. Biol. Reprod. 71, 1016–1025.
  12. Sassone-Corsi, P. (2002). Unique chromatin remodeling and transcriptional regulation in spermatogenesis. Science 296, 2176–2178.
  13. Noguchi, T. und Miller, K.G. (2003). A role for actin dynamics in individualization during spermatogenesis in Drosophila melanogaster. Development 130, 1805–1816.
  14. Barreau, C., Benson, E., Gudmannsdottir, E., Newton, F. und White- Cooper, H. (2008). Post-meiotic transcription in Drosophila testes. Development 135, 1897–1902.
  15. Rathke, C., Baarends, W.M., Jayaramaiah-Raja, S., Bartkuhn, M., Renkawitz, R. und Renkawitz-Pohl, R. (2007). Transition from a nucleosome-based to a protamine-based chromatin configuration during spermiogenesis in Drosophila.
  16. Brewer, L., Corzett, M., Lau, E.Y. und Balhorn, R. (2003). Dynamics of Protamine 1 binding to single DNA molecules. J. Biol. Chem. 278, 42403–42408.
  17. Balhorn, R., Brewer, L. und Corzett, M. (2000). DNA condensation by Protamine and arginine-rich peptides: Analysis of toroid stability using single DNA molecules. Mol. Reprod. Dev. 56, 230–234.
  18. Balasubramanyam, K., Altaf, M., Varier, R.A., Swaminathan, V., Ravindran, A., Sadhale, P.P. und Kundu, T.P. (2004). Polyisoprenylated benzophenone, garcinol, a natural histone acetyltransferase inhibitor, represses chromatin transcription and alters global gene expression. J. Biol. Chem. 279, 33716–33726.
  19. Bizzaro, D., Manicardi, G., Bianchi, P.G. und Sakkas, D. (2000). Sperm decondensation during fertilisation in the mouse: presence of DNase I hypersensitive sites in situ and a putative role for topoisomerase II. Zygote 8, 197–202.
  20. Mai, A., Massa, S., Rotili, D., Cerbara, I., Valente, S., Pezzi, R., Simeoni, S. und Ragno, R. (2005). Histone deacetylation in epigenetics: An attractive target for anticancer therapy. Med. Res. Rev. 25, 261–309.
  21. Venables, J.P. und Eperon, I.C. (1999). The roles of RNA-binding proteins in sperma- togenesis and male infertility. Curr. Opin. Genet. Dev. 9, 346–354.
  22. Clarkson, M. und Saint, R. (1999). A His2AvDGFP fusion gene complements a lethal His2AvD mutant allele and provides an in vivo marker for Drosophila chromosome behavior. DNA Cell. Biol. 18, 457–462.
  23. Sambrook, J., Fritsch, E.F. und Maniatis, T. (1989). Molecular Cloning: a Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.
  24. Shirley, C.R., Hayashi, S., Mounsey, S., Yangimachi, R. und Meistrich, M.L. (2004). Abnormalities and reduced reproductive potential of sperm from Tnp1-and Tnp2- null double mutant mice. Biol. Reprod. 71, 1220–1229.
  25. Pivot-Pajot, C., Caron, C., Govin, J., Vion, A., Rousseaux, S. und Khochbin, S. (2003). Acetylation-dependent chromatin reorganization by BRDT, a testis-specific bromodomain-containing protein. Mol. Cell. Biol. 23, 5354–5365.
  26. Kurtz, K., Martínez-Soler, F., Ausió, J. und Chiva, M. (2007). Acetylation of histone H4 in complex structural transitions of spermiogenic chromatin.
  27. Kennedy, B.P. und Davies, P.L. (1980). Acid-soluble nuclear proteins of the testis du- ring spermatogenesis in the winter flounder. Loss of the high mobility group proteins. J. Biol. Chem. 255, 2533–2539.
  28. Carrell, D.T., Emery, B.R. und Hammoud, S. (2007). Altered protamine expression and diminished spermatogenesis: what is the link? Hum. Reprod. Update 13, 313–327.
  29. Sasnauskas, G., Zakrys, L., Zaremba, M., Cosstick, R., Gaynor, J.W., Halford, S.E. und Siksnys, V. (2010). A novel mechanism for the scission of double-stranded DNA: Bfil cuts both 3'-5' and 5'-3' strands by rotating a single active site. Nucleic Acids Res. elektronische Vorabveröffentlichung.
  30. Raukas, E. und Mikelsaar, R.-H. (1999). Are there molecules of nucleoprotamine? BioEssays 21, 440–448.
  31. Kawamoto, T., Kawai, K., Kodama, T., Yokokura, T. und Niki, Y. (2008). Autonomous differentiation of Drosophila spermatogonia in vitro.
  32. Olivieri, G. und Olivieri, A. (1965). Autoradiographic study of nucleic acid synthesis during spermatogenesis in Drosophila melanogaster. Mutat. Res. 2, 366–380.
  33. Lewis, J.D., Song, Y., de Jong, M.E., Bagha, S.M. und Ausió, J. (2003). A walk though vertebrate and invertebrate protamines. Chromosoma 111, 473–482.
  34. Wang, J.C. (2002). Cellular roles of DNA topoisomerases: a molecular perspective. Nat. Rev. Mol. Cell Biol. 3, 430–440.
  35. Costelloe, T., FitzGerald, J., Murphy, N.J., Flaus, A. und Lowndes, N.F. (2006). Chro- matin modulation and the DNA damage response. Exp. Cell Res. 312, 2677–2686.
  36. Kimmins, S. und Sassonne-Corsi, P. (2005). Chromatin remodelling and epigenetic features of germ cells. Nature 434, 583–589.
  37. Ripp, C. (2009). Chromatin remodelling during spermiogenesis: Characterization of the mutant snowballs and analyses of topoisomerase expression during the switch from histones to protamines. Master-Arbeit, Philipps-Universität Marburg.
  38. McPherson, S. und Longo, F.J. (1993). Chromatin structure-functions alterations du- ring mammalian spermatogenesis: DNA nicking and repair in elongating spermatids. Eur. J. Histochem. 37, 109–128.
  39. (1997). Crystal structure of the nucleosome core particle at 2.8 ˚ A resolution.
  40. Ward, W.S. (1993). Deoxyribonucleic acid loop domain tertiary structure in mamma- lian spermatozoa. Biol. Reprod. 48, 1193–1201.
  41. Rathke, C., Barckmann, B., Burkhard, S., Jayaramaiah-Raja, S., Roote, J. und Renkawitz-Pohl, R. (2010). Distinct functions of Mst77F and protamines in Literaturverzeichnis 89 nuclear shaping and chromatin condensation during Drosophila spermiogenesis.
  42. Hammoud, S.S., Nix, D.A., Zhang, H., Purwar, J., Carrell, D.T. und Cairns, B.R. (2009). Distinctive chromatin in human sperm packages genes for embryo develop- ment. Nature 460, 473–478.
  43. Leduc, F., Maquennehan, V., Nkoma, G.B. und Boissonneault, G. (2008). DNA Da- mage response during chromatin remodelling in elongating spermatids of mice. Biol. Reprod. 78, 324–332.
  44. Nagy, Z. und Soutoglou, E. (2009). DNA repair: easy to visualize, difficult to elucidate. Trends Cell Biol. 19, 617–629.
  45. Belote, J.M. und Zhong, L. (2009). Duplicated proteasome subunit genes in Drosophila and their roles in spermatogenesis. Heredity 103, 23–31.
  46. Tokuyasu, K.T., Peacock, W.J. und Hardy, R.W. (1972). Dynamics of spermiogenesis in Drosophila melanogaster. II. Coiling process.
  47. Bendena, W.G., Ayme-Southgate, A., Garbe, J.C. und Pardue, M. L. (1991). Ex- pression of heat-shock locus hsr-omega in nonstressed cells during development in Drosophila melanogaster. Dev. Biol. 144, 65–77.
  48. Lee, K.K. und Workman, J.L. (2007). Histone acetyltransferase complexes: one size doesn't fit all. Nat. Rev. Mol. Cell Biol. 8, 284–295.
  49. Mehnert, J.M. und Kelly, W.K. (2007). Histone Deacetylase Inhibitors: Biology and Mechanism of Action. Cancer J. 13, 23–29.
  50. (Ed.), Histones and other basic nuclear proteins. CRC Press, Boca Raton USA, 347–373.
  51. Olsen, A-K., Lindemann, B., Wiger, R., Duale, N. und Brunborg, G. (2005). How do male germ cells handle DNA damage? Tox. Appl. Pharma. 207, S521–S531.
  52. Shields, G. und Sang, J.H. (1977). Improved medium for culture of Drosophila embryo- nic cells. Drosophila Information Service 52, 161.
  53. Sun, Y., Jiang, X., Chen, S. und Price, B.D. (2006). Inhibition of histone acetyl- transferase activity by anacardic acid sensitizes tumor cells to ionizing radiation. FEBS Lett. 580, 4353–4356.
  54. Liebrich, W. (1981). In vitro spermatogenesis in Drosophila. I. Development of isolated spermatocyte cysts from wild-type D. hydei. Cell Tissue Res. 220, 251–262.
  55. Wu, L. und Belasco, J.G. (2008). Let me count the ways: Mechanisms of gene regulation by miRNAs and siRNAs. Mol. Cell 29, 1–7.
  56. Sng, J.H., Heaton, V.J., Bell, M., Maini, P., Austin, C.A. und Fisher, L.M. (1999). Molecular cloning and characterization of the human topoisomerase IIal- pha and IIbeta genes: evidence for isoform evolution through gene duplication.
  57. Yamashita, Y.M., Jones, D.L. und Fuller, M.T. (2003). Orientation of asymmetric stem cell division by the APC tumor suppressor and centrosome. Science 301, 1547–1550.
  58. Miller, D., Brinkworth, M. und Iles, D. (2009). Paternal DNA packaging in spermatozoa: more than the sum of its parts? Reproduction elektronische Vorabveröffentlichung.
  59. Kennedy, B.P. und Davies, P.L. (1981). Phosphorylation of a group of high mole- cular weight basic nuclear proteins during spermatogenesis in the winter flounder. J. Biol. Chem. 256, 9254–9259.
  60. Haraguchi, C.M., Mabuchi, T., Hirata, S., Shoda, T., Tokumoto, T., Hoshi, K. und Yokota, S. (2007). Possible function of caudal nuclear pocket: Degradation of nucleo- proteins by ubiquitin-proteasome system in rat spermatids and human sperm.
  61. Saiki, R.K., Gelfand, D.H., Stoffel, S., Scharf, S.J., Higuchi, R., Horn, G.T., Mullis, K.B. und Erlich, H.A. (1988). Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239, 487–491.
  62. Torregrosa, N., Domínguez-Fandos, D., Camejo, M.I., Shirley, C.R., Meistrich, M.L., Ballescà, J.L. und Oliva, R. (2006). Protamine 2 precursors, protamine 1/prot- amine 2 ratio, DNA integrity and other sperm parameters in infertile patients. Hum. Reprod. 21, 2084–2089.
  63. Oliva, R. (2006). Protamines and male infertility. Hum. Reprod. Upd. 12, 417–435.
  64. McMurray, C.T. und Kortun, I.V. (2003). Repair in haploid male germ cells occurs late in differentiation as chromatin is condensing. Chromosoma 111, 505–508.
  65. Kuroda, Y. (1974). Spermatogenesis in pharate adult testes of Drosophila in tissue cultures without ecdysones. J. Insect. Physiol. 20, 637–640.
  66. Caron, N., Veilleux, S. und Boissonneault, G. (2001). Stimulation of DNA repair by the spermatidal TP1 protein. Mol. Reprod. Dev. 58, 437–443.
  67. Rabinowitz, M. (1941). Studies on the cytology and early embryology of the egg of Drosophila melanogaster. J. Morphol. 69, 1–49.
  68. Cabrero, J., Palomino-Morales, R.J. und Camacho, J.P.M. (2007). The DNA-repair Ku70 is located in the nucleus and tail of elongating spermatids in grasshoppers. Chromosome Res. 15, 1093–1100.
  69. Shang, E., Nickerson, H.D., Wen, D., Wang, X. und Wolgemuth, D.J. (2007). The first bromodomain of Brdt, a testis-specific member of the BET subfamily of double- bromodomain-containing proteins, is essential for male germ cell differentiation. Development 134, 3507–3515.
  70. Lindsley, D.L. und Lifschytz, E. (1972). The genetic control of spermatogenesis in Drosophila. In: Beatty, R.A. und Gluecksohn-Waelsch, S. (Eds.), Proc. Int. Symp. The genetics of the spermatozoon, Edinburgh. Edinburgh, New York, 203–222.
  71. The germinal proliferation center in the testis of Drosophila melanogaster.
  72. Loppin, B., Bonnefoy, E., Anselme, C., Laurencon, A., Karr, T.L. und Couble, P. (2005). The histone 3.3 chaperone HIRA is essential for chromatin assembly in the male pronucleus. Nature 437, 1386–1390.
  73. von Mikecz, A. (2006). The nuclear ubiquitin-proteasome system.
  74. Bodenstein, D. (1950). The postembryonic development of Drosophila. In: Demerec, M. (Ed. ), Biology of Drosophila. Wiley and Sons, USA, 275–364.
  75. Balhorn, R. (2007). The protamine family of sperm nuclear proteins.
  76. Govin, J., Caron, C., Lestrat, C., Rousseaux, S. und Khochbin, S. (2004). The role of histones in chromatin remodelling during mammalian spermiogenesis. Eur. J. Biochem. 271, 3459–3469.
  77. Gould-Somero, M. und Holland, L. (1974). The timing of RNA synthe- sis for spermiogenesis in organ cultures of Drosophila melanogaster testes.
  78. Samejima, K. und Earnshaw, W.C. (2005). Thrashing the genome: The role of nucleases during apoptosis. Nat. Rev. Mol. Cell Biol. 6, 677-688.
  79. Marcon, L. und Boissonneault, G. (2004). Transient DNA strand breaks during mouse and human spermiogenesis: New insights in stage specificity and link to chromatin remodelling. Biol. Reprod. 70, 910–918.
  80. van der Heijden, G.W., Derijck, A.A.H.A., Ramos, L., Giele, M., van der Vlag, J. und de Boer, P. (2006). Transmission of modified nucleosomes from the mouse male germline to the zygote and subsequent remodelling of paternal chromatin. Dev. Biol. 298, 458–469.
  81. Kiefer, B.I. (1966). Ultrastructural abnormalities in developing sperm of X/O Droso- phila melanogaster. Genetics 54, 1441–1452.
  82. Balhorn, R., Gledhill, B.L. und Wyrobek, A.J. (1977). Mouse sperm chromatin prote- ins: Quantitative isolation and partial characterization. Biochemistry 16, 4074–4080.
  83. Braun, R.E. (2001). Packaging paternal chromosomes with protamine. Nature Gen. 28, 10–12.
  84. Cho, C., Jung-Ha, H., Willis, W.D., Goulding, E.H., Stein, P., Xu, Z., Schultz, R.M., Hecht, N.B. und Eddy, E.M. (2003). Protamine 2 deficiency leads to sperm DNA damage and embryo death in mice. Biol. Reprod. 69, 211–217.
  85. Laberge, R.-M. und Boissonneault, G. (2005). On the nature and origin of DNA strand breaks in elongating spermatids. Biol. Reprod. 73, 289–296.
  86. Nandi, D., Tahiliani, P., Kumar, A. und Chandu, D. (2006). The ubiquitin-proteasome system. J. Biosci. 31, 137-155.
  87. Jayaramaiah Raja, S. und Renkawitz-Pohl, R. (2005). Replacement by Drosophila me- lanogaster Protamines and Mst77F of histones during chromatin condensation in late spermatids and role of Sesame in the removal of these proteins from the male pronucleus. Mol. Cell. Biol. 25, 6165–6177.
  88. Görisch, S.M., Wachsmuth, M., Tóth, K.F., Lichter, P. und Rippe, K. (2005). Histone acetylation increases chromatin accessibility. J. Cell Sci. 118, 5825–5834.
  89. Epoxomicin, a potent and selective proteasome inhibitor, exhibits in vivo antiinflam- matory activity. Proc. Natl. Acad. Sci. USA 96, 10403–10408.
  90. Boutanaev, A.M., Mikhaylova, L.M. und Nurminsky, D.I. (2007). Up-regulation of the Ku heterodimer in Drosophila testicular cyst cells. FEBS Lett. 581, 1707–1715.
  91. Nair, M., Nagamori, I., Sun, P., Mishra, D.P., Rhéaume, C., Li, B., Sassone-Corsi, P. und Dai, X. (2008). Nuclear regulator Pygo2 controls spermiogenesis and histone H3 acetylation. Dev. Biol. 320, 446–455.
  92. Mimitou, E.P. und Symington, L.S. (2009). DNA end resection: Many nucleases make light work. DNA Repair 8, 983–995.
  93. Oliva, R. und Mezquita, C. (1982). Histone H4 hyperacetylation and rapid tur- nover of its acetyl groups in transcriptionally inactive rooster testis spermatids. Nucleic Acids Res. 10, 8049-8059.
  94. Histone H1 and the origin of protamines. PNAS 101, 4148–4152.
  95. Grosse, F. und Manns, A. (1993). Terminal Deoxyribonucleotidyl Transferase (EC 6.5.1.3). In: Burrell, M.M. (Ed.), Enzymes of Molecular Biology. Humana Press, USA, 213–230.
  96. Lieber, M.R. (2008). The mechanism of human nonhomologous DNA end joining.
  97. Holstein, A.F., Schulze, W. und Davidoff, M. (2003). Understanding spermatogenesis is a prerequisite for treatment. Reprod. Biol. Endocrinol. 1, 107–661.
  98. Bassi, L. und Palitti, F. (2000). Anti-topoisomerase drugs as potent inducers of chro- mosomal aberrations. Gen. Mol. Biol. 23, 1065–1069.


* Das Dokument ist im Internet frei zugänglich - Hinweise zu den Nutzungsrechten