Skip to main content Accessibility help

Multiple histone site epigenetic modifications in nuclear transfer and in vitro fertilized bovine embryos

  • Xia Wu (a1), Yan Li (a1), Lian Xue (a1), Lingling Wang (a1), Yongli Yue (a1), Kehan Li (a1), Shorgan Bou (a1), Guang-Peng Li (a2) (a1) and Haiquan Yu (a2) (a1)...


During mammalian embryonic development, DNA methylation and histone modifications are important in gene expression regulation and epigenetic reprogramming. In cloned embryos, high levels of DNA methylation and abnormal demethylation were widely observed during the preimplantation period. Little is known whether there is a difference in histone modifications between in vitro fertilization (IVF) and cloned embryos during preimplantation development. In the present study, the distributions and intensity patterns of acetylations in H3 lysine 9, 18 and H4 lysine 8, 5 and tri-methyl lysine 4 and dimethyl-lysine 9 in histone H3 were compared in cloned and IVF bovine preimplantation embryos by using indirect immunofluorescence and scanning confocal microscopy. The results showed that the acetylation and methylation levels of H3K9ac, H3K18ac, H4K5ac, H4K8ac, H3K4me3 and H3K9me2 were abnormally high in the cloned embryos from the pronuclear to the 8-cell stage. H4K8ac and H4K5ac in the cloned embryos were particularly abnormal when compared with the IVF controls. At the blastocyst stage differences dissipated between cloned and IVF embryos and the distribution and intensity patterns of all histone modifications showed no obvious difference. These results suggest that somatic cells in recipient oocytes produced aberrant histone modifications at multiple sites before the donor cell genome is activated. After zygotic genome activation, distributions and intensity patterns of histone modifications were comparable with both cloned and IVF embryos.


Corresponding author

All correspondence to: Haiquan Yu and Guang-Peng Li. The Key Laboratory of Mammalian Reproductive Biology and Biotechnology of the Ministry of Education, Inner Mongolia University, Hohhot 010021, China. Tel: +86 471 4992495. Fax: +86 471 4995071. e-mail: and


Hide All
Adenot, P.G., Mercier, Y., Renard, J.P. & Thompson, E.M. (1997). Differential H4 acetylation of paternal and maternal chromatin precedes DNA replication and differential transcriptional activity in pronuclei of 1-cell mouse embryos. Development 124, 4615–25.
Arney, K.L., Bao, S., Bannister, A.J., Kouzarides, T. & Surani, M.A. (2002). Histone methylation defines epigenetic asymmetry in the mouse zygote. Int. J. Dev. Biol. 46, 317–20.
Beaujean, N., Hartshorne, G., Cavilla, J., Taylor, J., Gardner, J., Wilmut, I., Meehan, R. & Young, L. (2004). Non-conservation of mammalian preimplantation methylation dynamics. Curr. Biol. 14, R2667.
Bernstein, B.E., Kamal, M., Lindblad-Toh, K., Bekiranov, S., Bailey, D.K., Huebert, D.J., McMahon, S., Karlsson, E.K., Kulbokas, E.J., 3rd, Gingeras, T.R., Schreiber, S.L. & Lander, E.S. (2005). Genomic maps and comparative analysis of histone modifications in human and mouse. Cell 120, 169–81.
Bertos, N.R., Wang, A.H. & Yang, X.J. (2001). Class II histone deacetylases: structure, function and regulation. Biochem. Cell. Biol. 79, 243–52.
Biel, M., Wascholowski, V. & Giannis, A. (2005). Epigenetics—an epicenter of gene regulation: histones and histone-modifying enzymes. Angew. Chem. Int. Ed. Engl. 44, 3186–216.
Bjerling, P., Silverstein, R.A., Thon, G., Caudy, A., Grewal, S. & Ekwall, K. (2002). Functional divergence between histone deacetylases in fission yeast by distinct cellular localization and in vivo specificity. Mol. Cell. Biol. 22, 2170–81.
Brackett, B.G. & Oliphant, G. (1975). Capacitation of rabbit spermatozoa in vitro. Biol. Reprod. 12, 260–74.
Brevini, T.A., Cillo, F., Antonini, S., Tosetti, V. & Gandolfi, F. (2007). Temporal and spatial control of gene expression in early embryos of farm animals. Reprod. Fertil. Dev. 19, 3542.
Dean, W., Santos, F., Stojkovic, M., Zakhartchenko, V., Walter, J., Wolf, E. & Reik, W. (2001). Conservation of methylation reprogramming in mammalian development: aberrant reprogramming in cloned embryos. Proc. Natl. Acad. Sci. USA 98, 1373413738.
Dean, W., Santos, F. & Reik, W. (2003). Epigenetic reprogramming in early mammalian development and following somatic nuclear transfer. Semin. Cell. Dev. Biol. 14, 93100.
Fulka, H., Mrazek, M., Tepla, O. & Fulka, J. Jr. (2004). DNA methylation pattern in human zygotes and developing embryos. Reproduction 128, 703–8.
Fulka, J., Fulka, H., Slavik, T., Okada, K. & Fulka, J. Jr (2006). DNA methylation pattern in pig in vivo produced embryos. Histochem. Cell. Biol. 126, 213–7.
Grant, P.A., Eberharter, A., John, S., Cook, R.G., Turner, B.M. & Workman, J.L. (1999). Expanded lysine acetylation specificity of Gcn5 in native complexes. J. Biol. Chem. 274, 5895–900.
Gurdon, J.B., Laskey, R.A., De Robertis, E.M. & Partington, G.A. (1979). Reprogramming of transplanted nuclei in amphibia. Int. Rev. Cytol. 9, 161–78.
Holker, M., Petersen, B., Hassel, P., Kues, W.A., Lemme, E., Lucas-Hahn, A. & Niemann, H. (2005). Duration of in vitro maturation of recipient oocytes affects blastocyst development of cloned porcine embryos. Cloning Stem Cells 7, 3544.
Huang, J.C., Lei, Z.L., Shi, L.H., Miao, Y.L., Yang, J.W., Ouyang, Y.C., Sun, Q.Y. & Chen, D.Y. (2007). Comparison of histone modifications in in vivo and in vitro fertilization mouse embryos. Biochem. Biophys. Res. Commun. 354, 7783.
Kim, J.M., Ogura, A., Nagata, M. & Aoki, F. (2002). Analysis of the mechanism for chromatin remodeling in embryos reconstructed by somatic nuclear transfer. Biol. Reprod. 67, 760–6.
Kim, J.M., Liu, H., Tazaki, M., Nagata, M. & Aoki, F. (2003). Changes in histone acetylation during mouse oocyte meiosis. J. Cell. Biol. 162, 3746.
Kishigami, S., Mizutani, E., Ohta, H., Hikichi, T., Thuan, N.V., Wakayama, S., Bui, H.T. & Wakayama, T. (2006). Significant improvement of mouse cloning technique by treatment with trichostatin A after somatic nuclear transfer. Biochem. Biophys. Res. Commun. 340, 183–9.
Kouzarides, T. (2007). Chromatin modifications and their function. Cell 128, 693705.
Kurdistani, S.K., Tavazoie, S. & Grunstein, M. (2004). Mapping global histone acetylation patterns to gene expression. Cell 117, 721–33.
Lachner, M. & Jenuwein, T. (2002). The many faces of histone lysine methylation. Curr. Opin. Cell. Biol. 14, 286–98.
Lachner, M., O'Sullivan, R.J. & Jenuwein, T. (2003). An epigenetic road map for histone lysine methylation. J. Cell. Sci. 116, 2117–24.
Lepikhov, K. & Walter, J. (2004). Differential dynamics of histone H3 methylation at positions K4 and K9 in the mouse zygote. BMC Dev. Biol. 4, 12.
Liu, H., Kim, J.M. & Aoki, F. (2004). Regulation of histone H3 lysine 9 methylation in oocytes and early pre-implantation embryos. Development 131, 2269–80.
Liu, X.Y., Mal, S.F., Miao, D.Q., Liu, D.J., Bao, S. & Tan, J.H. (2005). Cortical granules behave differently in mouse oocytes matured under different conditions. Hum. Reprod. 20, 3402–13.
Marmorstein, R. & Roth, S.Y. (2001). Histone acetyltransferases: function, structure and catalysis. Curr. Opin. Genet. Dev. 11, 155–61.
Martin, C. & Zhang, Y. (2005). The diverse functions of histone lysine methylation. Nat. Rev. Mol. Cell. Biol. 6, 838–49.
McGraw, S., Robert, C., Massicotte, L. & Sirard, M.A. (2003). Quantification of histone acetyltransferase and histone deacetylase transcripts during early bovine embryo development. Biol. Reprod. 68, 383–9.
Meissner, A. & Jaenisch, R. (2006). Mammalian nuclear transfer. Dev. Dyn. 235, 2460–9.
Mizzen, C.A. & Allis, C.D. (1998). Linking histone acetylation to transcriptional regulation. Cell. Mol. Life Sci. 54, 620.
Noma, K., Allis, C.D. & Grewal, S.I. (2001). Transitions in distinct histone H3 methylation patterns at the heterochromatin domain boundaries. Science 293, 1150–5.
O'Neill, L.P. & Turner, B.M. (1995). Histone H4 acetylation distinguishes coding regions of the human genome from heterochromatin in a differentiation-dependent but transcription-independent manner. EMBO J. 14, 3946–57.
Park, J.S., Jeong, Y.S., Shin, S.T., Lee, K.K. & Kang, Y.K. (2007). Dynamic DNA methylation reprogramming: active demethylation and immediate remethylation in the male pronucleus of bovine zygotes. Dev. Dyn. 236, 2523–33.
Pereira, D.C., Dode, M.A. & Rumpf, R. (2005). Evaluation of different culture systems on the in vitro production of bovine embryos. Theriogenology 63, 1131–41.
Rybouchkin, A., Kato, Y. & Tsunoda, Y. (2006). Role of histone acetylation in reprogramming of somatic nuclei following nuclear transfer. Biol. Reprod. 74, 1083–9.
Santos-Rosa, H., Schneider, R., Bannister, A.J., Sherriff, J., Bernstein, B.E., Emre, N.C., Schreiber, S.L., Mellor, J. & Kouzarides, T. (2002). Active genes are tri-methylated at K4 of histone H3. Nature 419, 407–11.
Santos, F. & Dean, W. (2004). Epigenetic reprogramming during early development in mammals. Reproduction 127, 643–51.
Santos, F., Hendrich, B., Reik, W. & Dean, W. (2002). Dynamic reprogramming of DNA methylation in the early mouse embryo. Dev. Biol. 241, 172–82.
Santos, F., Peters, A.H., Otte, A.P., Reik, W. & Dean, W. (2005). Dynamic chromatin modifications characterise the first cell cycle in mouse embryos. Dev. Biol. 280, 225–36.
Santos, F., Zakhartchenko, V., Stojkovic, M., Peters, A., Jenuwein, T., Wolf, E., Reik, W. & Dean, W. (2003). Epigenetic marking correlates with developmental potential in cloned bovine preimplantation embryos. Curr. Biol. 13, 1116–21.
Sarmento, O.F., Digilio, L.C., Wang, Y., Perlin, J., Herr, J.C., Allis, C.D. & Coonrod, S.A. (2004). Dynamic alterations of specific histone modifications during early murine development. J. Cell Sci. 117, 4449–59.
Schiltz, R.L., Mizzen, C.A., Vassilev, A., Cook, R.G., Allis, C.D. & Nakatani, Y. (1999). Overlapping but distinct patterns of histone acetylation by the human coactivators p300 and PCAF within nucleosomal substrates. J. Biol. Chem. 274, 1189–92.
Schultz, R.M. (1993). Regulation of zygotic gene activation in the mouse. Bioessays 15, 531–8.
Shi, L.H., Ai, J.S., Ouyang, Y.C., Huang, J.C., Lei, Z.L., Wang, Q., Yin, S., Han, Z.M., Sun, Q.Y. & Chen, D.Y. (2008). Trichostatin A and nuclear reprogramming of cloned rabbit embryos. J. Anim. Sci. 86, 1106–13.
Solter, D. (2000). Mammalian cloning: advances and limitations. Nat. Rev. Genet. 1, 199207.
Stallcup, M.R. (2001). Role of protein methylation in chromatin remodeling and transcriptional regulation. Oncogene 20, 3014–20.
Stein, P., Worrad, D.M., Belyaev, N.D., Turner, B.M. & Schultz, R.M. (1997). Stage-dependent redistributions of acetylated histones in nuclei of the early preimplantation mouse embryo. Mol. Reprod. Dev. 47, 421–9.
Suteevun, T., Parnpai, R., Smith, S.L., Chang, C.C., Muenthaisong, S. & Tian, X.C. (2006). Epigenetic characteristics of cloned and in vitro-fertilized swamp buffalo (Bubalus bubalis) embryos. J. Anim. Sci. 84, 2065–71.
Turner, B.M. (1991). Histone acetylation and control of gene expression. J. Cell Sci. 99, 1320.
Turner, B.M. (1998). Histone acetylation as an epigenetic determinant of long-term transcriptional competence. Cell. Mol. Life Sci. 54, 2131.
Valls, E., Sanchez-Molina, S. & Martinez-Balbas, M.A. (2005). Role of histone modifications in marking and activating genes through mitosis. J. Biol. Chem. 280, 42592–600.
van der Heijden, G.W., Derijck, A.A., Ramos, L., Giele, M., van der Vlag, J. & de Boer, P. (2006). Transmission of modified nucleosomes from the mouse male germline to the zygote and subsequent remodeling of paternal chromatin. Dev. Biol. 298, 458–69.
Vigneault, C., McGraw, S., Massicotte, L. & Sirard, M.A. (2004). Transcription factor expression patterns in bovine in vitro-derived embryos prior to maternal-zygotic transition. Biol. Reprod. 70, 1701–9.
Wang, F., Kou, Z., Zhang, Y. & Gao, S. (2007). Dynamic reprogramming of histone acetylation and methylation in the first cell cycle of cloned mouse embryos. Biol. Reprod. 77, 1007–16.
Wilmut, I., Schnieke, A.E., McWhir, J., Kind, A.J. & Campbell, K.H. (1997). Viable offspring derived from fetal and adult mammalian cells. Nature 385, 810–13.
Worrad, D.M., Turner, B.M. & Schultz, R.M. (1995). Temporally restricted spatial localization of acetylated isoforms of histone H4 and RNA polymerase II in the 2-cell mouse embryo. Development 121, 2949–59.
Wrenzycki, C., Wells, D., Herrmann, D., Miller, A., Oliver, J., Tervit, R. & Niemann, H. (2001). Nuclear transfer protocol affects messenger RNA expression patterns in cloned bovine blastocysts. Biol. Reprod. 65, 309–17.
Yan, C. & Boyd, D.D. (2006). Histone H3 acetylation and H3 K4 methylation define distinct chromatin regions permissive for transgene expression. Mol. Cell. Biol. 26, 6357–71.
Yang, J., Yang, S., Beaujean, N., Niu, Y., He, X., Xie, Y., Tang, X., Wang, L., Zhou, Q. & Ji, W. (2007). Epigenetic marks in cloned rhesus monkey embryos: comparison with counterparts produced in vitro. Biol. Reprod. 76, 3642.


Multiple histone site epigenetic modifications in nuclear transfer and in vitro fertilized bovine embryos

  • Xia Wu (a1), Yan Li (a1), Lian Xue (a1), Lingling Wang (a1), Yongli Yue (a1), Kehan Li (a1), Shorgan Bou (a1), Guang-Peng Li (a2) (a1) and Haiquan Yu (a2) (a1)...


Full text views

Total number of HTML views: 0
Total number of PDF views: 0 *
Loading metrics...

Abstract views

Total abstract views: 0 *
Loading metrics...

* Views captured on Cambridge Core between <date>. This data will be updated every 24 hours.

Usage data cannot currently be displayed