title: Nucleoplasmin facilitates reprogramming and in vivo development of bovine nuclear transfer embryos.
abstract: Successful cloning by somatic cell nuclear transfer (NT) involves an oocyte-driven transition in gene expression from an inherited somatic pattern, to an embryonic form, during early development. This reprogramming of gene expression is thought to require the remodeling of somatic chromatin and as such, faulty and/or incomplete chromatin remodeling may contribute to the aberrant gene expression and abnormal development observed in NT embryos. We used a novel approach to supplement the oocyte with chromatin remodeling factors and determined the impact of these molecules on gene expression and development of bovine NT embryos. Nucleoplasmin (NPL) or polyglutamic acid (PGA) was injected into bovine oocytes at different concentrations, either before (pre-NT) or after (post-NT) NT. Pre-implantation embryos were then transferred to bovine recipients to assess in vivo development. Microinjection of remodeling factors resulted in apparent differences in the rate of blastocyst development and in pregnancy initiation rates in both NPL- and PGA-injected embryos, and these differences were dependent on factor concentration and/or the time of injection. Post-NT NPL-injected embryos that produced the highest rate of pregnancy also demonstrated differentially expressed genes relative to pre-NT NPL embryos and control NT embryos, both of which had lower pregnancy rates. Over 200 genes were upregulated following post-NT NPL injection. Several of these genes were previously shown to be downregulated in NT embryos when compared to bovine IVF embryos. These data suggest that addition of chromatin remodeling factors to the oocyte may improve development of NT embryos by facilitating reprogramming of the somatic nucleus.
authors.affiliation: Infigen, Inc., Madison, Wisconsin 53718, USA.
authors.forename: Jeffery M
authors.initials: JM
authors.lastname: Betthauser
authors.forename: Martha
authors.initials: M
authors.lastname: Pfister-Genskow
authors.forename: Hongzhi
authors.initials: H
authors.lastname: Xu
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Golueke
authors.forename: Jenine C
authors.initials: JC
authors.lastname: Lacson
authors.forename: Richard W
authors.initials: RW
authors.lastname: Koppang
authors.forename: Cena
authors.initials: C
authors.lastname: Myers
authors.forename: Bing
authors.initials: B
authors.lastname: Liu
authors.forename: Ina
authors.initials: I
authors.lastname: Hoeschele
authors.forename: Kenneth J
authors.initials: KJ
authors.lastname: Eilertsen
authors.forename: Gregory H
authors.initials: GH
authors.lastname: Leno
mesh_terms: D000818:Animals; D002417:Cattle; D002467:Cell Nucleus; D042002:Chromatin Assembly and Disassembly; D019976:Cloning, Organism; D004622:Embryo, Mammalian / Q000502:physiology*; D004625:Embryo, Nonmammalian; D005260:Female; D020869:Gene Expression Profiling; D009687:Nuclear Proteins / Q000378:metabolism*; D053652:Nuclear Transfer Techniques; D056489:Nucleoplasmins; D020411:Oligonucleotide Array Sequence Analysis; D010750:Phosphoproteins / Q000378:metabolism*; D011099:Polyglutamic Acid / Q000378:metabolism; D011247:Pregnancy; D014982:Xenopus laevis
chemical_list: D009687:Nuclear Proteins; D056489:Nucleoplasmins; D010750:Phosphoproteins; D011099:Polyglutamic Acid