title: Dynamic distribution of NuMA and microtubules in human fetal fibroblasts, developing oocytes and somatic cell nuclear transferred embryos.
abstract: BACKGROUND
  The nuclear mitotic apparatus (NuMA) plays a central role in the assembly and maintenance of spindle poles. Somatic cell nuclear transfer (SCNT) studies on non-human primates have shown that meiotic spindle removal during enucleation causes depletion of NuMA and the minus-end-directed motor protein (HSET) from the ooplasm, and this in turn leads to failure of embryo development. To determine whether NuMA from somatic cells could compensate for NuMA loss during enucleation, the distribution of NuMA and microtubule organization were investigated in human fibroblasts, developing oocytes and SCNT embryos.
  
  
  METHODS
  Human fetal fibroblasts, oocytes at various maturation stages and human embryos reconstructed by different SCNT methods were analyzed for NuMA and α-tubulin using immunofluorescent confocal microscopy.
  
  
  RESULTS
  NuMA was detected in interphase nuclei of fibroblasts and oocytes. During mitosis and meiosis, NuMA relocated to the domain surrounding the two spindle poles. During the enucleation process, NuMA was removed along with the meiotic spindle. At 2 h after injection into a donor cell, transitory bipolar spindles were organized and NuMA was detected in the reformed poles. NuMA could be detected spreading uniformly across the nucleoplasm of one pseudo-pronucleus in SCNT embryos but was excluded from the nucleolus. Regardless of the method used for SCNT (enucleation-injection or injection-pronuclei enucleation), NuMA aggregated and relocated to the reformed spindle poles at metaphase of the first mitotic event. At interphase, NuMA relocated throughout the nucleus in developmentally arrested SCNT embryos.
  
  
  CONCLUSIONS
  Our results show that donor cell nuclei contain NuMA, which might contribute to the maintenance of spindle morphology in SCNT embryos. Normal spindle and NuMA expression were found in human SCNT embryos at different developmental stages.
authors.affiliation: Institute of Reproductive and Stem Cell Engineering, Central South University, Changsha 410078, People's Republic of China.
authors.forename: Xiaoming
authors.initials: X
authors.lastname: Xu
authors.forename: Xin
authors.initials: X
authors.lastname: Duan
authors.forename: Changfu
authors.initials: C
authors.lastname: Lu
authors.forename: Ge
authors.initials: G
authors.lastname: Lin
authors.forename: Guangxiu
authors.initials: G
authors.lastname: Lu
mesh_terms: D034961:Antigens, Nuclear / Q000032:analysis / Q000378:metabolism*; D018797:Cell Cycle Proteins; D004622:Embryo, Mammalian / Q000378:metabolism* / Q000648:ultrastructure; D005333:Fetus / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D007399:Interphase; D008870:Microtubules / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure*; D034521:Nuclear Matrix-Associated Proteins / Q000032:analysis / Q000378:metabolism*; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000254:growth & development / Q000378:metabolism* / Q000648:ultrastructure; D008941:Spindle Apparatus / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure*; D014404:Tubulin / Q000032:analysis / Q000378:metabolism
chemical_list: D034961:Antigens, Nuclear; D018797:Cell Cycle Proteins; C050045:NUMA1 protein, human; D034521:Nuclear Matrix-Associated Proteins; D014404:Tubulin