A protein homologous to human Ku p70-protein is required for reconstitution of Xenopus sperm pronuclei.
The monoclonal antibody mAbH6, which recognizes one human Ku-protein (p70), cross-reacted with the counterpart protein from Xenopus eggs. The Xenopus antigen purified with a mAbH6-Sepharose column was a complex of 88 kDa and 72 kDa proteins. The role of Ku protein in nuclear structure formation was studied using a cell-free nuclear assembly extract derived from Xenopus interphase eggs. The protein was distributed on the surface of demembranated sperm chromatin and in the membrane vesicle fraction of the nuclear assembly extract. Addition of mAbH6 to the assembly extracts prevented the completion of reconstitution of pronuclei from demembranated sperm chromatins, although partial decondensation mediated by nucleoplasmin was not impeded. As a result, the sperm chromatin remained in partially swollen structures or formed round but small anomalous nuclei. Nuclear membranes were formed on the nuclei in mAbH6-inhibited extract systems, but DNA synthesis was largely decreased, suggesting the incomplete reconstitution of pronuclei. The incorporation of lamin to the nuclei was inhibited by mAbH6. It is suggested that the Xenopus Ku-homologous protein has a role in the formation of the lamin layer after nuclear membrane reconstitution.
Higashiura
M
M
Department of Biology, Faculty of Science, Kwansei Gakuin University, Hyogo, Japan.
Takasuga
Y
Y
Yamashita
J
J
Yagura
T
T
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D034961:Antigens, Nuclear; D002467:Cell Nucleus / Q000502:physiology* / Q000648:ultrastructure; D002843:Chromatin / Q000502:physiology / Q000648:ultrastructure*; D003429:Cross Reactions; D004265:DNA Helicases; D004268:DNA-Binding Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D000072200:Ku Autoantigen; D008297:Male; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013094:Spermatozoa / Q000166:cytology / Q000502:physiology*; D013938:Thymidine Monophosphate / Q000378:metabolism; D013942:Thymine Nucleotides / Q000378:metabolism; D014981:Xenopus
D000911:Antibodies, Monoclonal; D034961:Antigens, Nuclear; D002843:Chromatin; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D013942:Thymine Nucleotides; D013938:Thymidine Monophosphate; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen; C024157:thymidine 5'-triphosphate