Microinjection of an antibody to the Ku protein arrests development in sea urchin embryos.
Ku is the regulatory subunit of the DNA-dependent protein kinase (DNA-PK). This enzyme plays a role in DNA repair, recombination, and transcription. It is composed of a large catalytic subunit (p460), and a regulatory heterodimer, the Ku protein, which consists of 86-kDa and 70-kDa subunits. These various components of the enzyme have been found in both eggs and embryos of the sea urchin. When variable amounts of a specific monoclonal antibody to the Ku protein (Ku 162) were injected into one cell of a 2-cell embryo of Lytechinus pictus, they caused a dose-dependent developmental arrest of the injected cell. The non-injected cell continued to develop normally. In contrast, injection of an antibody (N3H10) raised against the 70-kDa subunit of the Ku protein had no effect on development when injected into 2-cell-stage embryos. Co-injection of purified DNA-PK with the antibody reversed the antibody-mediated inhibition of development. In the fertilized egg and during the early stages of development, the DNA-PK was localized largely in the cytoplasm, but in later developmental stages, it assumed a nuclear location. On the basis of these results, we postulate that the injection of the Ku antibody either prevents the translocation of the DNA-PK into the nucleus or interferes with its enzymatic activity either in the nucleus or in the cytoplasm. In either case, the results suggest that DNA-PK plays an important role in regulating the early stages of embryogenesis in this primitive organism.
Kanungo
J
J
Institute of Molecular Medicine and Genetics, Medical College of Georgia, Augusta 30912, USA.
Empson
R M
RM
Rasmussen
H
H
D000818:Animals; D000911:Antibodies, Monoclonal / Q000008:administration & dosage*; D034961:Antigens, Nuclear; D002455:Cell Division; D004265:DNA Helicases; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000502:physiology*; D047108:Embryonic Development; D006801:Humans; D000072200:Ku Autoantigen; D051379:Mice; D008845:Microinjections; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000008:administration & dosage / Q000502:physiology; D012617:Sea Urchins / Q000196:embryology*
D000911:Antibodies, Monoclonal; D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; C000606510:Xrcc6 protein, mouse; D000072200:Ku Autoantigen