Expression and segregation of nucleoplasmin during development in Xenopus

The spatial segregation of informational molecules in the unfertilized egg and embryo has been hypothesized to be a necessary phenomenon for the normal progression of development leading to the determination of cellular phenotypes. This study describes the selection of a monoclonal antibody (Mab: 2G6) that identifies an antigen (Ag: 2G6) which is localized in the germinal vesicle of oocytes and has a discrete pattern of inheritance during embryogenesis. The antigen displayed biochemical and physical characteristics very similar to nucleoplasmin, which is the histone-binding and nucleosome-assembly protein previously described. Immunoblot analysis with purified oocyte nucleoplasmin confirmed this relationship. Indirect immunofluorescence was used to study the temporal expression and spatial distribution of nucleoplasmin. From early cleavage stages through gastrulation, it is preferentially localized in nuclei of blastomeres at the animal pole. By tadpole stages, it was detected only in nuclei of postmitotic cells of the central nervous system and in nuclei of striated muscle. It was not detected in adult tissues. Western blot analysis during embryogenesis revealed at least five immunologically related polypeptides that displayed distinct patterns of expression during development. The different species observed most likely represent different levels of phosphorylation of nucleoplasmin. The more acidic forms, known to be more active in nucleosome assembly, were present during cleavage stages. Analysis of labelled oocyte proteins by two-dimensional immunoblots and autoradiography revealed that synthesis of nucleoplasmin was first detected in stage-2 oocytes, reached 60% maximum levels at stage 3, peaked at stage 4 and was undetectable in stage-6 oocytes. The amount of nucleoplasmin message present does not follow a similar pattern during oogenesis. These results suggest that the message undergoes pronounced changes in translational efficiency during oogenesis. A comparative immunoblot analysis using proteins from a variety of adult tissues revealed that, whereas the polyclonal antisera against amphibian vitellogenic oocyte nucleoplasmin recognized several different, tissue-specific polypeptides, two different monoclonal antibodies (Mab: b7-1D1, Mab: 2G6) failed to recognize any of the adult tissues tested. We conclude that nucleoplasmin is a family of closely related proteins with distinct embryonic and adult members.