Rediscovery of the Nucleolinus

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DESCRIPTION (provided by applicant): The nucleolinus is an organelle present in many, if not all cells. Its behavior during mitosis and neoplastic transformation led early investigators to propose a role in cell division. Yet, more than 100 years since its discovery, virtually nothing is known about the composition or function of the nucleolinus. Because it is closely apposed to the nucleolus and not easily visible in most cells, it is often mistaken as a part of the nucleolus. Fortunately, there are some cells, such as surf clam (Spisula solidissima) oocytes, in which the nucleolinus is prominent. Such exceptions provide clues to a role for the nucleolinus in cell physiology and opportunities to study function. Using Spisula oocytes as a model system, we have made three observations that potentially link the nucleolinus to the centrosome, the principle microtubule organizing center of animal cells. First is that centrosomal RNAs are specifically localized to the nucleolinus before centrosome formation. Second, the last visible position of the nucleolinus during meiosis coincides with the site of the meiotic spindle pole, where the centrosome organizes astral and spindle microtubules in preparation for cell division. Third, preliminary experiments in which the nucleolinus was ablated before centrosome formation resulted in a blockage of meiosis. These observations lead us to hypothesize that the nucleolinus is required for de novo centrosome formation. The Spisula oocyte model will be used to test this hypothesis in two complementary approaches: 1) laser ablation of the nucleolinus in situ followed by examination of its effects on assembly of centrosomes and other components of the cell division apparatus; 2) analysis of centrosome formation from isolated nucleolini in a cell-free system. We anticipate that these experiments will delineate a model in which the nucleolinus is activated by exposure to oocyte cytoplasm, and then functions in forming the first oocyte centrosome(s). Such a finding would represent a fundamental advance in our understanding of centrosome biogenesis and have broad implications for cell and biomedical sciences.