| Budget Amount *help |
¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 1988: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1987: ¥900,000 (Direct Cost: ¥900,000)
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| Research Abstract |
During aggregation of Dictyostelium discoideum, nucleoli protruded from the anterior of the nucleus and elongated to from nozzle-like structures. The structure and function of deformed nucleoli were analyzed by methods of rapid freezing, indirect-immunofluoresence cell fractionation as well as two-dimensional gel electrophoresis.
1. The mechanism of structural changes of the nucleolus.
(1) The formation of nozzles depends on microtubules (MTs) bound to the nuclear envelope (NE): In aggregating cells, the nucleus and nozzle were surrounded by more than 10 MTs elongated from the microtubule-organizing center (MTOC). Both nozzles and MTs were disappeared in the presence of nocodazole. The MTOC and MTs were both attached to the nuclei isolated from aggregating cells. In contrast, only the MTOC was bound to the nucleus at the growing stage.
(2) The position of protrusions is determined by MTs but their structures are possibly based on actin fibers: By 2-D gel analysis of nucleoli from both growing and aggregating cells, actin was one of the major component. In the presence of cytochalasin B, nozzles were found but not protrusions. In contrast, protrusions remained on the side of the nucleus by treatment with nocodazole.
(3) The assumption is as follows: At the aggregation stage, nucleoli attached to the NE become protruded depending on actin fibers. MTs link to the NE surface by molecules like kinesin. Then, protrusions move to the front of the nucleus and nozzles are elongated by anterograded movements of NE.
2. Function of protrusions and nozzles: In the presence of nocodazole or cytochalasin, aggregation was delayed, howeer, cells developed normally. Diffrentiation ceased just after cells aggregated nuder submerged conditions. In any case, either protrusions or nozzles did not appeared. Therefore, deformed nucleoli are not necessary for development, rather they may support effective cell migration and differentiation during aggregation.
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