Changes in the Distribution of Calcium-Sequestering Membranes during the First Cell Cycle of the Sea Urchin, Lytechinus variegatus

Chlortetracycline (CTC) has been used to study sequential changes in the distribution of calcium-sequestering membranes during the first cell cycle of fertilized sea urchin eggs CTC staining patterns first appear as a diffuse ring around the centered zygote nucleus at the time of syngamy. As development proceeds, the ring becomes brighter and then elongates concurrently with the formation of the streak apparatus. Fluorescence subsequently accumulates in the centrospheres of the developing mitotic apparatus and is present in mitotic asters throughout mitosis. When the mitotic apparatus disappears, the fluorescence associated with each aster condenses into a bright ring surrounding each daughter nucleus. Ultrastructural studies show that CTC-fluorescent areas are rich in membranes while experiments with rhodamine 123, a mitochondrion-specific laser dye, indicate that mitochondria are excluded from areas in which membranes accumulate. Microtubule inhibitors prevent the initial accumulation of fluorescence around the zygote nucleus and arrest the development of existing fluorescence patterns when applied at later stages. In contrast, changes in fluorescence patterns are unaffected by the microfilament inhibitor cytochalasin D. These observations show that calcium-sequestering membranes are associated consecutively with the sperm aster, streak, and mitotic apparatus and that the continual reorganization of these membranes during the first cell cycle depends on the assembly and disassembly of microtubules.