Studies on outgrowth of pseudopodial cables and spicule rod formation in cultured derived from micromeres of sea urchin embryos.

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Micromere-derived cells of sea urchin embryos are known to produce pseudopodial cables and spicule rods during culture with horse serum or blastocoelic fluid isolated from sea urchin embryos. In the present study, insulin was found to induce the cable growth without spicule rod formation. Insulin binding protein was found by SDS-PAGE of insulin treated cells. Insulin bound to the cells with Kd of about 0.5 pM and bound insulin was replaced by unknown compounds in horse serum and blastocoelic fluid. This indicates that horse serum and blastocoelic fluid contain insulin-like compounds. Insulin binding in the cells increased in 6 hr period due to translocation of the receptor protein from microsome to plasma membrane and was maintained high up to 40 hr of culture in the absence of horse serum or insulin. In the cells with high insulin binding capacity, insulin, as well as horse serum, augmentation of RNA synthesis occurred following activation of protein phosphorylation and finally resulted in the cable growth. Inhibition of protein phosphorylation reduced the rate of RNA synthesis and inhibition of RNA synthesis by actinomycin D after insulin-induced augmentation of protein phosphorylation resulted in blockage of pseudopodial cable grwoth. In the cells kept with horse serum, the rates of protein phosphorylation and RNA synthesis remained high even after the initiation of cable growth. Actinomycin D introduced after the cable growth initiation allowed the cells treated with horse serum or insulin to cause pseudopodial cables but blocked the spicule formation in horse serum-treated cells. Horse serum and blastocoelic fluid seems to contain signalling compounds other than insulin.