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

在从海胆胚胎中分离的马血清或囊胚液培养过程中，已知海胆胚胎微粒源性细胞产生假足索和针状杆。本研究发现胰岛素可诱导索生长，但不形成针状杆。胰岛素处理细胞的SDS-PAGE检测到胰岛素结合蛋白。与细胞结合的胰岛素Kd约为0.5 pM，结合胰岛素在马血清和囊胚液中被未知化合物所取代。这表明马血清和囊胚腔液含有胰岛素样化合物。由于受体蛋白从微粒体转位到质膜，细胞中的胰岛素结合在6小时内增加，并且在没有马血清或胰岛素的情况下保持高水平直至培养40小时。在具有高胰岛素结合能力的细胞、胰岛素和马血清中，RNA合成随着蛋白磷酸化激活而增加，最终导致索生长。抑制蛋白磷酸化降低RNA合成速率，胰岛素诱导的蛋白磷酸化增强导致假足索生长受阻，放线菌素D抑制RNA合成。在与马血清保存的细胞中，即使在电缆开始生长后，蛋白质磷酸化和RNA合成的速率仍然很高。放线菌素D可使经马血清或胰岛素处理的细胞产生假足索，但可阻断经马血清处理的细胞形成针状体。马血清和囊胚腔液似乎含有除胰岛素以外的信号化合物。

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.