Nameroff M, Trotter J A, Keller J M, Munar E
J Cell Biol. 1973 Jul;58(1):107-18. doi: 10.1083/jcb.58.1.107.
In cell culture, a partially purified commercial preparation of phospholipase C (PLC) from Clostridium welchii inhibited fusion of myoblasts at concentrations of 12-50 microg per ml. At lower concentrations, PLC-treated cultures were indistinguishable from controls, and at concentrations above 100 microg per ml, PLC-treated cells detached from their substrates. The effect was reversible and fusion resumed approximately one cell cycle time after removal of the enzyme. Neither the percent of cells in the mitotic cycle nor the duration of the different phases of the cycle were altered by PLC at concentrations which inhibited fusion. Cell motility was not reduced by the enzyme. Unfused, PLC-treated myoblasts were virtually indistinguishable in ultrastructure from untreated cells just before fusion. In the presence of PLC, mononucleated myogenic cells did not synthesize thick (150 A) filaments. Treatment of culture medium with insolubilized commercial PLC did not abolish the capacity of the medium to support myogenesis. Chondrocytes treated with PLC divided repeatedly but failed to synthesize metachromatic matrix and failed to incorporate labeled sulfate into chondroitin sulfate. PLC was further purified by chromatography on Sephadex G-100. The resulting preparation was free of detectable protease, yielded one band on SDS-acrylamide gel electrophoresis, and displayed all of the biological activities of the less pure material.
在细胞培养中，来自产气荚膜梭菌的部分纯化的商业磷脂酶C（PLC）制剂在浓度为每毫升12 - 50微克时抑制成肌细胞融合。在较低浓度下，经PLC处理的培养物与对照无差异，而在每毫升高于100微克的浓度下，经PLC处理的细胞从其底物上脱离。该作用是可逆的，去除酶后约一个细胞周期时间融合恢复。在抑制融合的浓度下，PLC对有丝分裂周期中的细胞百分比或周期不同阶段的持续时间均无改变。该酶未降低细胞运动性。未融合的、经PLC处理的成肌细胞在超微结构上与即将融合前未处理的细胞几乎没有区别。在有PLC存在的情况下，单核生肌细胞不合成粗（150埃）丝。用不溶性商业PLC处理培养基并未消除其支持肌生成的能力。经PLC处理的软骨细胞反复分裂，但未能合成异染性基质，也未能将标记的硫酸盐掺入硫酸软骨素中。通过在Sephadex G - 100上进行层析进一步纯化PLC。所得制剂不含可检测到的蛋白酶，在SDS - 丙烯酰胺凝胶电泳上产生一条带，并表现出较不纯物质的所有生物学活性。
