Hinssen H, Vandekerckhove J, Lazarides E
Division of Biology, California Institute of Technology, Pasadena 91125.
J Cell Biol. 1987 Sep;105(3):1425-33. doi: 10.1083/jcb.105.3.1425.
We have identified an approximately 85-kD protein in chicken erythrocytes which is immunologically, structurally, and functionally related to the gelsolin found in many muscle and nonmuscle cell types. Cell fractionation reveals a Ca2+-dependent partitioning of gelsolin into the soluble cytoplasm and the membrane-associated cytoskeleton of differentiating or mature erythrocytes. Depending on either the presence of Ca2+ during cell lysis or on the preincubation of the intact cells with the Ca2+-ionophore A23187, up to 40% of the total cellular gelsolin is found associated with the membrane skeleton. Expression of gelsolin shows a strong negative regulation during erythroid differentiation. From quantitations of its steady-state molar ratio to actin, gelsolin is abundant in early progenitor cells as revealed from avian erythroblastosis virus- and S13 virus-transformed cells which are arrested at the colony forming unit erythroid (CFU-e) stage of erythroid development. In these cells, which have a rudimentary and unstable membrane skeleton, gelsolin remains quantitatively cytoplasmic, irrespective of the Ca2+ concentration. During chicken embryo development and maturation, the expression of gelsolin decreases by a factor of approximately 10(3) in erythroid cells. This down regulation is independent from that of actin, which is considerably less, and is observed also when S13-transformed erythroid progenitor cells are induced to differentiate under conditions where the actin content of these cells does not change. In mature erythrocytes of the adult the amount of gelsolin is low, and significantly less than required for potentially capping of all membrane-associated actin filaments. We suggest that the gelsolin in erythroid cells is involved in the assembly of the actin filaments present in the membrane skeleton, and that it may provide for a mechanism, by means of its severing action on actin filaments, to extend the meshwork of the spectrin-actin-based membrane skeleton in erythroid cells during erythropoiesis.
我们在鸡红细胞中鉴定出一种约85-kD的蛋白质,它在免疫、结构和功能上与许多肌肉和非肌肉细胞类型中发现的凝溶胶蛋白相关。细胞分级分离显示,凝溶胶蛋白在分化或成熟红细胞的可溶性细胞质和膜相关细胞骨架中存在Ca2+依赖性分配。根据细胞裂解过程中Ca2+的存在情况或完整细胞与Ca2+离子载体A23187的预孵育情况,高达40%的总细胞凝溶胶蛋白与膜骨架相关。凝溶胶蛋白的表达在红细胞分化过程中表现出强烈的负调控。从其与肌动蛋白的稳态摩尔比定量分析可知,在禽成红细胞增多症病毒和S13病毒转化的细胞中,凝溶胶蛋白在早期祖细胞中含量丰富,这些细胞停滞在红细胞发育的集落形成单位红细胞(CFU-e)阶段。在这些具有原始且不稳定膜骨架的细胞中,无论Ca2+浓度如何,凝溶胶蛋白在数量上仍存在于细胞质中。在鸡胚胎发育和成熟过程中,红细胞中凝溶胶蛋白的表达降低了约10(3)倍。这种下调与肌动蛋白的下调无关,肌动蛋白的下调幅度要小得多,并且在S13转化的红细胞祖细胞在其肌动蛋白含量不变的条件下被诱导分化时也能观察到。在成年鸡的成熟红细胞中,凝溶胶蛋白的含量很低,明显低于潜在封闭所有膜相关肌动蛋白丝所需的量。我们认为,红细胞中的凝溶胶蛋白参与了膜骨架中肌动蛋白丝的组装,并且它可能通过对肌动蛋白丝的切断作用,为红细胞生成过程中扩展基于血影蛋白-肌动蛋白的膜骨架网络提供一种机制。