Lewis A K, Bridgman P C
Department of Anatomy and Neurobiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Cell Motil Cytoskeleton. 1996;33(2):130-50. doi: 10.1002/(SICI)1097-0169(1996)33:2<130::AID-CM5>3.0.CO;2-G.
To determine if unconventional myosins play a role in nerve outgrowth, antibodies specific for rat brain derived mammalian myosin I alpha (MMI alpha) were used to label cultured rat superior cervical ganglion nerve cells. Observations were made at both the light and electron microscopic level of resolution using preparative procedures designed to enhance the ability to precisely determine the relationship between antibody label and cellular structures in order to map the distribution and structural association of this myosin. Immunofluorescence showed that MMI alpha has a punctate distribution throughout the nerve cell body, neurites, and growth cones. In growth cones, MMI alpha staining is sometimes elevated in thin peripheral regions of high actin content at the leading edge. Immunoelectron microscopy using colloidal gold conjugated antibodies showed that in growth cones MMI alpha is absent from membranous organelles and is concentrated primarily in the cell cortex adjacent to the cell membrane. The cortical label is equally distributed between upper and lower membranes. The plasma membrane association of the MMI alpha label persists under conditions in which the actin cytoskeleton is perturbed or removed, suggesting a direct association between a fraction of MMI alpha and the plasma membrane. MMI alpha label is also associated with the non-cortical actin cytoskeleton. Partial disruption of the actin cytoskeleton using cytochalasin B causes redistribution of only a subset of MMI alpha label. These data suggest a complex relationship between MMI alpha, the actin cytoskeleton, and the plasma membrane in the growth cone. In contrast to its localization in the growth cone, in neuronal cell bodies MMI alpha is also associated with tubulovesicular structures. This suggests that at this location MMI alpha may either act as an organelle motor or is passively transported to the plasma membrane on vesicles.
为了确定非常规肌球蛋白是否在神经生长中发挥作用，使用针对大鼠脑源性哺乳动物肌球蛋白Iα（MMIα）的特异性抗体对培养的大鼠颈上神经节神经细胞进行标记。使用旨在增强精确确定抗体标记与细胞结构之间关系的能力的制备程序，在光学和电子显微镜分辨率水平上进行观察，以绘制这种肌球蛋白的分布和结构关联图谱。免疫荧光显示，MMIα在整个神经细胞体、神经突和生长锥中呈点状分布。在生长锥中，MMIα染色有时在前沿高肌动蛋白含量的薄周边区域升高。使用胶体金偶联抗体的免疫电子显微镜显示，在生长锥中，MMIα不存在于膜性细胞器中，主要集中在与细胞膜相邻的细胞皮质中。皮质标记在上膜和下膜之间均匀分布。在肌动蛋白细胞骨架受到干扰或去除的条件下，MMIα标记与质膜的关联仍然存在，这表明一部分MMIα与质膜之间存在直接关联。MMIα标记也与非皮质肌动蛋白细胞骨架相关。用细胞松弛素B部分破坏肌动蛋白细胞骨架仅导致MMIα标记的一个子集重新分布。这些数据表明在生长锥中MMIα、肌动蛋白细胞骨架和质膜之间存在复杂的关系。与其在生长锥中的定位相反，在神经元细胞体中MMIα也与微管泡状结构相关。这表明在这个位置，MMIα可能要么作为细胞器马达起作用，要么被动地通过囊泡运输到质膜。
