The organization of F-actin and microtubules in growth cones exposed to a brain-derived collapsing factor.
The organization of F-actin and microtubules in growth cones exposed to a brain-derived collapsing factor.
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DOI：
10.1083/jcb.121.4.867
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发表时间：
1993-05
期刊：
影响因子：
--
通讯作者：
Raper JA
中科院分区：
文献类型：
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作者：
Fan J;Mansfield SG;Redmond T;Gordon-Weeks PR;Raper JA
在先前的工作中，我们对一种脑源性溃变因子进行了特征描述，该因子在培养过程中可诱导背根神经节生长锥的溃变（雷珀和卡普夫哈默，1990年）。为了确定生长锥细胞骨架在溃变过程中是如何重新排列的，我们比较了正常生长锥和部分溃变生长锥中F - 肌动蛋白和微管的分布。通过将F - 肌动蛋白的罗丹明 - 鬼笔环肽染色强度与一种通用蛋白质染色强度进行比值，可以测量生长锥中F - 肌动蛋白相对于所有蛋白质的相对浓度。在暴露于溃变因子5分钟的生长锥中，F - 肌动蛋白的相对浓度降低了约一半，此时生长锥刚刚开始溃变。在此期间，生长锥前沿的F - 肌动蛋白相对浓度急剧下降，而中心的F - 肌动蛋白浓度下降很少。这些结果表明，溃变与生长锥前沿F - 肌动蛋白的净损失有关。用抗α - 微管蛋白酪氨酸化和去酪氨酸化异构体的抗体检测了正常生长锥和经溃变因子处理的生长锥中微管的分布。酪氨酸化形式存在于新聚合的微管中，而去酪氨酸化形式则不存在。在溃变过程中，这些异构体的相对近 - 远端分布没有改变，这表明微管聚合和解聚的速率不受溃变因子存在的很大影响。对溃变前后微管分布的分析表明，微管发生了重新排列，但在溃变过程中其聚合状态未受影响。这些结果与以下假设一致：脑源性溃变因子对微管聚合或解聚几乎没有影响。相反，它似乎诱导了生长锥前沿F - 肌动蛋白的净损失。
In previous work we characterized a brain derived collapsing factor that induces the collapse of dorsal root ganglion growth cones in culture (Raper and Kapfhammer, 1990). To determine how the growth cone cytoskeleton is rearranged during collapse, we have compared the distributions of F-actin and microtubules in normal and partially collapsed growth cones. The relative concentration of F-actin as compared to all proteins can be measured in growth cones by rationing the intensity of rhodamine-phalloidin staining of F-actin to the intensity of a general protein stain. The relative concentration of F- actin is decreased by about one half in growth cones exposed to collapsing factor for five minutes, a time at which they are just beginning to collapse. During this period the relative concentration of F-actin in the leading edges of growth cones decreases dramatically while the concentration of F-actin in the centers decreases little. These results suggest that collapse is associated with a net loss of F- actin at the leading edge. The distributions of microtubules in normal and collapsing factor treated growth cones were examined with antibodies to tyrosinated and detyrosinated isoforms of alpha-tubulin. The tyrosinated form is found in newly polymerized microtubules while the detyrosinated form is not. The relative proximal-distal distributions of these isoforms are not altered during collapse, suggesting that rates of microtubule polymerization and depolymerization are not greatly affected by the presence of collapsing factor. An analysis of the distributions of microtubules before and after collapse suggests that microtubules are rearranged, but their polymerization state is unaffected during collapse. These results are consistent with the hypothesis that the brain derived collapsing factor has little effect on microtubule polymerization or depolymerization. Instead it appears to induce a net loss of F-actin at the leading edge of the growth cone.