Yao R, Osada H
Laboratory of Antibiotics, The Institute of Physical and Chemical Research (RIKEN), Wako, Saitama, Japan.
Exp Cell Res. 1997 Aug 1;234(2):233-9. doi: 10.1006/excr.1997.3615.
Rat pheochromocytoma cells, PC12 cells, undergo differentiation in response to nerve growth factor (NGF). Although the Ras-MAP kinase signaling pathway has been shown to play a central role in the response to NGF, the precise mechanism which induces differentiation remains unclarified. Recently, several gamma-lactam-related microbial products were identified to induce neurite outgrowth in neuroblastoma cells. Therefore, we synthesized a series of gamma-lactam-related compounds and tested for their ability to induce neurite outgrowth in PC12 cells. We found that two compounds, MT-19 and MT-20, induced neurite outgrowth at concentrations as low as 1 microg/ml. MT-19 and MT-20 have an n-hexadecyl group and an n-dodecyl group, respectively, at the position N-1 of the gamma-lactam ring, and the modification of this group leads to partial or complete loss of activity. In addition, the modification of the methyl and hydroxyl group at C-5 leads to complete loss of activity, indicating a strict structure-activity relationship. Interestingly, MT-19 and MT-20 induced neurite outgrowth of PC12 cells which lack normal Ras function. Furthermore, these compounds did not induce MAP kinase activation, suggesting that MT-19 and MT-20 do not require the Ras-MAP kinase signaling pathway which is shown to be necessary and sufficient for NGF-induced neurite outgrowth. Consistent with this, none of the early- or late-response genes tested, which include fos, zif268, Nur77, vgf, and transin, was induced. However, the protein level of three neurofilaments was increased after the incubation with these compounds. Since the level of other cytoskeleton proteins including actin and tubulin remained constant, MT-19 and MT-20 specifically affected neurofilament synthesis and/or turnover. Taken together, these findings indicate that MT-19 and MT-20 induce neurite outgrowth by activating the downstream target of MAP kinase or by a novel mechanism which is distinct from the NGF-activated pathway.
大鼠嗜铬细胞瘤细胞（PC12细胞）会对神经生长因子（NGF）作出反应而发生分化。尽管已表明Ras - MAP激酶信号通路在对NGF的反应中起核心作用，但诱导分化的确切机制仍不清楚。最近，几种与γ-内酰胺相关的微生物产物被鉴定出可诱导神经母细胞瘤细胞长出神经突。因此，我们合成了一系列与γ-内酰胺相关的化合物，并测试了它们在PC12细胞中诱导神经突生长的能力。我们发现两种化合物MT - 19和MT - 20在低至1微克/毫升的浓度下就能诱导神经突生长。MT - 19和MT - 20在γ-内酰胺环的N - 1位分别有一个正十六烷基和一个正十二烷基，该基团的修饰会导致活性部分或完全丧失。此外，C - 5位甲基和羟基的修饰会导致活性完全丧失，这表明存在严格的构效关系。有趣的是，MT - 19和MT - 20可诱导缺乏正常Ras功能的PC12细胞长出神经突。此外，这些化合物不会诱导MAP激酶激活，这表明MT - 19和MT - 20不需要对NGF诱导神经突生长而言必要且充分的Ras - MAP激酶信号通路。与此一致的是，所测试的早期或晚期反应基因（包括fos、zif268、Nur77、vgf和转胶酶）均未被诱导。然而，与这些化合物孵育后，三种神经丝的蛋白水平升高。由于包括肌动蛋白和微管蛋白在内的其他细胞骨架蛋白水平保持不变，MT - 19和MT - 20特异性地影响神经丝的合成和/或周转。综上所述，这些发现表明MT - 19和MT - 20通过激活MAP激酶的下游靶点或通过一种不同于NGF激活途径的新机制来诱导神经突生长。
