Inhibition of N1-Src kinase by a specific SH3 peptide ligand reveals a role for N1-Src in neurite elongation by L1-CAM.
Inhibition of N1-Src kinase by a specific SH3 peptide ligand reveals a role for N1-Src in neurite elongation by L1-CAM.
复制标题
DOI：
10.1038/srep43106
复制
发表时间：
2017-02-21
影响因子：
4.6
通讯作者：
Evans GJ
中科院分区：
文献类型：
--
作者：
Keenan S;Wetherill SJ;Ugbode CI;Chawla S;Brackenbury WJ;Evans GJ
在哺乳动物脑中，普遍存在的酪氨酸激酶C-Src经历剪接以在SH 3结构域中插入短序列以产生N1-Src和N2-Src。我们和其他人之前已经证明，与C-Src相比，N-Src改变了底物特异性和激酶活性。然而，N-Src的确切功能是未知的，并且很可能N-Src信号传导事件被错误地归因于C-Src，因为它们不能通过靶向激酶结构域的常规Src抑制剂来区分。通过筛选噬菌体肽库，我们发现了一个新的配体（PDN 1），其靶向N1-Src的独特SH 3结构域，并抑制细胞中的N1-Src。在培养的神经元中，融合到荧光蛋白的PDN 1抑制了神经突的生长，这种作用被靶向N1-Src微外显子的shRNA所模仿。PDN 1还抑制小脑颗粒神经元中L1-CAM依赖的神经突伸长，这是一种先前在Src−/−小鼠中被破坏的途径。因此，PDN 1代表了区分神经元中N1-Src和C-Src功能的新工具，并且是开发N1-Src小分子抑制剂的起点。
In the mammalian brain the ubiquitous tyrosine kinase, C-Src, undergoes splicing to insert short sequences in the SH3 domain to yield N1- and N2-Src. We and others have previously shown that the N-Srcs have altered substrate specificity and kinase activity compared to C-Src. However, the exact functions of the N-Srcs are unknown and it is likely that N-Src signalling events have been misattributed to C-Src because they cannot be distinguished by conventional Src inhibitors that target the kinase domain. By screening a peptide phage display library, we discovered a novel ligand (PDN1) that targets the unique SH3 domain of N1-Src and inhibits N1-Src in cells. In cultured neurons, PDN1 fused to a fluorescent protein inhibited neurite outgrowth, an effect that was mimicked by shRNA targeting the N1-Src microexon. PDN1 also inhibited L1-CAM-dependent neurite elongation in cerebellar granule neurons, a pathway previously shown to be disrupted in Src−/− mice. PDN1 therefore represents a novel tool for distinguishing the functions of N1-Src and C-Src in neurons and is a starting point for the development of a small molecule inhibitor of N1-Src.