Department of Biological Sciences and the Center for Neuroscience Research, University at Albany, State University of New York, Albany, NY 12222, USA.
J Neurosci. 2012 Mar 7;32(10):3563-74. doi: 10.1523/JNEUROSCI.5197-11.2012.
Axotomized optic axons of Xenopus laevis, in contrast to those of mammals, retain their ability to regenerate throughout life. To better understand the molecular basis for this successful regeneration, we focused on the role of an RNA-binding protein, heterogeneous nuclear ribonucleoprotein (hnRNP) K, because it is required for axonogenesis during development and because several of its RNA targets are under strong post-transcriptional control during regeneration. At 11 d after optic nerve crush, hnRNP K underwent significant translocation into the nucleus of retinal ganglion cells (RGCs), indicating that the protein became activated during regeneration. To suppress its expression, we intravitreously injected an antisense Vivo-Morpholino oligonucleotide targeting hnRNP K. In uninjured eyes, it efficiently knocked down hnRNP K expression in only the RGCs, without inducing either an axotomy response or axon degeneration. After optic nerve crush, staining for multiple markers of regenerating axons showed no regrowth of axons beyond the lesion site with hnRNP K knockdown. RGCs nonetheless responded to the injury by increasing expression of multiple growth-associated RNAs and experienced no additional neurodegeneration above that normally seen with optic nerve injury. At the molecular level, hnRNP K knockdown during regeneration inhibited protein, but not mRNA, expression of several known hnRNP K RNA targets (NF-M, GAP-43) by compromising their efficient nuclear transport and disrupting their loading onto polysomes for translation. Our study therefore provides evidence of a novel post-transcriptional regulatory pathway orchestrated by hnRNP K that is essential for successful CNS axon regeneration.
非洲爪蟾的轴突被切断后，与哺乳动物不同的是，它们在整个生命过程中都保持着再生的能力。为了更好地理解这种成功再生的分子基础，我们将重点放在 RNA 结合蛋白异质核核糖核蛋白（hnRNP）K 上，因为它在发育过程中轴突发生所必需的，并且其几个 RNA 靶标在再生过程中受到强烈的转录后调控。在视神经挤压后 11 天，hnRNP K 显著转位到视网膜神经节细胞（RGC）的核内，表明该蛋白在再生过程中被激活。为了抑制其表达，我们通过玻璃体腔内注射靶向 hnRNP K 的反义 Vivo-Morpholino 寡核苷酸来抑制其表达。在未受伤的眼睛中，它仅在 RGC 中有效地敲低了 hnRNP K 的表达，而不会诱导轴突切断反应或轴突退化。视神经挤压后，用多种再生轴突标志物染色显示，hnRNP K 敲低后，轴突没有在损伤部位以外的地方再生。然而，RGC 对损伤做出反应，增加了多种生长相关 RNA 的表达，并且没有超过视神经损伤通常观察到的额外神经退行性变。在分子水平上，hnRNP K 在再生过程中的敲低通过损害其有效的核运输并破坏其加载到多核糖体进行翻译来抑制几种已知的 hnRNP K RNA 靶标（NF-M、GAP-43）的蛋白质表达，但不抑制其 mRNA 表达。因此，我们的研究提供了证据表明 hnRNP K 协调了一种新的转录后调控途径，该途径对成功的中枢神经系统轴突再生是必不可少的。
