†Glycochemistry & Glycobiology Lab, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
*School of Life Sciences, Northeast Normal University, Changchun 130024, China.
Biochem J. 2014 Apr 15;459(2):405-15. doi: 10.1042/BJ20131360.
Neurodevelopment is orchestrated by a series of growth factor-HS (heparan sulfate) interactions which are involved in neuritogenesis. GLCE (glucuronic acid epimerase) is a critical enzyme involved in HS synthesis, which converts GlcA (D-glucuronic acid) into IdoA (L-iduronic acid). However, the function of GLCE in neuritogenesis is largely unknown. In the present study we showed that GLCE depletion caused arrested PC12 cell growth and promoted the cell neuritogenesis and differentiation induced by NGF (nerve growth factor). PC12 cell growth was boosted by overexpression of GLCE, and neuritogenesis was impaired when GLCE depletion was rescued. Interestingly, overexpression of wild-type GLCE with Y168A and Y222A mutations led to enhanced PC12 cell growth and attenuated the neuritogenesis triggered by GLCE silencing. We showed further that GLCE depletion blocked SMAD1/5/8 phosphorylation; however, this signalling could be restored by GLCE or the mutation of its active enzymatic site. In addition, the downstream effector of SMAD1/5/8, ID3 (inhibitor of DNA binding/differentiation 3) was induced by GLCE. ID3 silencing inhibited PC12 cell growth and induced cell neuritogenesis and differentiation. In addition, ectopic expression of ID3 partially rescued the phenotype caused by GLCE silencing. The results of the present study suggest that GLCE plays a key role in PC12 cell growth and neuritogenesis through SMAD/ID3 signalling.
神经发育是由一系列生长因子-HS（硫酸乙酰肝素）相互作用来协调的，这些相互作用参与了神经突生成。GLCE（葡萄糖醛酸差向异构酶）是参与 HS 合成的关键酶，它将 GlcA（D-葡萄糖醛酸）转化为 IdoA（L-艾杜糖醛酸）。然而，GLCE 在神经突生成中的功能在很大程度上是未知的。在本研究中，我们表明 GLCE 耗竭导致 PC12 细胞生长停滞，并促进了 NGF（神经生长因子）诱导的细胞神经突生成和分化。GLCE 的过表达促进了 PC12 细胞的生长，而 GLCE 耗竭的挽救则损害了神经突生成。有趣的是，野生型 GLCE 的 Y168A 和 Y222A 突变体的过表达导致了 PC12 细胞生长的增强，并减弱了 GLCE 沉默触发的神经突生成。我们进一步表明，GLCE 耗竭阻断了 SMAD1/5/8 的磷酸化；然而，这种信号可以通过 GLCE 或其活性酶位点的突变来恢复。此外，SMAD1/5/8 的下游效应物 ID3（DNA 结合/分化抑制因子 3）被 GLCE 诱导。ID3 的沉默抑制了 PC12 细胞的生长，并诱导了细胞的神经突生成和分化。此外，ID3 的异位表达部分挽救了 GLCE 沉默引起的表型。本研究的结果表明，GLCE 通过 SMAD/ID3 信号通路在 PC12 细胞的生长和神经突生成中发挥关键作用。
