帕金森病相关基因α-突触核蛋白和PINK1对Nurr1的调节作用

Regulatory effects of α-synuclein and PINK1 on Nurr1 activity

目的探讨帕金森病相关基因α-突触核蛋白(α-synuclein,α-syn)和PTEN诱导的蛋白激酶1(PTEN induced putative kinase 1,PINK1)对转录因子Nurr1的调节作用。方法利用荧光共振能量转移(fluorescence resonance energy transfer,FRET)的方法检测α-syn与Nurr1的相互作用。同时,在PINK1基因敲减的细胞模型中,通过双荧光素酶报告系统检测PINK1对Nurr1转录激活活性的调节作用。结果 FRET检测显示,CFP-α-syn与YFP-Nurr1之间发生了荧光共振能量转移的现象。其FRET效率大约为20.52%。提示α-syn与Nurr1之间可能存在直接的相互作用。对于PINK1基因敲减的细胞进行研究发现,在PINK1被抑制表达24 h或48 h后,Nurr1的活性显著下调。结论以往研究提示α-syn可能是Nurr1的抑制因子。研究显示α-syn可能与Nurr1直接结合而抑制其活性。而PINK1对Nurr1可能存在一个正性调节作用。因此,二者对Nurr1的调节可能存在一个相互协调、相互制约的关系。

Objective To investigate if α-synuclein（ α-syn） and PTEN induced putative kinase 1（ PINK1） have an effect on nuclear receptor related 1（ Nurr1） activity. Methods Two plasmids p Am Cyan1-N1-α-syn（ CFP-α-syn） and p Zs Yellow1-N1-Nurr1（ YFP-Nurr1）were constructed to detect the interaction between α-syn and Nurr1 using fluorescence resonance energy transfer（ FRET） technique.PINK1 gene was silenced with siRNA in MN9 D cells followed by Nurr1 activity detection. Results After transfected with the above two plasmids,SK-N-SH cells were observed under the confocal microscopy. The fluorescence of CFP and YFP did not become weak with fusion of target protein. FRET phenomenon was observed and the FRET efficiency between CFP-α-syn and YFP-Nurr1 was about 21%,indicating that there was an interaction between α-syn and Nurr1. The results showed that Nurr1 activity was down-regulated when PINK1 gene was knocked-down for 24 h or 48 h,indicating that PINK1 had a regulatory effect on Nurr1 activity. Conclusion Therefore α-syn may be as a transcriptional repressor to inhibit Nurr1 activity by interacting with the latter. In contrary,PINK1 positively regulates Nurr1 activity. The coordination between PINK1 and α-syn may play an important role in DA homeostasis.