KCNQ/M(Kv7)钾通道的转录抑制调控及其在大鼠骨癌痛中的作用研究

Our previous studies have documented that the down-regulated KCNQ/M channels in cancer rat DRG neurons contributed to the hyperexcitability of the nociceptive DRG neurons and to the cancer-induced bone pain. It is well accepted that the transcription factor REST functions as a repressor for the regulation of neural-specific gene expression. In our pilot study, we have found that in cancer rat DRGs, the mRNA levels of KCNQ2 and KCNQ3 decreased significantly, whereas the expression of REST mRNA and protein increased dramatically, indicating that the up-regulated REST likely resulted in the bone cancer pain via the transcription repression to KCNQ genes expression. In a recent study, the authors have found that IGF-I can elevate the REST expression. Our preliminary experimental results also showed that the expression of IGF-I and its receptor IGF-IR increased markedly in cancer rat DRGs, and the elevated IGF-I likely caused the bone cancer pain through its receptor IGF-IR. On the basis of these findings, we plan to investigate the mechanisms underlying the down-regulated KCNQ/M channels in cancer rat DRG neurons as well as their roles in the bone cancer pain. We will carry out the following studies to validate: (1) the transcriptional repression of KCNQ genes by REST and its contribution to the development of bone cancer pain, and the mechanisms underlying the transcriptional repression of KCNQ genes by REST; (2) Up-regulation of REST by IGF-I and its potential signaling pathway(s); (3) roles of the transcriptional repression of KCNQ genes by IGF-I-upregulated REST in the development of peripheral sensitization and the bone cancer pain. This study will be beneficial to elucidate the pathogenic mechanisms of bone cancer pain, and provide new target for the treatment of bone cancer pain.

我们前期的研究已经证实,KCNQ/M钾通道在DRG神经元的下调可以引起神经元的超兴奋性和骨癌痛。研究表明,转录抑制因子REST对许多神经元的基因表达起着重要的负性调控作用。我们的研究发现,在骨癌大鼠的DRG中,KCNQ2和KCNQ3的mRNA水平显著降低而REST的表达却显著增加,提示上调的REST可能通过对KCNQ 的转录抑制而引起骨癌痛。有研究报道IGF-I可以上调REST的表达。我们的研究也发现IGF-I在骨癌大鼠的DRG中升高且与骨癌痛的发生有关。本项目拟通过以下研究来进一步明确KCNQ/M通道在骨癌大鼠下调的分子机制及其在骨癌痛中的作用：1)REST对KCNQ转录抑制的分子机制及其在骨癌痛中的作用；2)IGF-I对REST的上调作用及其信号通路；3)由IGF-I上调REST所介导的KCNQ转录抑制在骨癌痛中的作用。该研究将有助于阐明骨癌痛的发病机理,为骨癌痛的药物治疗提供新靶点。

本项目主要从表观遗传学的角度分以下部分研究了KCNQ/M钾通道的转录抑制调控及其在大鼠骨癌痛中的作用与机制：1)组蛋白去乙酰化对KCNQ转录抑制的分子机制及其在骨癌痛中的作用；2) EGF对HDAC2的上调作用及其信号通路；3)由EGF上调HDAC2所介导的KCNQ转录抑制在骨癌痛中的作用。主要研究结果如下：1)在骨癌大鼠的DRG中,KCNQ2和KCNQ3的蛋白表达和电流密度下调、mRNA水平降低；相反，HDAC2、EGF和EGF受体EGFR的蛋白表达显著增加。 2)在骨癌大鼠DRG中,HDAC2与kcnq2和kcnq3基因启动子区域的结合能力增加。上调的HDAC2可以通过对kcnq基因启动子区域的组蛋白去乙酰化而发挥其对KCNQ的转录抑制作用，进而导致KCNQ的表达减少和骨癌痛的发生。3)在Kcnq2基因启动子区域存在2个可以与MeCP2结合的位点，其中位点2可以与MeCP2结合而位点1不能与MeCP2结合。在Kcnq3基因启动子区域存在1个可以与MeCP2结合的位点。在癌症大鼠DRG中，MeCP2与Kcnq2基因启动子区位点2、以及与Kcnq3基因启动子区的结合能力都显著增加。4)在骨癌大鼠DRG中，MeCP2可以与HDAC2形成复合体结合在kcnq2/3基因启动子区上，进而导致Kcnq2/3基因启动子区的组蛋白乙酰化降低，并抑制kcnq基因的转录。5)在骨癌大鼠DRG神经元中上调的的EGF及其受体EGFR可能通过HDAC2实现对KCNQ的表观遗传学调控，即通过HDAC2对kcnq基因启动子区域的组蛋白去乙酰化而发挥其对KCNQ 的转录抑制作用，进而导致KCNQ的表达减少和骨癌痛的发生。本课题首次从表观遗传学的角度探究了KCNQ/M钾通道的转录调控机制及其在大鼠骨癌痛发生中的作用，为阐明癌症痛的发病机理和临床治疗提供了新的理论切入点。