Department of Pharmacology & Experimental Therapeutics, Laboratory of Translational Epilepsy, Boston University School of Medicine, Boston, MA 02118, USA.
BMC Pharmacol Toxicol. 2013 Jul 23;14:37. doi: 10.1186/2050-6511-14-37.
The γ-aminobutyric acid (GABA) type A receptor (GABA(A)R) contains the recognition sites for a variety of agents used in the treatment of brain disorders, including anxiety and epilepsy. A better understanding of how receptor expression is regulated in individual neurons may provide novel opportunities for therapeutic intervention. Towards this goal we have studied transcription of a GABA(A)R subunit gene (GABRB1) whose activity is autologously regulated by GABA via a 10 base pair initiator-like element (β(1)-INR).
By screening a human cDNA brain library with a yeast one-hybrid assay, the Polycomblike (PCL) gene product PHD finger protein transcript b (PHF1b) was identified as a β(1)-INR associated protein. Promoter/reporter assays in primary rat cortical cells demonstrate that PHF1b is an activator at GABRB1, and chromatin immunoprecipitation assays reveal that presence of PHF1 at endogenous Gabrb1 is regulated by GABA(A)R activation.
PCL is a member of the Polycomb group required for correct spatial expression of homeotic genes in Drosophila. We now show that PHF1b recognition of β(1)-INR is dependent on a plant homeodomain, an adjacent helix-loop-helix, and short glycine rich motif. In neurons, it co-immunoprecipitates with SUZ12, a key component of the Polycomb Repressive Complex 2 (PRC2) that regulates a number of important cellular processes, including gene silencing via histone H3 lysine 27 trimethylation (H3K27me3).
The observation that chronic exposure to GABA reduces PHF1 binding and H3K27 monomethylation, which is associated with transcriptional activation, strongly suggests that PHF1b may be a molecular transducer of GABA(A)R function and thus GABA-mediated neurotransmission in the central nervous system.
γ-氨基丁酸（GABA）A型受体（GABA(A)R）包含用于治疗大脑疾病的各种药物的识别位点，包括焦虑和癫痫。更好地了解个体神经元中受体表达是如何调节的，可能为治疗干预提供新的机会。为此，我们研究了 GABA(A)R 亚基基因（GABRB1）的转录，该基因的活性通过 10 个碱基对起始样元件（β(1)-INR）被 GABA 自动调节。
通过酵母单杂交分析筛选人 cDNA 脑文库，鉴定出多梳样（PCL）基因产物 PHF1b 是 β(1)-INR 相关蛋白。在原代大鼠皮质细胞中的启动子/报告基因检测表明，PHF1b 是 GABRB1 的激活剂，染色质免疫沉淀检测显示 PHF1 在内源性 Gabrb1 中的存在受 GABA(A)R 激活的调节。
PCL 是多梳组的成员，对于果蝇同源基因的正确空间表达是必需的。我们现在表明，PHF1b 对 β(1)-INR 的识别依赖于植物同源域、相邻的螺旋-环-螺旋和短甘氨酸丰富的基序。在神经元中，它与 SUZ12 共免疫沉淀，SUZ12 是多梳抑制复合物 2（PRC2）的关键组成部分，调节许多重要的细胞过程，包括通过组蛋白 H3 赖氨酸 27 三甲基化（H3K27me3）进行基因沉默。
慢性暴露于 GABA 会降低 PHF1 结合和 H3K27 单甲基化的观察结果，这与转录激活有关，强烈表明 PHF1b 可能是 GABA(A)R 功能的分子转导者，因此也是中枢神经系统中 GABA 介导的神经传递的分子转导者。
