Laboratory of Molecular Neurobiology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan.
Department of Anatomy, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan.
J Neurochem. 2019 Jan;148(2):204-218. doi: 10.1111/jnc.14596. Epub 2018 Nov 12.
The expression of immediate early genes (IEGs) is thought to be an essential molecular basis of neuronal plasticity for higher brain function. Many IEGs contain serum response element in their transcriptional regulatory regions and their expression is controlled by serum response factor (SRF). SRF is known to play a role in concert with transcriptional cofactors. However, little is known about how SRF cofactors regulate IEG expression during the process of neuronal plasticity. We hypothesized that one of the SRF-regulated neuronal IEGs, activity-regulated cytoskeleton-associated protein (Arc; also termed Arg3.1), is regulated by an SRF coactivator, megakaryoblastic leukemia (MKL). To test this hypothesis, we initially investigated which binding site of the transcription factor or SRF cofactor contributes to brain-derived neurotrophic factor (BDNF)-induced Arc gene transcription in cultured cortical neurons using transfection and reporter assays. We found that BDNF caused robust induction of Arc gene transcription through a cAMP response element, binding site of myocyte enhancer factor 2, and binding site of SRF in an Arc enhancer, the synaptic activity-responsive element (SARE). Regardless of the requirement for the SRF-binding site, the binding site of a ternary complex factor, another SRF cofactor, did not affect BDNF-mediated Arc gene transcription. In contrast, chromatin immunoprecipitation revealed occupation of MKL at the SARE. Furthermore, knockdown of MKL2, but not MKL1, significantly decreased BDNF-mediated activation of the SARE. Taken together, these findings suggest a novel mechanism by which MKL2 controls the Arc SARE in response to BDNF stimulation.
立即早期基因（IEGs）的表达被认为是高等大脑功能神经元可塑性的重要分子基础。许多 IEG 在其转录调控区域包含血清反应元件，其表达受血清反应因子（SRF）控制。已知 SRF 与转录共因子协同发挥作用。然而，关于 SRF 共因子如何在神经元可塑性过程中调节 IEG 的表达知之甚少。我们假设，SRF 调节的神经元 IEG 之一，活性调节细胞骨架相关蛋白（Arc；也称为 Arg3.1），受 SRF 共激活因子巨核细胞白血病（MKL）调节。为了验证这一假设，我们最初使用转染和报告基因分析，研究了转录因子或 SRF 共因子的哪个结合位点有助于培养的皮质神经元中脑源性神经营养因子（BDNF）诱导的 Arc 基因转录。我们发现，BDNF 通过 cAMP 反应元件、肌细胞增强因子 2 结合位点和 Arc 增强子中的 SRF 结合位点（突触活性反应元件（SARE）），引起 Arc 基因转录的强烈诱导。无论是否需要 SRF 结合位点，另一个 SRF 共因子三元复合物因子的结合位点都不会影响 BDNF 介导的 Arc 基因转录。相比之下，染色质免疫沉淀显示 MKL 占据了 SARE。此外，MKL2 的敲低，而不是 MKL1 的敲低，显著降低了 BDNF 介导的 SARE 激活。总之，这些发现表明了一种新的机制，即 MKL2 控制 MKL2 对 BDNF 刺激的 Arc SARE 的反应。
