Queipo Mª José, Gil-Redondo Juan C, Morente Verónica, Ortega Felipe, Miras-Portugal Mª Teresa, Delicado Esmerilda G, Pérez-Sen Raquel
Departamento de Bioquímica y Biología Molecular IV, Facultad de Veterinaria, Instituto Universitario de Investigación en Neuroquímica, Instituto de Investigación Sanitaria del Hospital Clínico San Carlos, Universidad Complutense Madrid, Madrid, Spain.
Front Mol Neurosci. 2018 Jan 10;10:448. doi: 10.3389/fnmol.2017.00448. eCollection 2017.
Extracellular signal-regulated kinases 1 and 2 (ERK1/2) play a central role in the intracellular signaling of P2X7 nucleotide receptors in neurons and glial cells. Fine spatio-temporal tuning of mitogen-activated protein (MAP) kinases is essential to regulate their biological activity. MAP kinase phosphatases (MKPs) are dual specificity protein phosphatases (DUSPs) that dephosphorylate phosphothreonine and phosphotyrosine residues in MAP kinases. This study focuses on how DUSP, DUSP6/MKP3, a phosphatase specific for ERK1/2 is regulated by the P2X7 nucleotide receptor in cerebellar granule neurons and astrocytes. Stimulation with the specific P2X7 agonist, BzATP, or epidermal growth factor (EGF) (positive control for ERK activation) regulates the levels of DUSP6 in a time dependent manner. Both agonists promote a decline in DUSP6 protein, reaching minimal levels after 30 min yet recovering to basal levels after 1 h. The initial loss of protein occurs through proteasomal degradation, as confirmed in experiments with the proteasome inhibitor, MG-132. Studies carried out with Actinomycin D demonstrated that the enhanced transcription of the gene is responsible for recovering the DUSP6 protein levels. Interestingly, ERK1/2 proteins are involved in the biphasic regulation of the protein phosphatase, being required for both the degradation and the recovery phase. We show that direct Ser phosphorylation of DUSP6 by ERK1/2 proteins could be part of the mechanism regulating their cytosolic levels, at least in glial cells. Thus, the ERK1/2 activated by P2X7 receptors exerts positive feedback on these kinase's own activity, promoting the degradation of one of their major inactivators in the cytosolic compartment, DUSP6, both in granule neurons and astrocytes. This feedback loop seems to function as a common universal mechanism to regulate ERK signaling in neural and non-neural cells.
细胞外信号调节激酶1和2（ERK1/2）在神经元和神经胶质细胞中P2X7核苷酸受体的细胞内信号传导中起核心作用。丝裂原活化蛋白（MAP）激酶的精细时空调节对于调节其生物学活性至关重要。MAP激酶磷酸酶（MKPs）是双特异性蛋白磷酸酶（DUSPs），可使MAP激酶中的磷酸苏氨酸和磷酸酪氨酸残基去磷酸化。本研究聚焦于特异性作用于ERK1/2的磷酸酶DUSP6/MKP3如何在小脑颗粒神经元和星形胶质细胞中受P2X7核苷酸受体调控。用特异性P2X7激动剂BzATP或表皮生长因子（EGF）（ERK激活的阳性对照）刺激可随时间依赖性方式调节DUSP6的水平。两种激动剂均促使DUSP6蛋白水平下降，30分钟后降至最低水平，但1小时后恢复至基础水平。蛋白的初始丢失是通过蛋白酶体降解发生的，蛋白酶体抑制剂MG-132的实验证实了这一点。用放线菌素D进行的研究表明，该基因转录增强负责DUSP6蛋白水平的恢复。有趣的是，ERK1/2蛋白参与了该蛋白磷酸酶的双相调节，在降解和恢复阶段均有需求。我们表明，ERK1/2蛋白对DUSP6的直接丝氨酸磷酸化可能是调节其胞质水平机制的一部分，至少在神经胶质细胞中如此。因此，由P2X7受体激活的ERK1/2对这些激酶自身的活性施加正反馈，促进其在胞质区室中的主要失活剂之一DUSP6在颗粒神经元和星形胶质细胞中的降解。这种反馈回路似乎作为一种常见的普遍机制来调节神经和非神经细胞中的ERK信号传导。
