酪氨酸磷酸酶Shp2对于NMDA受体酪氨酸磷酸化和突触可塑性调控的机制研究
发布时间：2019-06-21 05:48
【摘要】：包含SH2结构域的酪氨酸磷酸酶2(Src homology 2 domain containing phosphotyrosine phosphatase 2,Shp2)是非受体酪氨酸磷酸酶的一种,在进化中高度保守。在患病人群中大约50%的Noonan综合征(Noonan syndrom,NS)是由Shp2的激活突变引起的,而大约80%的LEOPARD综合征(LEOPARD syndrom,LS)则是由它的失活突变引起。这两种疾病都是全身性的发育障碍病,并且均有一定比例的病人表现出智力障碍。虽然已有报道称Shp2能够通过调控下游Ras/ERKMAP激酶(MAPK)来参与记忆过程,但是内在机制依然不是很清楚。有报道称Shp2存在于N-甲基-D-门冬氨酸(N-methyl-D-aspartate,NMDA)受体复合物中,并且有它和NMDA受体的GluN2B亚基直接相互作用的证据。NMDA受体是一类兴奋性离子型谷氨酸受体,主要通透Ca2+。能够发挥功能的NMDA受体由2个组构性的GluN1亚基和两个调节性的GluN2或者GluN3亚基构成。它是配体和电压双门控的通道,能够响应突触前的谷氨酸释放和突触后的去极化,在突触可塑性中发挥了重要的作用。基于此,我们采用Shp2 FloxP/FloxP小鼠与CaMKⅡα-Cre小鼠杂交得到在前脑兴奋性神经元中特异敲除Shp2的小鼠(Shp2 Floxp/Floxp:CaMKⅡα-Cre +/-,CaSKO)来研究 Shp2 是否通过调节 NMDA 受体亚基磷酸化来调控突触可塑性乃至学习记忆功能。我们进行的蛋白检测表明CaSKO小鼠海马组织中Src激酶416位酪氨酸(Src Y416)磷酸化水平升高,这一位点代表了其酶活性,相应的NMDA受体的GluN2B亚基的1472位酪氨酸(GluN2B Y1472)和GluN2A亚基的1325位酪氨酸(GluN2A Y1325)的磷酸化水平也特异性的升高。这使得突触部位NMDA受体的电流增加并且CaSKO小鼠海马CA1的锥体神经元上兴奋性突触传递也增加。我们进一步的分析表明,这些在CaSKO小鼠突触上增加的NMDA受体是三异聚体GluN1/GluN2A/GluN2B,并且这些受体对于锌离子抑制的敏感性增强,导致在高频刺激时通过该受体的电量减少。在场电位长时程突触增强(long-term potentiation,LTP)的测试中,我们发现敲除Shp2会削弱海马CA1由100 Hz/s-高频刺激诱导的LTP,而这种LTP的削弱正是与突触部位的NMDA受体对于锌离子抑制的敏感性增加相关。此外,我们进一步发现CaSKO小鼠的长时程场景恐惧记忆被削弱,而场景恐惧记忆的形成和短时记忆则不受影响。对于场景恐惧记忆前后小鼠海马组织中这些蛋白的磷酸化检测发现WT小鼠在场景恐惧记忆检测后SrcY416的磷酸化升高,并且GluN2B Y1472的磷酸化也相应上升,而在CaSKO小鼠中这些位点已经升高的磷酸化水平并未出现活性依赖的升高。通过对CaSKO小鼠进行Src激酶抑制剂PP2的处理,我们发现能够反转Shp2缺失所引起的一系列后果。据此,我们推断是Shp2敲除引起的Src激酶活性增高导致了 NMDA受体亚基磷酸化水平的升高,从而使得更多的NMDA受体在突触部位表达,且功能性突触数目和其强度也增多。这种饱和作用导致了 LTP的下降,并进一步影响长时程场景恐惧记忆。这为我们理解酪氨酸磷酸酶在突触可塑性调控中的作用以及NS和LS中智力障碍的表型提供了新的思路。
[Abstract]:The tyrosine phosphatase 2 (SH2), which contains the SH2 domain, is one of the non-receptor tyrosine phosphatases and is highly conserved in the evolution. Noonan syndrome (NS), which is about 50% of the affected population, is caused by the activation mutation of Shp2, while about 80% of the LEOPARD syndrom (LS) is caused by its inactivation mutation. Both of these diseases are systemic developmental disorders and there are a proportion of patients exhibiting mental disorders. Although it has been reported that Shp2 is able to participate in the memory process by regulating downstream Ras/ ERKMAP kinase (MAPK), the intrinsic mechanism remains unclear. It is reported that Shp2 is present in the N-methyl-D-aspartate (NMDA) receptor complex and has evidence of its direct interaction with the GluN2B subunit of the NMDA receptor. The NMDA receptor is an excitable ionic glutamate receptor, which is mainly permeable to Ca2 +. The NMDA receptor capable of functioning is composed of 2 fabric GluN1 subunits and two regulatory GluN2 or GluN3 subunits. It is a double-gated channel of ligand and voltage, which can respond to the glutamate release and postsynaptic depolarization before the synapse, and play an important role in the synaptic plasticity. Based on this, we used the SH2 FloxP/ FloP mice to cross with CaMK 鈪，
本文编号：2503839
[Abstract]:The tyrosine phosphatase 2 (SH2), which contains the SH2 domain, is one of the non-receptor tyrosine phosphatases and is highly conserved in the evolution. Noonan syndrome (NS), which is about 50% of the affected population, is caused by the activation mutation of Shp2, while about 80% of the LEOPARD syndrom (LS) is caused by its inactivation mutation. Both of these diseases are systemic developmental disorders and there are a proportion of patients exhibiting mental disorders. Although it has been reported that Shp2 is able to participate in the memory process by regulating downstream Ras/ ERKMAP kinase (MAPK), the intrinsic mechanism remains unclear. It is reported that Shp2 is present in the N-methyl-D-aspartate (NMDA) receptor complex and has evidence of its direct interaction with the GluN2B subunit of the NMDA receptor. The NMDA receptor is an excitable ionic glutamate receptor, which is mainly permeable to Ca2 +. The NMDA receptor capable of functioning is composed of 2 fabric GluN1 subunits and two regulatory GluN2 or GluN3 subunits. It is a double-gated channel of ligand and voltage, which can respond to the glutamate release and postsynaptic depolarization before the synapse, and play an important role in the synaptic plasticity. Based on this, we used the SH2 FloxP/ FloP mice to cross with CaMK 鈪，
本文编号：2503839
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