ABIN-1 Negatively Regulates m-Opioid Receptor Function
ABIN-1 Negatively Regulates m-Opioid Receptor Function
复制标题
ABIN-1 负向调节 m-阿片受体功能
作者：
周培岚;江洁冰;颜慧;苏瑞斌
M-阿片受体(MOR)是一种Gi/o蛋白偶联受体，介导镇痛、兴奋和奖赏效应。通过细菌双杂交筛选，我们报道了大鼠MOR的羧基尾巴与核因子kB的20结合抑制物(ABIN-1)有关。细胞裂解物中MOR和ABIN-1蛋白的直接蛋白-蛋白结合和免疫共沉淀证实了这种相互作用。饱和结合研究表明，ABIN-1对MOR结合没有影响。但ABIN-1和MOR的相互作用可抑制DAMGO([D-Ala2，N-Me-Phe4，Gly5-ol]-Enkephalin)诱导的G蛋白的激活。在共表达MOR和ABIN-1的中国仓鼠卵巢细胞中，DAMGO诱导的MOR磷酸化、泛素化和内化减少。ABIN-1与MOR的相互作用也抑制了DAMGO对Forsklin刺激的腺苷酸环化酶的抑制作用。此外，细胞外信号调节的激酶激活也受到ABIN-1过表达的负调控。这些数据表明，ABIN-1在体外是MOR激活、磷酸化和内化的负协同调节因子。Abin-1还抑制吗啡诱导的斑马鱼幼体(AB系)的超速运动。利用反义吗啉寡核苷酸(MO)基因敲除技术，注射了ABIN-1的斑马鱼幼体在急性吗啡处理后，其活动距离显著增加(约60%)(P<0.01)。综上所述，ABIN-1在体内外对MOR功能具有负性调节作用
The m-opioid receptor (MOR) is a Gi/o protein-coupled receptor.that mediates analgesic, euphoric, and reward effects. Using a.bacterial two-hybrid screen, we reported that the carboxyl tail of.the rat MOR associates with A20-binding inhibitor of nuclear.factor kB (ABIN-1). This interaction was confirmed by direct.protein-protein binding and coimmunoprecipitation of MOR and.ABIN-1 proteins in cell lysates. Saturation binding studies showed.that ABIN-1 had no effect on MOR binding. However, the interaction.of ABIN-1 and MOR inhibited the activation of G proteins induced.by DAMGO ([D-Ala2,N-Me-Phe4,Gly5-ol]-Enkephalin). MOR phos-.phorylation, ubiquitination, and internalization induced by DAMGO.were decreased in Chinese hamster ovary cells that coexpressed.MOR and ABIN-1. The suppression of forskolin-stimulated adenylyl..cyclase by DAMGO was also inhibited by the interaction of ABIN-1.with MOR. In addition, extracellular signal-regulated kinase activa-.tion was also negatively regulated by overexpression of ABIN-1..These data suggest that ABIN-1 is a negative coregulator of MOR.activation, phosphorylation, and internalization in vitro. ABIN-1 also.inhibited morphine-induced hyperlocomotion in zebrafish larvae.(AB strain). By utilization of an antisense morpholino oligo-.nucleotide (MO) gene knockdown technology, the ABIN-1.MO-injected zebrafish larvae showed a significant increase.(approximately 60%) in distance moved compared with control.MO-injected larvae after acute morphine treatment (P ,0.01)..Taken together, ABIN-1 negatively regulates MOR function in.vitro and in vivo