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Novel interaction between the human 5-HT7 receptor isoforms and PLAC-24/eIF3k

机译：人5-HT7受体亚型与PLAC-24 / eIF3k之间的新型相互作用

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Three 5-HT7 receptor isoforms are expressed in rat and man, which differ in the amino acid sequence of their C-terminus. Thus far, no changes have been observed in the pharmacological profile of all three isoforms. To further elucidate the signal transduction pathway specific for these receptor variants, we screened for possible interacting proteins of the C-terminus of the h5-HT7(a) variant in a human foetal brain cDNA library. Using a yeast two-hybrid assay, we isolated PLAC-24/eIF3k as a possible interacting candidate. The association of PLAC-24 with all three receptor variants was observed and further reconfirmed in vivo by co-immunoprecipitation of PLAC-24 with the full-length receptor isoforms in transfected COS-7 cells. Studies with different deletion mutants of the receptor showed that the interaction between PLAC-24 and the receptor is not restricted to the C-terminus of the receptor. PLAC-24/eIF3k consists of 3 domains: an N-terminal HAM domain, a central WH domain and a C-terminal tail. We generated different domain constructs of PLAC-24, which indicated that the HAM and WH domain both interact with the 5-HT7(a) receptor. Overexpression of PLAC-24 in FIEK293 cells, stably expressing the h5-HT7(a) receptor, caused a threefold augmentation in the expression levels of the receptor. Co-localisation studies in COS-7 cells showed that PLAC-24 relocates from the nucleus and perinuclear sites towards the plasma membrane upon co-expression with the receptor. On the other hand, the expression of domain variants of PLAC-24 seems to block the translocation of the receptor towards the membrane. These observations suggest that PLAC-24 may play a role in the transport and the stabilisation of newly synthesised 5-HT7 receptor towards the plasma membrane. (c) 2006 Elsevier Inc. All rights reserved.
机译：在大鼠和人类中表达了三种5-HT7受体同工型，它们的C端氨基酸序列不同。迄今为止，尚未观察到所有三种同工型的药理学变化。为了进一步阐明对这些受体变体特异的信号转导途径，我们在人胎脑cDNA文库中筛选了h5-HT7（a）变体C末端可能相互作用的蛋白。使用酵母双杂交测定法，我们分离了PLAC-24 / eIF3k作为可能的相互作用候选物。观察到PLAC-24与所有三个受体变体的关联，并通过在转染的COS-7细胞中对PLAC-24与全长受体同工型进行共免疫沉淀在体内进一步证实。用受体的不同缺失突变体进行的研究表明，PLAC-24与受体之间的相互作用不限于受体的C端。 PLAC-24 / eIF3k由3个域组成：N末端HAM域，中央WH结构域和C末端尾巴。我们生成了PLAC-24的不同结构域构建体，表明HAM和WH结构域均与5-HT7（a）受体相互作用。稳定表达h5-HT7（a）受体的FIEK293细胞中PLAC-24的过表达导致该受体表达水平增加了三倍。在COS-7细胞中的共定位研究表明，与受体共表达后，PLAC-24从细胞核和核周位置向质膜迁移。另一方面，PLAC-24结构域变体的表达似乎阻止了受体向膜的转运。这些观察结果表明，PLAC-24可能在新合成的5-HT7受体向质膜的运输和稳定中发挥作用。 （c）2006 Elsevier Inc.保留所有权利。

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