Yoon Eunju, Beom Sunryeo, Cheong Ho, Kim Soyoung, Oak Minho, Cho Dongim, Kim Kyeong-Man
Department of Pharmacology, College of Pharmacy, Chonnam National University, Kwang-Ju 500-757, Republic of Korea.
Biochem Biophys Res Commun. 2004 Dec 3;325(1):117-23. doi: 10.1016/j.bbrc.2004.09.216.
The high affinity IgE receptor (FcepsilonRI) usually exists as a tetramer composed of alphabetagamma2 subunits. The COOH-tail of beta and gamma subunits contains consensus sequence termed 'immunoreceptor tyrosine-based activation motif' (ITAM). Tyrosine phosphorylated ITAM interacts with signaling proteins that contain the Src homology domain, forming a main amplifying and signaling route for FcepsilonRI. Unlike the COOH-tail, the functional role of NH(2)-tail of beta subunit in the signaling of FcepsilonRI is not clear because it lacks the ITAM sequences. To study the roles of NH(2)-tail of beta subunit, the cDNA library of RBL-2H3 cells was screened by yeast two-hybrid assay, and the NH(2)-tail of the beta subunit was found to interact with phospholipase Cgamma2 (PLCgamma2) but not with PLCgamma1. Since both PLCgamma1 and PLCgamma2 are expressed in RBL-2H3 cells and they possess identical cellular functions, the functional meaning of the protein-protein interaction between PLCgamma2 and NH(2)-tail of beta subunit was studied by comparing the regulatory pathways that control the FcepsilonRI-mediated tyrosine phosphorylation of the two enzymes. Our study shows that PI3-kinase and PMA-sensitive PKCs were required exclusively for the FcepsilonRI-mediated tyrosine phosphorylation of PLCgamma1. Also the FcepsilonRI-mediated tyrosine phosphorylation of PLCgamma1 was more sensitive to the inhibitors of Src and Syk kinases. These results therefore suggest that PLCgamma1 is involved in dynamic regulation of protein kinase C activity and inositol triphosphate levels in response to cellular needs. In contrast, PLCgamma2, through continuous interaction with the NH(2)-tail of beta subunit, co-localizes with FcepsilonRI in the same signaling domain, and maintains the basal cellular PLC activity.
高亲和力IgE受体（FcepsilonRI）通常以由αβγ2亚基组成的四聚体形式存在。β和γ亚基的COOH末端包含称为“基于免疫受体酪氨酸的激活基序”（ITAM）的共有序列。酪氨酸磷酸化的ITAM与含有Src同源结构域的信号蛋白相互作用，形成FcepsilonRI的主要放大和信号传导途径。与COOH末端不同，β亚基的NH(2)末端在FcepsilonRI信号传导中的功能作用尚不清楚，因为它缺乏ITAM序列。为了研究β亚基NH(2)末端的作用，通过酵母双杂交试验筛选了RBL-2H3细胞的cDNA文库，发现β亚基的NH(2)末端与磷脂酶Cγ2（PLCγ2）相互作用，但不与PLCγ1相互作用。由于PLCγ1和PLCγ2都在RBL-2H3细胞中表达且它们具有相同的细胞功能，通过比较控制两种酶的FcepsilonRI介导的酪氨酸磷酸化的调节途径，研究了PLCγ2与β亚基NH(2)末端之间蛋白质-蛋白质相互作用的功能意义。我们的研究表明，PI3激酶和PMA敏感的蛋白激酶C专门用于FcepsilonRI介导的PLCγ1酪氨酸磷酸化。此外，FcepsilonRI介导的PLCγ1酪氨酸磷酸化对Src和Syk激酶的抑制剂更敏感。因此，这些结果表明PLCγ1参与响应细胞需求对蛋白激酶C活性和肌醇三磷酸水平的动态调节。相比之下，PLCγ2通过与β亚基的NH(2)末端持续相互作用，与FcepsilonRI共定位于相同的信号域，并维持基础细胞PLC活性。
