Petti L M, Irusta P M, DiMaio D
Department of Genetics, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
Oncogene. 1998 Feb 19;16(7):843-51. doi: 10.1038/sj.onc.1201590.
We replaced the transmembrane domain of the wild type murine PDGF beta receptor with that of p185neu*, the oncogenic form of p185neu, thereby generating a constitutively activated chimeric receptor PR/neu*. Unlike the wild type PDGF beta receptor or a chimeric receptor containing the transmembrane domain of wild type p185neu (PR/neu), PR/neu* induced morphologic transformation, focus formation, and tumorigenicity in mouse C127 fibroblasts. Expression of PR/neu* in mouse Ba/F3 hematopoietic cells, which normally depend on IL-3 for survival and sustained proliferation, induced proliferation in the absence of IL-3. The PR/neu chimera conferred limited IL-3-independent growth of Ba/F3 cells. Only PR/neu* and not PR/neu displayed significantly increased levels of phosphotyrosine compared to the wild type PDGF receptor in C127 and Ba/F3 cells. In addition, PR/neu* immune complexes displayed increased levels of kinase activity in vitro compared to immune complexes of the wild type receptor. Furthermore, novel tyrosine phosphorylated proteins of approximately 60 kDa appeared to specifically complex with PR/neu*, suggesting that PR/neu* may activate distinct signaling pathways. We speculate that the p185neu* transmembrane domain in the context of the PDGF beta receptor facilitates receptor homodimerization, thereby inducing tyrosine autophosphorylation followed by association with important signaling substrates and transforming activity. Thus, PR/neu* should be a useful reagent for further characterizing activation and signaling mechanisms of the PDGF beta receptor.
我们将野生型小鼠血小板衍生生长因子β受体（PDGF beta受体）的跨膜结构域替换为p185neu*（p185neu的致癌形式）的跨膜结构域，从而产生了一种组成型激活的嵌合受体PR/neu*。与野生型PDGF beta受体或含有野生型p185neu跨膜结构域的嵌合受体（PR/neu）不同，PR/neu在小鼠C127成纤维细胞中诱导了形态转化、集落形成和致瘤性。PR/neu在小鼠Ba/F3造血细胞中的表达（这些细胞通常依赖白细胞介素-3生存和持续增殖）在没有白细胞介素-3的情况下诱导了增殖。PR/neu嵌合体赋予Ba/F3细胞有限的不依赖白细胞介素-3的生长能力。与C127和Ba/F3细胞中的野生型PDGF受体相比，只有PR/neu而非PR/neu显示出磷酸酪氨酸水平显著升高。此外，与野生型受体的免疫复合物相比，PR/neu免疫复合物在体外显示出更高的激酶活性水平。此外，约60 kDa的新型酪氨酸磷酸化蛋白似乎与PR/neu特异性结合，这表明PR/neu可能激活不同的信号通路。我们推测，在PDGF beta受体背景下的p185neu跨膜结构域促进了受体同源二聚化，从而诱导酪氨酸自身磷酸化，随后与重要的信号底物结合并产生转化活性。因此，PR/neu应该是进一步表征PDGF beta受体激活和信号传导机制的有用试剂。
