Hafizi Sassan, Gustafsson Anna, Stenhoff Jonas, Dahlbäck Björn
Department of Laboratory Medicine, Section for Clinical Chemistry, Lund University, Wallenberg Laboratory, University Hospital Malmö, SE-205 02 Malmö, Sweden.
Int J Biochem Cell Biol. 2005 Nov;37(11):2344-56. doi: 10.1016/j.biocel.2005.05.006.
Axl belongs to a particular subfamily of transmembrane receptor tyrosine kinases, the biological ligand for which is the growth/survival factor Gas6. However, little is known about the molecular mechanisms for Axl activation and signal transduction. We have previously identified a novel interaction between the intracellular domain of Axl and Ran binding protein in microtubule organising centre (RanBPM). In the present study, we investigated further the nature of the RanBPM interaction with Axl. A wide distribution of RanBPM mRNA expression in human tissues and various human cancer cell lines was detected. The strength of interaction of both proteins in yeast was comparable to that with the other Axl-binding proteins phosphatidylinositol 3-kinase and Grb2. A truncated version of RanBPM with the SPRY-LisH domain region omitted failed to interact with Axl in yeast. RanBPM was also found to interact in yeast with the Axl homologue, Sky/Tyro3. The interaction between Axl intracellular domain and RanBPM was reproduced in coimmunoprecipitation experiments in both cell-free and mammalian cell systems. Furthermore, coimmunoprecipitation revealed endogenous Axl and RanBPM to interact in several mammalian cell lines in a constitutive manner. Stimulation of COS cells with Gas6 caused increased Axl tyrosine phosphorylation although appeared not to influence the RanBPM-Axl association. In conclusion, we have identified and characterised a novel interaction between RanBPM and the related receptor tyrosine kinases, Axl and Sky. This novel insight into the signalling interactions of Axl and Sky may shed further light on their suspected roles in tumourigenesis, inflammation as well as other cell proliferative diseases.
Axl属于跨膜受体酪氨酸激酶的一个特定亚家族，其生物学配体是生长/存活因子Gas6。然而，关于Axl激活和信号转导的分子机制知之甚少。我们之前已经确定了Axl细胞内结构域与微管组织中心的Ran结合蛋白（RanBPM）之间存在一种新的相互作用。在本研究中，我们进一步研究了RanBPM与Axl相互作用的性质。检测到RanBPM mRNA在人类组织和各种人类癌细胞系中广泛表达。两种蛋白在酵母中的相互作用强度与其他与Axl结合的蛋白磷脂酰肌醇3激酶和Grb2相当。缺失SPRY-LisH结构域区域的RanBPM截短版本在酵母中无法与Axl相互作用。还发现RanBPM在酵母中与Axl同源物Sky/Tyro3相互作用。Axl细胞内结构域与RanBPM之间的相互作用在无细胞和哺乳动物细胞系统的共免疫沉淀实验中得到重现。此外，共免疫沉淀显示内源性Axl和RanBPM在几种哺乳动物细胞系中以组成型方式相互作用。用Gas6刺激COS细胞导致Axl酪氨酸磷酸化增加，尽管似乎不影响RanBPM-Axl的结合。总之，我们已经鉴定并表征了RanBPM与相关受体酪氨酸激酶Axl和Sky之间的一种新的相互作用。对Axl和Sky信号相互作用的这一新见解可能会进一步揭示它们在肿瘤发生、炎症以及其他细胞增殖性疾病中所怀疑的作用。
