Departments of Chemistry and Physics, McGill University, Montreal, QC, Canada H3G 0B1.
Proc Natl Acad Sci U S A. 2011 Apr 26;108(17):7016-21. doi: 10.1073/pnas.1018280108. Epub 2011 Apr 11.
Cell signaling involves dynamic changes in protein oligomerization leading to the formation of different signaling complexes and modulation of activity. Spatial intensity distribution analysis (SpIDA) is an image analysis method that can directly measure oligomerization and trafficking of endogenous proteins in single cells. Here, we show the use of SpIDA to quantify dimerization/activation and surface transport of receptor protein kinases--EGF receptor and TrkB--at early stages of their transactivation by several G protein-coupled receptors (GPCRs). Transactivation occurred on the same timescale and was directly limited by GPCR activation but independent of G-protein coupling types. Early receptor protein kinase transactivation and internalization were not interdependent for all receptor pairs tested, revealing heterogeneity between groups of GPCRs. SpIDA also detected transactivation of TrkB by dopamine receptors in intact neurons. By allowing for time and space resolved quantification of protein populations with heterogeneous oligomeric states, SpIDA provides a unique approach to undertake single cell multivariate quantification of signaling processes involving changes in protein interactions, trafficking, and activity.
细胞信号转导涉及蛋白质寡聚化的动态变化，导致不同信号复合物的形成和活性的调节。空间强度分布分析（SpIDA）是一种图像分析方法，可以直接测量内源性蛋白质在单细胞中的寡聚化和运输。在这里，我们展示了 SpIDA 的使用，以量化受体蛋白激酶——表皮生长因子受体（EGF 受体）和 TrkB——在其被几种 G 蛋白偶联受体（GPCR）早期转激活时的二聚化/激活和表面转运。转激活发生在相同的时间尺度上，直接受到 GPCR 激活的限制，但与 G 蛋白偶联类型无关。对于所有测试的受体对，早期受体蛋白激酶的转激活和内化并不相互依赖，这揭示了 GPCR 组之间的异质性。SpIDA 还检测到多巴胺受体在完整神经元中对 TrkB 的转激活。通过允许对具有不同寡聚状态的蛋白质群体进行时间和空间分辨的定量，SpIDA 为涉及蛋白质相互作用、运输和活性变化的信号转导过程的单细胞多变量定量提供了一种独特的方法。
