Paul Suman, Schaefer Brian C
Department of Microbiology and Immunology, Uniformed Services University, Bethesda, MD, 20814, USA.
Methods Mol Biol. 2015;1280:219-38. doi: 10.1007/978-1-4939-2422-6_12.
T cells are an immune cell lineage that play a central role in protection against pathogen infection. Antigen, in the form of pathogen-derived peptides, stimulates the T-cell receptor (TCR), leading to activation of the transcription factor, nuclear factor kappa B (NF-κB). The subsequent NF-κB-dependent gene expression program drives expansion and effector differentiation of antigen-specific T cells, leading to the adaptive anti-pathogen immune response. The cell surface TCR transmits activating signals to cytosolic NF-κB by a complex signaling cascade, in which the adapter protein Bcl10 plays a key role. We have previously demonstrated that TCR engagement leads to the formation of cytosolic Bcl10 clusters, called POLKADOTS, that provide a platform for the assembly of the terminal signaling complex that ultimately mediates NF-κB activation. In this chapter, we describe the methods utilized to visualize the formation of TCR-induced POLKADOTS and to study the temporal association between POLKADOTS formation and nuclear translocation of the NF-κB subunit, RelA/p65.
T细胞是一种免疫细胞谱系，在抵御病原体感染中发挥核心作用。病原体衍生肽形式的抗原刺激T细胞受体（TCR），导致转录因子核因子κB（NF-κB）活化。随后依赖NF-κB的基因表达程序驱动抗原特异性T细胞的扩增和效应分化，从而引发适应性抗病原体免疫反应。细胞表面的TCR通过复杂的信号级联将激活信号传递至胞质中的NF-κB，其中衔接蛋白Bcl10发挥关键作用。我们之前已经证明，TCR结合会导致胞质中形成称为POLKADOTS的Bcl10簇，其为最终介导NF-κB活化的末端信号复合物的组装提供了一个平台。在本章中，我们描述了用于可视化TCR诱导的POLKADOTS形成以及研究POLKADOTS形成与NF-κB亚基RelA/p65核转位之间时间关联的方法。
