Mechanisms of positive and negative thymic selection of CD8 T cells

 描述（由申请方提供）：胸腺中TCR与自身肽-MHC复合物的相互作用控制着阳性选择、谱系定型和阴性选择的关键发育检查点，这些检查点对于功能性CD 8 T细胞库的生成至关重要。这些步骤背后的机制仍然不完全清楚。最初假设MHC I呈递的自身肽在所有选择检查点都是相同的。然而，一个令人兴奋的发现是，皮质胸腺上皮细胞（cTEC）表达一个独特的蛋白酶体活性位点亚基，ss 5 t，它纳入蛋白酶体产生胸腺蛋白酶体。ss 5 t的基因失活减少了CD 8 T细胞的发育。cTEC是呈递肽-MHC复合物用于阳性选择的关键细胞，并且本领域的普遍观点是胸腺蛋白酶体的作用是产生对于该选择步骤更优化的肽。另一种不受欢迎的模型是胸腺蛋白酶体不需要产生用于阳性选择的特殊肽，而是产生与参与阴性选择的肽不同的肽，从而防止阴性选择抵消阳性选择。到目前为止，研究人员已经使用了部分基因“敲除”来试图阐明胸腺蛋白酶体在正选择和负选择中的作用。我们已经建立了一个更完整的遗传模型，其中小鼠缺乏所有专门的蛋白酶体，并且在CD 8 T细胞发育中具有明显更强的阻断作用。与早期模型相比，这些小鼠在cTEC中缺乏所有三种胸腺蛋白酶体亚基，也完全不能在阳性和阴性选择步骤之间切换肽。我们建议使用我们的新模型与其他国家的最先进的遗传模型，文化系统，和质谱来阐明胸腺选择的潜在机制。我们的初步数据显示了可行性，并表明目前青睐的模型可能是不正确的。本提案有三个目的来测试这些问题.

 DESCRIPTION (provided by applicant): TCR interactions with self-peptide-MHC complexes in the thymus control the key developmental check points of positive selection, lineage commitment and negative selection that are essential for the generation of a functional CD8 T cell repertoire. The mechanisms underlying these various steps are still incompletely understood. It was originally assumed that the MHC I-presented self-peptides were identical at all the selection check points. However, an exciting discovery was that cortical thymic epithelial cells (cTEC) express a unique proteasome active site subunit, ss5t, which incorporates into proteasomes to generate thymoproteasomes. Genetic inactivation of ss5t reduces the development of CD8 T cells. cTECs are the key cells that present peptide-MHC complexes for positive selection and the prevailing opinion in the field is that the role of thymoproteasomes is to generate peptides that are more optimal for this selection step. An alternate and disfavored model is that thymoproteasomes are not required to generate special peptides for positive selection but rather to make different peptides from those involved in negative selection and thereby prevent negative selection from cancelling out positive selection. So far investigators have used partial genetic "knock outs" to try to elucidate the role of thymoproteasomes in positive and negative selection. We have generated a more complete genetic model in which mice lack all specialized proteasomes and turn out to have a significantly stronger block in CD8 T cell development. Compared to earlier models, these mice lack all three thymoproteasome subunits in cTECs are also completely unable to switch peptides between the positive and negative selection steps. We propose to use our novel model together with other state of the art genetic models, culture systems, and mass spectrometry to elucidate the underlying mechanisms of thymic selection. Our preliminary data show feasibility and suggest that the currently favored model may be incorrect. This proposal has 3 aims to test these elucidate these issues.