Development of the reconstituted antigen presenting system and its medical application

蛋白酶体是一种大的多亚基聚合酶，是内源性抗原的加工酶，参与获得性免疫中自身与非自身的识别，在细胞免疫的启动中起着不可或缺的作用。我们发现干扰素-γ（IFN-γ）可诱导三对同源性较高的蛋白酶体亚基发生亚基置换，产生“免疫蛋白酶体”，强调其在抗原加工中的特殊功能，表明抗原加工过程可受多种细胞外信号的调控。对编码IFN-γ相关蛋白酶体亚单位的基因的遗传位点分析表明，位于MHC区域的各种基因，如TAP（抗原加工相关转运蛋白）和免疫蛋白酶体LMP 2/LMP 7亚单位已通过古老的染色体复制产生，为MHC基因的起源提供了新的假设。此外，我们发现一种新的蛋白酶体激活蛋白PA 28，由三个同源家族蛋白组成，也受到IFN-γ的调节，表明PA 28在免疫应答中起着重要作用。最后，我们发现蛋白酶体可以通过严格识别目标多肽的长度和序列，将合成的覆盖CTL表位序列的前体肽在体外产生抗原肽。此外，发现PA 28与蛋白酶体以双切割方式合作，正确地切割抗原肽。这些观察结果为我们理解内源性抗原的加工和呈递的调节机制提供了新的见解。这些结果将有助于体外重组抗原提呈系统的开发。

The proteasome, a large multisubunit cimplex, has been implicated to be a processing enzyme of endogeneous antigens, which is involved in the recognition between self and non-self in the adaptive immunity, implying that it plays an indispensable role for triggering the cell-mediated immunity. We found that interferon-gamma (IFN-gamma) induces subunit replacement of three pair of proteasomal subunits with high homology, producing "immunoproteasomes" termed to emphasize their specialized functions for antigen procesisng, indicating that the antigen processing process can be regulated in response to various extracellular signals. Analysis of genetic loci of the genes encoding IFN-gamma responsible proteasomal subunits revealed that various genes located on the MHC regions, such as TAP (transporter associated with antigen processing) and immunoproteasomal LMP2/LMP7 subunits have been produced by an ancient chromosomal duplication, providing a new hypothesis for the origin of the MHC genes. In addition, we found that a novel proteasomal activator protein PA28, consisting of three homologous family proteins, also are regulated by IFN-gamma, indicating that PA28 plays an essential role for the immune response. Finally, we found that the proteasome can generate antigenic peptide in vitro from the synthetic precursor plypeptide covering the CTL epitopic sequences by recognizing strictly the length and sequence of the targe polypeptides Moreover, PA28 was found to collaborate with the proteasome to excise correctly antigenic peptide in a dual-cleavage fashion. These observations provide a new insight for our understanding of the regulatory mechanisms of processing and presentation of endogenous antigens. These findings would contribute to developement of the reconstituted antigen presenting system in vitro.