Higuchi Megumi, Matsuo Aya, Shingai Masashi, Shida Kyoko, Ishii Akihiro, Funami Kenji, Suzuki Yasuhiko, Oshiumi Hiroyuki, Matsumoto Misako, Seya Tsukasa
Department of Microbiology and Immunology, Hokkaido University Graduate School of Medicine, Kita-15, Nishi-7, Kita-ku, Sapporo 060-8638, Japan.
Dev Comp Immunol. 2008;32(2):147-55. doi: 10.1016/j.dci.2007.05.003. Epub 2007 Jun 15.
Human Toll-like receptor 2 (TLR2) subfamily recognizes bacterial lipoproteins (BLP) and peptidoglycan (PGN). According to the genome information, chicken has structural orthologs of TLRs1 and 2, in addition to TLRs3, 4, 5 and 7. Chicken has two additional TLRs, TLR15 and TLR21, whose orthologs human lacks. The chicken (ch)TLR1 and 2 genes are individually duplicated to encode for four different proteins, chTLR1-1, 1-2, 2-1 and 2-2, of the TLR2 subfamily. Here we investigated the functional profile of these TLR2 subfamily proteins of chicken. By NF-kappaB reporter assay using HEK293 cells, we found that chTLR2-1 and chTLR1-2 cooperatively signal the presence of PGN. A combination of chTLR2-1 and chTLR1-2 also most efficiently recognized diacylated BLP, macrophage-activating lipopeptide 2kDa (Malp-2), while the combination of chTLR2-1 and chTLR1-1 failed to recognize Malp-2. All combinations, however, recognized triacylated BLP, Pam3. Consistent with these results, human TLR2-stimulating mycobacteria preparations, BCG-cell wall and cell lysate of Mycobacterium avium, induced activation of NF-kappaB in cells expressing chTLR2-1 and 1-2 and to lesser extents, cells with chTLR2-2 and either of chTLR1. Strikingly, expression of either of these alone did not activate the reporter for NF-kappaB. These chTLRs are likely to have the combination functional feature as in the human TLR2 subfamily. Confocal and immunoprecipitation analyses of human cell transfectants showed that they cluster on the cell surface by a physical molecular association, causing all of them to merge and coprecipitate. These results suggest that chTLR2 subfamily members discriminate between their ligands by combinational events.
人类 Toll 样受体 2（TLR2）亚家族可识别细菌脂蛋白（BLP）和肽聚糖（PGN）。根据基因组信息，鸡除了有 TLR3、4、5 和 7 外，还有 TLR1 和 2 的结构直系同源物。鸡还有另外两个 TLR，即 TLR15 和 TLR21，人类没有它们的直系同源物。鸡（ch）的 TLR1 和 2 基因各自发生复制，以编码 TLR2 亚家族的四种不同蛋白质，即 chTLR1-1、1-2、2-1 和 2-2。在此，我们研究了鸡的这些 TLR2 亚家族蛋白的功能概况。通过使用 HEK293 细胞进行 NF-κB 报告基因检测，我们发现 chTLR2-1 和 chTLR1-2 协同发出 PGN 存在的信号。chTLR2-1 和 chTLR1-2 的组合也最有效地识别二酰化 BLP，即巨噬细胞激活脂肽 2kDa（Malp-2），而 chTLR2-1 和 chTLR1-1 的组合无法识别 Malp-2。然而，所有组合都能识别三酰化 BLP，即 Pam3。与这些结果一致，刺激人类 TLR2 的分枝杆菌制剂、卡介苗细胞壁和鸟分枝杆菌细胞裂解物，在表达 chTLR2-1 和 1-2 的细胞中诱导 NF-κB 激活，在程度较轻的情况下，在表达 chTLR2-2 和任一 chTLR1 的细胞中也能诱导 NF-κB 激活。令人惊讶的是，单独表达这些蛋白中的任何一种都不会激活 NF-κB 报告基因。这些鸡 TLR 可能具有与人类 TLR2 亚家族相同的组合功能特征。对人类细胞转染体的共聚焦和免疫沉淀分析表明，它们通过物理分子缔合在细胞表面聚集，导致它们全部融合并共沉淀。这些结果表明，chTLR2 亚家族成员通过组合事件区分其配体。
