A single base mutation in the PRAT4A gene reveals differential interaction of PRAT4A with Toll-like receptors
A single base mutation in the PRAT4A gene reveals differential interaction of PRAT4A with Toll-like receptors
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DOI：
10.1093/intimm/dxn098
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发表时间：
2008-11-01
影响因子：
4.4
通讯作者：
Miyake, Kensuke
中科院分区：
文献类型：
--
作者：
Kiyokawa, Takashi;Akashi-Takamura, Sachiko;Miyake, Kensuke
Toll样受体（TLR）在防御反应中起重要作用。免疫细胞表达多种TLR，这些TLR同时被微生物病原体激活。Tlr 4A相关蛋白（PRAT 4A）是一种分子伴侣样内质网（ER）驻留蛋白，是多种TLR正确亚细胞分布所必需的。PRAT 4A（-/-）小鼠显示细胞表面上TLR 2/4的表达受损，并且缺乏配体诱导的TLR 9从ER到内溶酶体的重新定位。因此，TLR对整个细菌以及TLR 2、4和9配体的应答受损。我们在这里比较这些TLR与PRAT 4A的相互作用。内源性PRAT 4A的关联仅与TLR 4容易检测。负责与PRAT 4A强相互作用的TLR 4区域非常接近与MD-2相互作用所必需的位点。通过使用瞬时表达，我们能够检测PRAT 4A与TLR 2和TLR 9的相互作用。用赖氨酸替换甲硫氨酸145的PRAT 4A单核苷酸突变体（M145 K）与TLR 9缔合，但不拯救配体依赖性TLR 9运输。相比之下，M145 K突变体与TLR 2和TLR 4弱相关（如果有的话）。M145 K突变体明显地挽救了PRAT 4A（-/-）骨髓来源的树突状细胞中的细胞表面TLR 2表达及其应答，而细胞表面TLR 4/MD-2表达及其应答几乎没有挽救。这些结果表明，PRAT 4A差异与每个TLR的相互作用，并表明在PRAT 4A基因的单核苷酸的变化不仅影响TLR反应的强度，但也可以改变每个TLR的相对活性。
Toll-like receptors (TLRs) play an essential role in defense responses. Immune cells express multiple TLRs which are simultaneously activated by microbial pathogens. PRotein Associated with Tlr4 A (PRAT4A) is a chaperone-like endoplasmic reticulum (ER)-resident protein required for the proper subcellular distribution of multiple TLRs. PRAT4A(-/-) mice show impaired expression of TLR2/4 on the cell surface and the lack of ligand-induced TLR9 relocation from the ER to endolysosome. Consequently, TLR responses to whole bacteria as well as to TLR2, 4 and 9 ligands are impaired. We here compare the interaction of these TLRs with PRAT4A. Association of endogenous PRAT4A was easily detected only with TLR4. The TLR4 region responsible for strong interaction with PRAT4A is very close to the site necessary for interaction with MD-2. By using transient expression, we were able to detect PRAT4A interaction with TLR2 and TLR9. The PRAT4A single-nucleotide mutant replacing methionine 145 with lysine (M145K) associates with TLR9 but does not rescue ligand-dependent TLR9 trafficking. By contrast, the M145K mutant weakly, if at all, associates with TLR2 and TLR4. The M145K mutant appreciably rescues cell-surface TLR2 expression and its responses in PRAT4A(-/-) bone marrow-derived dendritic cells, whereas little if any rescue of cell-surface TLR4/MD-2 expression and its responses occurs. These results demonstrate that PRAT4A differentially interacts with each TLR and suggest that a single-nucleotide change in the PRAT4A gene influences not only the strength of TLR responses but can also alter the relative activity of each TLR.