School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Kanagawa 226-8501, Japan.
Department of Emergency and Disaster Medicine Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan.
Food Funct. 2022 Oct 31;13(21):11021-11033. doi: 10.1039/d2fo00886f.
Commensal intestinal microbiota interacts with gut epithelial cells in the host by binding to specific host receptors. Several pattern recognition receptors on the gut that sense conserved microbial-associated molecular patterns have been reported; however, many of the gut receptor molecules involved in bacterial binding have not yet been identified. In this study, commensal intestinal bacteria interacting with mouse gut surface proteins were screened from fecal bacterial samples, to identify novel receptors on the epithelial cells in the mouse gut. Among the screened intestinal lactic acid bacteria, the frequently isolated MG was used for the purification of gut receptor proteins. An approximately 30 kDa protein was purified using affinity resin coupled surface layer proteins isolated from MG. The purified gut protein was identified as a member of the tight junction protein family, junctional adhesion molecule-2 (JAM-2). As expected, the tight junctions of Caco-2 cells damaged by HO were repaired by incubation with MG. RNA sequence analysis showed significant upregulation of the expression of genes for tight junctions, anti-inflammatory effects, transcriptional regulation, and apoptosis in Caco-2 cells, following MG treatment. In MG, the surface layer 40 kDa protein was purified with gut protein-coupled affinity resin and identified as the moonlighting protein glyceraldehyde-3-phosphate dehydrogenase (GAPDH). These results suggest that MG promotes the barrier function integrity in Caco-2 cells GAPDH-JAM-2 binding. Here, we propose a promising approach to identify novel gut receptor molecules based on commensal bacterial interactions and understand host-bacterial communication in a mouse model.
肠道共生菌群通过与宿主特定的受体结合与肠道上皮细胞相互作用。已经报道了几种能够识别保守的微生物相关分子模式的肠道模式识别受体；然而，许多参与细菌结合的肠道受体分子尚未被鉴定。在这项研究中，从粪便细菌样本中筛选了与小鼠肠道表面蛋白相互作用的共生肠道细菌，以鉴定小鼠肠道上皮细胞上的新型受体。在所筛选的肠道乳酸菌中，经常分离到的 MG 用于纯化肠道受体蛋白。使用从 MG 中分离的表面层蛋白偶联的亲和树脂纯化了约 30 kDa 的蛋白质。纯化的肠道蛋白被鉴定为紧密连接蛋白家族的成员，即连接粘附分子-2（JAM-2）。正如预期的那样，用 MG 孵育可以修复由 HO 损伤的 Caco-2 细胞的紧密连接。RNA 序列分析显示，MG 处理后，Caco-2 细胞中紧密连接、抗炎作用、转录调节和细胞凋亡相关基因的表达显著上调。在 MG 中，用与肠道蛋白偶联的亲和树脂纯化了表面层 40 kDa 蛋白，并鉴定为多功能蛋白甘油醛-3-磷酸脱氢酶（GAPDH）。这些结果表明，MG 通过 GAPDH-JAM-2 结合促进了 Caco-2 细胞中屏障功能的完整性。在这里，我们提出了一种有前途的方法，基于共生细菌的相互作用来鉴定新型肠道受体分子，并在小鼠模型中理解宿主-细菌的通讯。
