Key Laboratory of Molecular Biophysics of the Ministry of Education, National Engineering Research Center for Nanomedicine, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, China.
Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, China.
Autophagy. 2021 Oct;17(10):3030-3047. doi: 10.1080/15548627.2020.1851496. Epub 2020 Dec 17.
Although genome-wide association studies have identified the gene encoding an E3 ubiquitin-protein ligase as conferring susceptibility to ulcerative colitis, the exact function of this protein remains unclear. In the present study, we demonstrate an important role for RNF186 in macroautophagy/autophagy activation in colonic epithelial cells and intestinal homeostasis. Mechanistically, RNF186 acts as an E3 ubiquitin-protein ligase for EPHB2 and regulates the ubiquitination of EPHB2. Upon stimulation by ligand EFNB1 (ephrin B1), EPHB2 is ubiquitinated by RNF186 at Lys892, and further recruits MAP1LC3B for autophagy. Compared to control mice, and mice have a more severe phenotype in the DSS-induced colitis model, which is due to a defect in autophagy in colon epithelial cells. More importantly, treatment with ephrin-B1-Fc recombinant protein effectively relieves DSS-induced mouse colitis, which suggests that ephrin-B1-Fc may be a potential therapy for human inflammatory bowel diseases.: ACTB: actin beta; ATG5: autophagy related 5; ATG16L1: autophagy related 16 like 1; ATP: adenosine triphosphate; Cas9: CRISPR associated protein 9; CD: Crohn disease; CQ: chloroquine; : colony stimulating factor 2; : c-x-c motif chemokine ligand 1; DMSO: dimethyl sulfoxide; DSS: dextran sodium sulfate; EFNB1: ephrin B1; EPHB2: EPH receptor B2; EPHB3: EPH receptor B3; EPHB2: lysine 788 mutated to arginine in EPHB2; EPHB2: lysine 892 mutated to arginine in EPHB2; ER: endoplasmic reticulum; FITC: fluorescein isothiocyanate; GFP: green fluorescent protein; GWAS: genome-wide association studies; HRP: horseradish peroxidase; HSPA5/BiP: heat shock protein family A (Hsp70) member 5; IBD: inflammatory bowel diseases; : interleukin 1 beta; : interleukin 6; IRGM:immunity related GTPase M; i.p.: intraperitoneally; IPP: inorganic pyrophosphatase; KD: knockdown; KO: knockout; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; NOD2: nucleotide binding oligomerization domain containing 2; PI3K: phosphoinositide 3-kinase; PtdIns3K: class III phosphatidylinositol 3-kinase; RNF186: ring finger protein 186; RNF186: alanine 64 mutated to threonine in RNF186; RNF186: arginine 179 mutated to X in RNF186; RPS6: ribosomal protein S6; : tumor necrosis factor; SQSTM1: sequestosome 1; Ub: ubiquitin; UBE2D2: ubiquitin conjugating enzyme E2 D2; UBE2H: ubiquitin conjugating enzyme E2 H; UBE2K: ubiquitin conjugating enzyme E2 K; UBE2N: ubiquitin conjugating enzyme E2 N; UC: ulcerative colitis; ULK1:unc-51 like autophagy activating kinase 1; WT: wild type.
虽然全基因组关联研究已经确定了编码 E3 泛素蛋白连接酶的基因赋予溃疡性结肠炎易感性，但该蛋白质的确切功能仍不清楚。在本研究中，我们证明了 RNF186 在结肠上皮细胞中的巨自噬/自噬激活以及肠道稳态中的重要作用。在机制上，RNF186 作为 EPHB2 的 E3 泛素蛋白连接酶，调节 EPHB2 的泛素化。在配体 EFNB1（ephrin B1）刺激下，EPHB2 在赖氨酸 892 处被 RNF186 泛素化，并进一步募集 MAP1LC3B 进行自噬。与对照小鼠相比，和 小鼠在 DSS 诱导的结肠炎模型中表现出更严重的表型，这是由于结肠上皮细胞中的自噬缺陷所致。更重要的是，用 Ephrin-B1-Fc 重组蛋白治疗可有效缓解 DSS 诱导的小鼠结肠炎，这表明 Ephrin-B1-Fc 可能是人类炎症性肠病的潜在治疗方法。: ACTB: actin beta; ATG5: autophagy related 5; ATG16L1: autophagy related 16 like 1; ATP: adenosine triphosphate; Cas9: CRISPR associated protein 9; CD: Crohn disease; CQ: chloroquine; : colony stimulating factor 2; : c-x-c motif chemokine ligand 1; DMSO: dimethyl sulfoxide; DSS: dextran sodium sulfate; EFNB1: ephrin B1; EPHB2: EPH receptor B2; EPHB3: EPH receptor B3; EPHB2: lysine 788 mutated to arginine in EPHB2; EPHB2: lysine 892 mutated to arginine in EPHB2; ER: endoplasmic reticulum; FITC: fluorescein isothiocyanate; GFP: green fluorescent protein; GWAS: genome-wide association studies; HRP: horseradish peroxidase; HSPA5/BiP: heat shock protein family A (Hsp70) member 5; IBD: inflammatory bowel diseases; : interleukin 1 beta; : interleukin 6; IRGM:immunity related GTPase M; i.p.: intraperitoneally; IPP: inorganic pyrophosphatase; KD: knockdown; KO: knockout; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; NOD2: nucleotide binding oligomerization domain containing 2; PI3K: phosphoinositide 3-kinase; PtdIns3K: class III phosphatidylinositol 3-kinase; RNF186: ring finger protein 186; RNF186: alanine 64 mutated to threonine in RNF186; RNF186: arginine 179 mutated to X in RNF186; RPS6: ribosomal protein S6; : tumor necrosis factor; SQSTM1: sequestosome 1; Ub: ubiquitin; UBE2D2: ubiquitin conjugating enzyme E2 D2; UBE2H: ubiquitin conjugating enzyme E2 H; UBE2K: ubiquitin conjugating enzyme E2 K; UBE2N: ubiquitin conjugating enzyme E2 N; UC: ulcerative colitis; ULK1:unc-51 like autophagy activating kinase 1; WT: wild type.
