Human Colon Tumors Express a Dominant-Negative Form of SIGIRR That Promotes Inflammation and Colitis-Associated Colon Cancer in Mice.
Human Colon Tumors Express a Dominant-Negative Form of SIGIRR That Promotes Inflammation and Colitis-Associated Colon Cancer in Mice.
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DOI：
10.1053/j.gastro.2015.08.051
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发表时间：
2015-12
期刊：
影响因子：
29.4
通讯作者：
Li X
中科院分区：
文献类型：
--
作者：
Zhao J;Bulek K;Gulen MF;Zepp JA;Karagkounis G;Martin BN;Zhou H;Yu M;Liu X;Huang E;Fox PL;Kalady MF;Markowitz SD;Li X
单一免疫球蛋白和 Toll 白细胞介素 1 受体 (SIGIRR) 是 Toll 样和白细胞介素 1 受体 (IL1R) 信号通路的负调节因子，可控制小鼠肠道炎症并抑制结肠肿瘤发生。然而，SIGIRR 在人类结直肠癌发展中的重要性尚未确定。我们研究了 SIGIRR 在人类结直肠癌发展中的作用。我们对来自 68 名患者的结肠肿瘤和非肿瘤组织进行了 RNA 序列分析。使用免疫印迹和免疫荧光分析来测定原代人结肠上皮细胞、肿瘤组织和结肠癌细胞系中 SIGIRR 蛋白的水平。我们在 Vaco 细胞系中表达了 SIGIRR 和该蛋白的突变形式。我们创建并分析了在肠上皮中特异性表达全长（对照）或 Sigirr 突变形式（编码 SIGIRRN86/102S，未糖基化）的小鼠。一些小鼠被给予氧化偶氮甲烷和葡聚糖硫酸钠来诱发结肠炎相关癌症。收集肠组织并通过免疫组织化学和基因表达谱分析进行分析。 RNA 序列分析显示，与配对的非肿瘤组织相比，结直肠癌组织中 SIGIRR mRNA 亚型 SIGIRRΔE8 的表达增加。 SIGIRRΔE8 未被复杂聚糖修饰，因此保留在细胞质中 - 它不能定位于细胞膜或减少 IL1R 信号传导。 SIGIRRΔE8 与 SIGIRR 相互作用并对 SIGIRR 产生显性负效应，减少其糖基化、细胞表面定位和功能。在人类结肠癌组织中检测到的大多数 SIGIRR 是在细胞质中，而在非肿瘤组织中，它是在细胞膜上发现的。与对照小鼠相比，表达 SIGIRRN86/102S 的小鼠在给予氧化偶氮甲烷和葡聚糖硫酸钠后出现更多炎症并形成更大的肿瘤；这些突变小鼠的结肠组织表达较高水平的炎症细胞因子 IL17A 和 IL6，并激活转录因子 STAT3 和 NFκB。在小鼠结肠中表达的 SIGIRRN86/102S 并未定位于上皮细胞表面。与非肿瘤组织相比，人类结直肠肿瘤中的 SIGIRR 水平较低；肿瘤含有显性失活亚型 SIGIRRΔE8。这种突变蛋白阻断全长 SIGIRR 在结肠上皮细胞表面的定位及其下调 IL1R 信号传导的能力。 SIGIRRN86/102S 在小鼠结肠上皮中的表达会增加炎症细胞因子的表达以及结肠炎相关肿瘤的形成和大小。
Single immunoglobulin and toll-interleukin 1 receptor (SIGIRR), a negative regulator of the Toll-like and interleukin-1 receptor (IL1R) signaling pathways, controls intestinal inflammation and suppresses colon tumorigenesis in mice. However, the importance of SIGIRR in human colorectal cancer development has not been determined. We investigated the role of SIGIRR in development of human colorectal cancer. We performed RNA sequence analyses of pairs of colon tumor and non-tumor tissues, each collected from 68 patients. Immunoblot and immunofluorescence analyses were used to determine levels of SIGIRR protein in primary human colonic epithelial cells, tumor tissues, and colon cancer cell lines. We expressed SIGIRR and mutant forms of the protein in Vaco cell lines. We created and analyzed mice that expressed full-length (control) or a mutant form of Sigirr (encoding SIGIRRN86/102S, which is not glycosylated) specifically in the intestinal epithelium. Some mice were given azoxymethane and dextran sulfate sodium to induce colitis-associated cancer. Intestinal tissues were collected and analyzed by immunohistochemical and gene expression profile analyses. RNA sequence analyses revealed increased expression of a SIGIRR mRNA isoform, SIGIRRΔE8, in colorectal cancer tissues compared to paired non-tumor tissues. SIGIRRΔE8 is not modified by complex glycans and is therefore retained in the cytoplasm—it cannot localize to the cell membrane or reduce IL1R signaling. SIGIRRΔE8 interacts with and has a dominant-negative effect on SIGIRR, reducing its glycosylation, localization to the cell surface, and function. Most SIGIRR detected in human colon cancer tissues was cytoplasmic, whereas in non-tumor tissues it was found at the cell membrane. Mice that expressed SIGIRRN86/102S developed more inflammation and formed larger tumors after administration of azoxymethane and dextran sulfate sodium than control mice; colon tissues from these mutant mice expressed higher levels of the inflammatory cytokines IL17A and IL6 had activation of the transcription factors STAT3 and NFκB. SIGIRRN86/102S expressed in colons of mice did not localize to the epithelial cell surface. Levels of SIGIRR are lower in human colorectal tumors, compared with non-tumor tissues; tumors contain the dominant-negative isoform SIGIRRΔE8. This mutant protein blocks localization of full-length SIGIRR to the surface of colon epithelial cells and its ability to downregulate IL1R signaling. Expression of SIGIRRN86/102S in the colonic epithelium of mice increases expression of inflammatory cytokines and formation and size of colitis-associated tumors.