Ihara Yoshito, Manabe Shino, Kanda Munetake, Kawano Hiroaki, Nakayama Toshiyuki, Sekine Ichiro, Kondo Takahito, Ito Yukishige
Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan.
Glycobiology. 2005 Apr;15(4):383-92. doi: 10.1093/glycob/cwi012. Epub 2004 Nov 3.
C-Mannosylation is a novel type of glycosylation in proteins. There are several examples of proteins in which the specific motif Trp-X-X-Trp is mannosylated at the first Trp to produce C-mannosylated Trp (CMW). Although C-mannosylation modifies Trp-X-X-Trp, predicted to be a functional motif of various integral proteins such as cytokine receptors, the physiological or pathological relevance of C-mannosylation in the cell is still not known. In this study, to characterize C-mannosylation in biological samples, we generated specific polyclonal antibodies against CMW by using a chemically synthesized CMW as an antigen. Using the antibody, we investigated the effect of hyperglycemic conditions on protein C-mannosylation in cultured cells and diabetic Zucker fatty rats. We found that protein C-mannosylation was increased in macrophage-like RAW264.7 cells under hyperglycemic conditions compared to low-glucose conditions. Furthermore, C-mannosylation was increased in the aortic vessel wall of Zucker fatty rats. Thrombospondin-1 was identified as a protein modified with C-mannosylation, and its expression was also increased in the aortic tissues of Zucker fatty rats. These results indicate that C-mannosylation is increased in specific tissues or cell types under hyperglycemic conditions, suggesting a pathological role for the increased C-mannosylation in the development of diabetic complications.
C-甘露糖基化是蛋白质中一种新型的糖基化修饰。有几个蛋白质的例子，其中特定基序Trp-X-X-Trp在第一个色氨酸处发生甘露糖基化，产生C-甘露糖基化色氨酸（CMW）。尽管C-甘露糖基化修饰Trp-X-X-Trp，预计其为细胞因子受体等多种整合蛋白的功能基序，但C-甘露糖基化在细胞中的生理或病理相关性仍不清楚。在本研究中，为了表征生物样品中的C-甘露糖基化，我们以化学合成的CMW为抗原，制备了针对CMW的特异性多克隆抗体。使用该抗体，我们研究了高糖条件对培养细胞和糖尿病Zucker肥胖大鼠中蛋白质C-甘露糖基化的影响。我们发现，与低糖条件相比，高糖条件下巨噬细胞样RAW264.7细胞中的蛋白质C-甘露糖基化增加。此外，Zucker肥胖大鼠的主动脉血管壁中C-甘露糖基化也增加。血小板反应蛋白-1被鉴定为一种发生C-甘露糖基化修饰的蛋白质，其在Zucker肥胖大鼠主动脉组织中的表达也增加。这些结果表明，在高糖条件下，特定组织或细胞类型中的C-甘露糖基化增加，提示C-甘露糖基化增加在糖尿病并发症发生发展中具有病理作用。
