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Monomeric C-mannosyl tryptophan is a degradation product of autophagy in cultured cells.
Glycoconjugate Journal ( IF 3 ) Pub Date : 2020-08-17 , DOI: 10.1007/s10719-020-09938-8
Shiho Minakata 1 , Yoko Inai 1 , Shino Manabe 2, 3 , Kazuchika Nishitsuji 1 , Yukishige Ito 4 , Yoshito Ihara 1
Affiliation  

C-Mannosyl tryptophan (C-Man-Trp) is a unique glycosylated amino acid present in various eukaryotes. The C-Man-Trp structure can be found as a monomeric form or a part of post-translational modifications within polypeptide chains in living organisms. However, the mechanism of how monomeric C-Man-Trp is produced has not been fully investigated. In this study, we assessed levels of cellular C-Man-Trp by ultra performance liquid chromatography with a mass spectrometry assay system, and investigated whether the cellular C-Man-Trp is affected by autophagy induction. The intracellular C-Man-Trp level was significantly increased under serum and/or amino acid starvation in A549, HaCaT, HepG2, NIH3T3, and NRK49F cells. The increase in C-Man-Trp was also observed in NIH3T3 cells treated with rapamycin, an autophagy inducer. The up-regulation of C-Man-Trp caused by starvation was reversed by the inhibition of lysosomal enzymes. We further showed that C-Man-Trp is produced by incubating a synthetic C-mannosylated peptide (C-Man-Trp-Ser-Pro-Trp) or thrombospondin (TSP) in a lysosomal fraction that was prepared from a mouse liver, which provides supporting evidence that C-Man-Trp is a degradation product of the C-mannosylated peptide or protein following lysosome-related proteolysis. Taken together, we propose that the autophagic pathway is a novel pathway that at least partly contributes to intracellular C-Man-Trp production under certain conditions, such as nutrient starvation.



中文翻译:

单体 C-甘露糖基色氨酸是培养细胞中自噬的降解产物。

C-甘露糖基色氨酸 ( C - Man -Trp) 是一种独特的糖基化氨基酸,存在于各种真核生物中。所述Ç -Man-色氨酸结构可以发现作为单体形式或在活生物体中的多肽链内的翻译后修饰的一部分。然而,单体C -Man-Trp如何产生的机制尚未得到充分研究。在这项研究中,我们通过超高效液相色谱和质谱分析系统评估了细胞C -Man-Trp 的水平,并研究了细胞C -Man-Trp 是否受自噬诱导的影响。细胞内C-Man-Trp 水平在 A549、HaCaT、HepG2、NIH3T3 和 NRK49F 细胞中的血清和/或氨基酸饥饿下显着增加。在用雷帕霉素(一种自噬诱导剂)处理的 NIH3T3 细胞中也观察到C -Man-Trp的增加。由饥饿引起的C -Man-Trp的上调被溶酶体酶的抑制逆转。我们进一步表明,C - Man -Trp 是通过在从小鼠肝脏制备的溶酶体部分中孵育合成的C-甘露糖基化肽 ( C - Man -Trp-Ser-Pro-Trp) 或血小板反应蛋白 (TSP) 产生的。提供支撑的证据表明,ç -Man-Trp为所述的降解产物ç溶酶体相关蛋白水解后的甘露糖基化肽或蛋白质。总之,我们提出自噬途径是一种新途径,在某些条件下,例如营养饥饿,它至少部分有助于细胞内C -Man-Trp 的产生。

更新日期:2020-08-20
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