Takahashi Noriko, Ohba Toshihiro, Imai Masahiko, Hasegawa Shinya, Takahashi Katsuhiko, Yamasaki Masahiro, Kameoka Yuri
Laboratory of Physiological Chemistry, Institute of Medicinal Chemistry, Hoshi University, Shinagawa, Tokyo 142-8501, Japan.
Laboratory of Physiological Chemistry, Institute of Medicinal Chemistry, Hoshi University, Shinagawa, Tokyo 142-8501, Japan.
Biochim Biophys Acta. 2016 Dec;1861(12 Pt A):2011-2019. doi: 10.1016/j.bbalip.2016.10.001. Epub 2016 Oct 6.
Retinoic acid (RA) has a variety of biological effects in mammalian cells and tissues. It is well known that RA induces differentiation of human acute promyelocytic leukemia (APL) HL60 cells, fresh human APL cells, and clinical remission in patients with APL. Retinoylation (acylation of proteins by RA) is a possible pathway for RA action. However, an understanding of the role that retinoylation plays in the actions of RA is lacking. In the current study, several retinoylated proteins were detected in RA-treated HL60 fractions following Mono Q anion exchange chromatography and analysis using two-dimensional polyacrylamide gel electrophoresis. One of the retinoylated proteins was identified as Rho-GDIβ (28kDa) by TOF-MS and co-migration with Rho-GDIβ (28kDa). Truncated Rho-GDIβ (23kDa, N∆19), a product of cleavage by caspase-3, was not retinoylated. RA covalently bound to the Thr2 residue in Rho-GDIβ (5kDa), which is the second product resulting from the cleavage of Rho-GDIβ (28kDa) by caspase-3. RA treatment increased the level of Rho-GDIβ (28kDa) and decreased the level of Rho-GDIβ (23kDa). RA did not induce caspase-3 activity or Rho-GDIβ mRNA expression. It is likely that retinoylation of Rho-GDIβ increases its metabolic stability.
维甲酸（RA）在哺乳动物细胞和组织中具有多种生物学效应。众所周知，RA可诱导人急性早幼粒细胞白血病（APL）HL60细胞、新鲜人APL细胞分化，并使APL患者实现临床缓解。维甲酰化作用（蛋白质被RA酰化）是RA发挥作用的一种可能途径。然而，目前对于维甲酰化在RA作用中所起的作用尚缺乏了解。在本研究中，经Mono Q阴离子交换色谱和二维聚丙烯酰胺凝胶电泳分析，在经RA处理的HL60组分中检测到了几种维甲酰化蛋白。其中一种维甲酰化蛋白经飞行时间质谱（TOF-MS）鉴定为Rho-GDIβ（28kDa），且与Rho-GDIβ（28kDa）共迁移。截短的Rho-GDIβ（23kDa，N∆19），即caspase-3切割产物，未发生维甲酰化。RA与Rho-GDIβ（5kDa）中的苏氨酸2残基共价结合，该残基是caspase-3切割Rho-GDIβ（28kDa）产生的第二个产物。RA处理可提高Rho-GDIβ（28kDa）水平，并降低Rho-GDIβ（23kDa）水平。RA并未诱导caspase-3活性或Rho-GDIβ mRNA表达。Rho-GDIβ的维甲酰化可能会提高其代谢稳定性。
