Hamada Toshiyuki, Miyakawa Kazuko, Kushige Hiroko, Shibata Shigenobu, Kurachi Sumiko
Applied Molecular-Imaging Physics, Hokkaido University Graduate School of Medicine, Sapporo, Hokkaido, 060-8638, Japan,
J Physiol Sci. 2015 Jul;65(4):349-57. doi: 10.1007/s12576-015-0373-8. Epub 2015 Apr 7.
In mammals, both circadian rhythm and aging play important roles in regulating time-dependent homeostasis. We previously discovered an age-related increase element binding protein, hnRNP A3, which binds to the 3'-untranslated region (UTR) of blood coagulation factor IX (FIX). Here, we describe other members of this protein family, hnRNP C and hnRNP H, which bind to the 3'-UTR of the mouse circadian clock gene Period 2 (mPer2). RNA electrophoretic mobility shift assays using a (32)P-labeled Per2 RNA probe coupled with two-dimensional gel electrophoresis followed by MALDI-TOF/MS peptide mass fingerprint analysis was used to analyze these proteins. Western blotting suggested that the total expression of these proteins in mouse liver cell nuclei does not increase with age. Two-dimensional gel electrophoresis analysis of age-related protein expression showed that many isoforms of these proteins exist in the liver and that each protein exhibits a complex age-related expression pattern. These results suggest that many isoforms of proteins are regulated by different aging systems and that many age regulation systems function in the liver.
在哺乳动物中，昼夜节律和衰老在调节时间依赖性内稳态方面都发挥着重要作用。我们之前发现了一种与年龄相关的增加元件结合蛋白hnRNP A3，它与凝血因子IX（FIX）的3'非翻译区（UTR）结合。在这里，我们描述了这个蛋白家族的其他成员，hnRNP C和hnRNP H，它们与小鼠生物钟基因Period 2（mPer2）的3'UTR结合。使用(32)P标记的Per2 RNA探针结合二维凝胶电泳，随后进行MALDI-TOF/MS肽质量指纹分析的RNA电泳迁移率变动分析用于分析这些蛋白质。蛋白质印迹法表明，这些蛋白质在小鼠肝细胞核中的总表达量不会随着年龄的增长而增加。对与年龄相关的蛋白质表达进行二维凝胶电泳分析表明，这些蛋白质的许多同工型存在于肝脏中，并且每种蛋白质都呈现出复杂的与年龄相关的表达模式。这些结果表明，蛋白质的许多同工型受不同的衰老系统调节，并且许多年龄调节系统在肝脏中发挥作用。
