1. Cytosolic LC3 ratio as a sensitive index of macroautophagy in isolated rat hepatocytes and H4-II-E cells.
- Author
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Karim MR, Kanazawa T, Daigaku Y, Fujimura S, Miotto G, and Kadowaki M
- Subjects
- Animals, Biomarkers metabolism, Blotting, Western, Carcinoma, Hepatocellular pathology, Cell Line, Tumor, Hydrolysis, Liver Neoplasms pathology, Male, Models, Biological, Rats, Rats, Wistar, Sensitivity and Specificity, Subcellular Fractions metabolism, Autophagy, Cytosol metabolism, Hepatocytes metabolism, Microtubule-Associated Proteins metabolism
- Abstract
Macroautophagy, an intracellular bulk degradation process in eukaryotes, is sensitive to nutrient supply and deprivation. Microtubule-associated protein 1 light chain 3 (LC3), a mammalian homologue of yeast Atg8, plays an indispensable role in macroautophagy formation and is a suitable marker for this process. Through analysis of the subcellular distribution of LC3, we determined that the cytosolic fraction contained not only a precursor form (LC3-I), but also an apparent active form (LC3-IIs). Both cytosolic LC3-I and LC3-IIs were more responsive to amino acids than those of total homogenate. Moreover, changes in the LC3-IIs/I ratio reflected those in the total proteolytic flux remarkably in both fresh rat hepatocytes and H4-II-E cell lines. Thus, in addition to a sensitive index of macroautophagy, calculating the cytosolic LC3 ratio became an easy and quick quantitative method for monitoring its regulation in hepatocytes and H4-II-E cells.
- Published
- 2007
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