title: p62 Promotes Amino Acid Sensitivity of mTOR Pathway and Hepatic Differentiation in Adult Liver Stem/Progenitor Cells.
abstract: Autophagy is a homeostatic process regulating turnover of impaired proteins and organelles, and p62 (sequestosome-1, SQSTM1) functions as the autophagic receptor in this process. p62 also functions as a hub for intracellular signaling such as that in the mammalian target of rapamycin (mTOR) pathway. Liver stem/progenitor cells have the potential to differentiate to form hepatocytes or cholangiocytes. In this study, we examined effects of autophagy, p62, and associated signaling on hepatic differentiation. Adult stem/progenitor cells were isolated from the liver of mice with chemically induced liver injury. Effects of autophagy, p62, and related signaling pathways on hepatic differentiation were investigated by silencing the genes for autophagy protein 5 (ATG5) and/or SQSTM1/p62 using small interfering RNAs. Hepatic differentiation was assessed based on increased albumin and hepatocyte nuclear factor 4α, as hepatocyte markers, and decreased cytokeratin 19 and SOX9, as stem/progenitor cell markers. These markers were measured using quantitative RT-PCR, immunofluorescence, and Western blotting. ATG5 silencing decreased active LC3 and increased p62, indicating inhibition of autophagy. Inhibition of autophagy promoted hepatic differentiation in the stem/progenitor cells. Conversely, SQSTM1/p62 silencing impaired hepatic differentiation. A suggested mechanism for p62-dependent hepatic differentiation in our study was activation of the mTOR pathway by amino acids. Amino acid activation of mTOR signaling was enhanced by ATG5 silencing and suppressed by SQSTM1/p62 silencing. Our findings indicated that promoting amino acid sensitivity of the mTOR pathway is dependent on p62 accumulated by inhibition of autophagy and that this process plays an important role in the hepatic differentiation of stem/progenitor cells. J. Cell. Physiol. 232: 2112-2124, 2017. © 2016 Wiley Periodicals, Inc.
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Masakazu
authors.initials: M
authors.lastname: Sugiyama
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Tomoharu
authors.initials: T
authors.lastname: Yoshizumi
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Yoshida
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Bekki
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Matsumoto
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Shohei
authors.initials: S
authors.lastname: Yoshiya
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Takeo
authors.initials: T
authors.lastname: Toshima
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Ikegami
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Shinji
authors.initials: S
authors.lastname: Itoh
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Norifumi
authors.initials: N
authors.lastname: Harimoto
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Shinji
authors.initials: S
authors.lastname: Okano
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Soejima
authors.affiliation: Department of Hepato-Biliary-Pancreatic Surgery, Graduate School of Medicine, Gunma University, Maebashi, Japan.
authors.forename: Ken
authors.initials: K
authors.lastname: Shirabe
authors.affiliation: Department of Surgery and Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Yoshihiko
authors.initials: Y
authors.lastname: Maehara
mesh_terms: D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D001343:Autophagy; D000071187:Autophagy-Related Protein 5 / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D019070:Cell Lineage; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D056486:Chemical and Drug Induced Liver Injury / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D022781:Hepatocytes / Q000201:enzymology* / Q000473:pathology; D008099:Liver / Q000201:enzymology* / Q000473:pathology; D008297:Male; D008810:Mice, Inbred C57BL; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D034622:RNA Interference; D000071456:Sequestosome-1 Protein / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D013234:Stem Cells / Q000201:enzymology* / Q000473:pathology; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection
chemical_list: D000596:Amino Acids; C495773:Atg5 protein, mouse; D000071187:Autophagy-Related Protein 5; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D000071456:Sequestosome-1 Protein; C101343:Sqstm1 protein, mouse; C546843:mTOR protein, mouse; D058570:TOR Serine-Threonine Kinases