文献詳細
文献概要
今月の主題 オートファジー 総論
オートファジーの検出方法―原理と課題
著者: 野田健司1
所属機関: 1大阪大学微生物学研究所細胞制御分野
ページ範囲:P.1527 - P.1534
文献購入ページに移動 オートファジーとは基本的に微量な分解過程であり,分解そのものを検出することは容易ではない.そこで,その活性を捉えるために,様々な方法が考案されてきている.本稿では,歴史的な方法から今日幅広く用いられている方法,さらに最新の方法まで,オートファジーの検出方法の原理,およびその課題について概説する.オートファジーにより輸送・分解される基質の量を測定する定量的な方法と,オートファジーの進行に付随して起こるLC3などの分子の変動を検出する半定量的な方法がある.オートファジーの調節機構の詳細な解析には定量的な方法が望ましい.一方,近年開発された半定量的な方法により様々な生命現象にオートファジーがかかわることが明らかにされてきた.
参考文献
1) Kopitz J, Kisen GO, Gordon PB, et al:Nonselective autophagy of cytosolic enzymes by isolated rat hepatocytes. J Cell Biol 111:941-953, 1990
2) Seglen PO, Gordon PB, Tolleshaug H, et al:Use of [3H] raffinose as a specific probe of autophagic sequestration. Exp Cell Res 162:273-277, 1986
3) Høyvik H, Gordon PB, Seglen PO:Use of a hydrolysable probe, [14C] lactose, to distinguish between pre-lysosomal and lysosomal steps in the autophagic pathway. Exp Cell Res 166:1-14, 1986
4) Mortimore GE, Mondon CE:Inhibition by insulin of valine turnover in liver. Evidence for a general control of proteolysis. J Biol Chem 245:2375-2383, 1970
5) Noda T, Matsuura A, Wada Y, et al:Novel system for monitoring autophagy in the yeast Saccharomyces cerevisiae. Biochem Biophys Res Commun 210:126-132, 1995
6) Kabeya Y, Mizushima N, Ueno T, et al:LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing. EMBO J 19:5720-5728, 2000
7) Mizushima N, Yamamoto A, Matsui M, et al:In vivo analysis of autophagy in response to nutrient starvation using transgenic mice expressing a fluorescent autophagosome marker. Mol Biol Cell 15:1101-1111, 2004
8) Kuma A, Matsui M, Mizushima N:LC3, an autophagosome marker, can be incorporated into protein aggregates independent of autophagy:caution in the interpretation of LC3 localization. Autophagy 3:323-328, 2007
9) Kimura S, Noda T, Yoshimori T:Dissection of the autophagosome maturation process by a novel reporter protein, tandem fluorescent-tagged LC3. Autophagy 3:452-460, 2007
10) Shvets E, Fass E, Elazar Z:Utilizing flow cytometry to monitor autophagy in living mammalian cells. Autophagy 4:621-628, 2008
11) Mizushima N, Yoshimori T:How to interpret LC3 immunoblotting. Autophagy 3:542-545, 2007
12) Tanida I, Minematsu-Ikeguchi N, Ueno T, et al:Lysosomal turnover, but not a cellular level, of endogenous LC3 is a marker for autophagy. Autophagy 1:84-91, 2005
13) Mizushima N, Yamamoto A, Hatano M, et al:Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells. J Cell Biol 152:657-668, 2001
14) Komatsu M, Waguri S, Koike M, et al:Homeostatic levels of p62 control cytoplasmic inclusion body formation in autophagy-deficient mice. Cell 131:1149-1163, 2007
15) Itakura E, Kishi C, Inoue K, et al:Beclin 1 forms two distinct phosphatidylinositol 3-kinase complexes with mammalian Atg14 and UVRAG. Mol Biol Cell 19:5360-5372, 2008
16) Matsunaga K, Saitoh T, Tabata K, et al:Two Beclin 1-binding proteins, Atg14L and Rubicon, reciprocally regulate autophagy at different stages. Nat Cell Biol 11:385-396, 2009
17) Hosokawa N, Hara Y, Mizushima N:Generation of cell lines with tetracycline-regulated autophagy and a role for autophagy in controlling cell size. FEBS Lett 581:2623-2629, 2007
18) Fujita N, Hayashi-Nishino M, Fukumoto H, et al:An Atg4B mutant hampers the lipidation of LC3 paralogues and causes defects in autophagosome closure. Mol Biol Cell 19:4651-4659, 2008
19) Petiot A, Ogier-Denis E, Blommaart EF, et al:Distinct classes of phosphatidylinositol 3'-kinases are involved in signaling pathways that control macroautophagy in HT-29 cells. J Biol Chem 275:992-998, 2000
20) Klionsky DJ, Elazar Z, Seglen PO, et al:Does bafilomycin A1 block the fusion of autophagosomes with lysosomes? Autophagy 4:849-850, 2008
21) Noda T, Ohsumi Y:Tor, a phosphatidylinositol kinase homologue, controls autophagy in yeast. J Biol Chem 273:3963-3966, 1998
22) Thoreen CC, Kang SA, Chang JW, et al:An ATP-competitive mammalian target of rapamycin inhibitor reveals rapamycin-resistant functions of mTORC1. J Biol Chem 284:8023-8032, 2009
23) Mizushima N:Methods for monitoring autophagy. Int J Biochem Cell Biol 36:2491-2502, 2004
24) Klionsky DJ, Abeliovich H, Agostinis P, et al:Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes. Autophagy 4:151-175, 2008
25) Klionsky DJ, Cuervo AM, Seglen PO:Methods for monitoring autophagy from yeast to human. Autophagy 3:181-206, 2007
26) Klionsky DJ, ed.:Methods Enzymol 452, Autophagy in mammalian systems, Part B. Preface, Elsevier, Amsterdam, 2009
27) Deretic V, ed.:Methods in Molecular Biology 445, Autophagosome and phagosome, Springer, Heidelberg, 2008
掲載誌情報