Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes

Daniel J. Klionsky, Hagai Abeliovich, Patrizia Agostinis, Devendra K. Agrawal, Gjumrakch Aliev, David S. Askew, Misuzu Baba, Eric H. Baehrecke, Ben A. Bahr, Andrea Ballabio, Bruce A. Bamber, Diane C. Bassham, Ettore Bergamini, Xiaoning Bi, Martine Biard-Piechaczyk, Janice S. Blum, Dale E. Bredesen, Jeffrey L. Brodsky, John H. Brumell, Ulf T. BrunkWilfried Bursch, Nadine Camougrand, Eduardo Cebollero, Francesco Cecconi, Yingyu Chen, Lih Shen Chin, Augustine Choi, Charleen T. Chu, Jongkyeong Chung, Peter G.H. Clarke, Robert S.B. Clark, Steven G. Clarke, Corinne Clavé, John L. Cleveland, Patrice Codogno, María I. Colombo, Ana Cotomontes, James M. Cregg, Ana Maria Cuervo, Jayanta Debnath, Francesca Demarchi, Patrick B. Dennis, Phillip A. Dennis, Vojo Deretic, Rodney J. Devenish, Federica Di Sano, J. Fred Dice, Marian DiFiglia, Savithramma Dinesh-Kumar, Clark W. Distelhorst, Mojgan Djavaheri-Mergny, Frank C. Dorsey, Wulf Dröge, Michel Dron, William A. Dunn, Michael Duszenko, N. Tony Eissa, Zvulun Elazar, Audrey Esclatine, Eeva Liisa Eskelinen, László Fésüs, Kim D. Finley, José M. Fuentes, Juan Fueyo, Kozo Fujisaki, Brigitte Galliot, Fen Biao Gao, David A. Gewirtz, Spencer B. Gibson, Antje Gohla, Alfred L. Goldberg, Ramon Gonzalez, Cristina González-Estévez, Sharon Gorski, Roberta A. Gottlieb, Dieter Häussinger, You Wen He, Kim Heidenreich, Joseph A. Hill, Maria Høyer-Hansen, Xun Hu, Wei Pang Huang, Akiko Iwasaki, Marja Jäättelä, William T. Jackson, Xuejun Jiang, Shengkan Jin, Terje Johansen, Jae U. Jung, Motoni Kadowaki, Chanhee Kang, Ameeta Kelekar, David H. Kessel, Jan A.K.W. Kiel, Pyo Kim Hong, Adi Kimchi, Timothy J. Kinsella, Kirill Kiselyov, Katsuhiko Kitamoto, Erwin Knecht, Masaaki Komatsu, Eiki Kominami, Seiji Kondo, Attila L. Kovács, Guido Kroemer, Chia Yi Kuan, Rakesh Kumar, Mondira Kundu, Jacques Landry, Marianne Laporte, Weidong Le, Huan Yao Lei, Michael J. Lenardo, Beth Levine, Andrew Lieberman, Kah Leong Lim, Fu Cheng Lin, Willisa Liou, Leroy F. Liu, Gabriel Lopez-Berestein, Carlos López-Otín, Bo Lu, Kay F. Macleod, Walter Malorni, Wim Martinet, Ken Matsuoka, Josef Mautner, Alfred J. Meijer, Alicia Meléndez, Paul Michels, Giovanni Miotto, Wilhelm P. Mistiaen, Noboru Mizushima, Baharia Mograbi, Iryna Monastyrska, Michael N. Moore, Paula I. Moreira, Yuji Moriyasu, Tomasz Motyl, Christian Münz, Leon O. Murphy, Naweed I. Naqvi, Thomas P. Neufeld, Ichizo Nishino, Ralph A. Nixon, Takeshi Noda, Bernd Nürnberg, Michinaga Ogawa, Nancy L. Oleinick, Laura J. Olsen, Bulent Ozpolat, Shoshana Paglin, Glen E. Palmer, Issidora Papassideri, Miles Parkes, David H. Perlmutter, George Perry, Mauro Piacentini, Ronit Pinkas-Kramarski, Mark Prescott, Tassula Proikascezanne, Nina Raben, Abdelhaq Rami, Fulvio Reggiori, Bärbel Rohrer, David C. Rubinsztein, Kevin M. Ryan, Junichi Sadoshima, Hiroshi Sakagami, Yasuyoshi Sakai, Marco Sandri, Chihiro Sasakawa, Miklos Sass, Claudio Schneider, Per O. Seglen, Oleksandr Seleverstov, Jeffrey Settleman, John J. Shacka, Irving M. Shapiro, Andrei Sibirny, Elaine C.M. Silva-Zacarin, Hans Uwe Simon, Crisfiano Simone, Anne Simonsen, Mark A. Smith, Katharina Spanel-Borowski, Vickram Srinivas, Meredith Steeves, Harald Stenmark, Per E. Stromhaug, Carlos S. Subauste, Seiichiro Sugimoto, David Sulzer, Toshihiko Suzuki, Michele S. Swanson, Ira Tabas, Fumihiko Takeshita, Nicholas J. Talbot, Zsolt Tallóczy, Keiji Tanaka, Kozo Tanaka, Isei Tanida, Graham S. Taylor, J. Paul Taylor, Alexei Terman, Gianluca Tettamanti, Craig B. Thompson, Michael Thumm, Aviva M. Tolkovsky, Sharon A. Tooze, Ray Truant, Lesya V. Tumanovska, Yasuo Uchiyama, Takashi Ueno, Néstor L. Uzcátegui, Ida Van Der Klei, Eva C. Vaquero, Tibor Vellai, Michael W. Vogel, Hong Gang Wang, Paul Webster, John W. Wiley, Zhijun Xi, Gutian Xiao, Joachim Yahalom, Jin Ming Yang, George Yap, Xiao Ming Yin, Tamotsu Yoshimori, Li Yu, Zhenyu Yue, Michisuke Yuzaki, Olga Zabirnyk, Xiaoxiang Zheng, Xiongwei Zhu, Russell L. Deter

Research output: Contribution to journalReview article

1788 Scopus citations

Abstract

Research in autophagy continues to accelerate,1 and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.2,3 There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from fully functional autophagy that includes delivery to, and degradation within, lysosomes (in most higher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for the selection and interpretation of the methods that can be used by investigators who are attempting to examine macroautophagy and related processes, as well as by reviewers who need to provide realistic and reasonable critiques of papers that investigate these processes. This set of guidelines is not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to verify an autophagic response.

Original languageEnglish (US)
Pages (from-to)151-175
Number of pages25
JournalAutophagy
Volume4
Issue number2
DOIs
StatePublished - Feb 16 2008

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Keywords

  • Autolysosome
  • Autophagosome
  • Flux
  • Lysosome
  • Phagophore
  • Stress
  • Vacuole

ASJC Scopus subject areas

  • Cell Biology
  • Molecular Biology

Cite this

Klionsky, D. J., Abeliovich, H., Agostinis, P., Agrawal, D. K., Aliev, G., Askew, D. S., Baba, M., Baehrecke, E. H., Bahr, B. A., Ballabio, A., Bamber, B. A., Bassham, D. C., Bergamini, E., Bi, X., Biard-Piechaczyk, M., Blum, J. S., Bredesen, D. E., Brodsky, J. L., Brumell, J. H., ... Deter, R. L. (2008). Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes. Autophagy, 4(2), 151-175. https://doi.org/10.4161/auto.5338