Abstract
A common way to measure the induction of autophagy in yeast and mammalian cells is to compare the amount of Atg8/LC3-I with that of Atg8-PE/LC3-II by using western blot analysis. This is because changes in the amount of LC3-II correlate closely with changes in the number of autophagosomes present in cells. Atg8/LC3 is initially synthesized as an unprocessed form, which is proteolytically processed to form Atg8/LC3-I, and then this is modified into the phosphatidylethanolamine (PE)-conjugated Atg8-PE/LC3-II form. Atg8/LC3-II is membrane bound, whereas Atg8-PE/LC3-I is cytosolic. By associating with both the inner and outer membranes of the autophagosome, Atg8-PE/LC3-II is the only autophagy reporter that is reliably associated with completed autophagosomes. In the nematode Caenorhabditis elegans, the ortholog of Atg8/LC3 is LGG-1. Here, we discuss how changes in the levels of LGG-1-II (and the paralog LGG-2) protein can, with appropriate controls, be used to monitor autophagy activity in nematodes and present a protocol for monitoring changes in the protein levels of different forms of LGG-1 by western blotting.
References
Feb 26, 2000·Molecular Biology of the Cell·C BucciB van Deurs
Feb 22, 2000·The Journal of Biological Chemistry·W P HuangD J Klionsky
Jun 10, 2000·The Journal of Biological Chemistry·T Oka, M Futai
Oct 19, 2000·The Journal of Cell Biology·T KirisakoY Ohsumi
Nov 4, 2000·The EMBO Journal·Y KabeyaT Yoshimori
Sep 13, 2002·Current Biology : CB·Xiaomeng LongJoseph Avruch
Apr 4, 2006·Cell Metabolism·Young-jai YouLeon Avery
May 11, 2006·FEBS Letters·Yon Ju JiJoohong Ahnn
Jul 6, 2007·Autophagy·Noboru Mizushima, Tamotsu Yoshimori
Sep 6, 2007·Genes & Development·Chanhee KangLeon Avery
May 13, 2009·Biology of the Cell·Xavier MicheletRenaud Legouis
Mar 13, 2010·The Journal of Pathology·Sandra BarthKay F Macleod
Jun 5, 2010·Autophagy·Adriana AlbertiRenaud Legouis
Jun 17, 2010·Cell·Ye TianHong Zhang
Aug 2, 2011·Developmental Cell·Qun LuHong Zhang
Nov 16, 2011·Autophagy·Shensi ShenGuido Kroemer
Mar 6, 2012·Journal of Cell Science·Abderazak DjeddiRenaud Legouis
Aug 14, 2012·Autophagy·Qianqian LiangHong Zhang
Jan 1, 2014·Developmental Cell·Marion Manil-SégalenRenaud Legouis
Citations
Sep 5, 2017·Cells·Yanfang ChenRenaud Legouis
Aug 9, 2019·Journal of Experimental & Clinical Cancer Research : CR·Lei YinBo Peng
Sep 3, 2016·Oncology Letters·Qinghua XiaXunbo Jin
Feb 3, 2016·Cold Spring Harbor Protocols·Nicholas J Palmisano, Alicia Meléndez