Abstract
LC3 is widely used marker for macroautophagy assays. After translation pro-LC3 is processed by Atg4 to expose C-terminal glycine residue for downstream conjugation reactions to accomplish the conversion of LC3-I to LC3-II. SDS-PAGE based Western blot (Wb) is generally utilized to quantify LC3-II levels where the LC3-I band migrates slower than LC3-II. We found that pro-human LC3B migrated at similar rate as LC3B-II in SDS-PAGE. The carboxyl-terminal five amino acids, particularly Lysine122 and Leucine123 of human LC3B play a major role in the faster migration of unprocessed LC3B, rendering it indistinguishable from LC3B-II in Wb assays. The unique faster migration of unprocessed LC3B than LC3B-I is also revealed in mouse LC3B, rat LC3B and rat LC3 but not in human LC3C. Our findings for the first time define pro-LC3 migration patterns for LC3 family member from human, mouse and rat species in SDS-PAGE. These findings provide a reference for pro-LC3 band patterns when Atg4 function is inhibited.
Highlights
Macroautophagy is a highly conserved cellular degradation process
Human LC3BG120A Migrates at a Similar Rate as LC3B-II
We demonstrated that unprocessed human LC3B migrates at a similar rate as LC3B-II in SDS- polyacrylamide gel
Summary
Macroautophagy ( referred to as autophagy) is a highly conserved cellular degradation process. In contrast to the ubiquitin-proteasome degradation system, two ubiquitin-like conjugation systems are essential for autophagy: Atg12-Atg and Atg8/LC3- phosphatidylethanolamine (PE) [1,2]. Atg8/LC3 lipidation involves a cysteine protease, Atg, to process pro-LC3 exposing a C-terminal glycine residue to generate LC3-I. LC3-I is activated by Atg as a thioester bond is formed between a catalytic cysteine residue of Atg and the C-terminal glycine of LC3-I. LC3-Atg intermediate afterward exchanges with Atg, an E2-like enzyme, to form a second thioester intermediate, LC3-Atg3 [3]. The lipidation process is completed by the putative E3-like enzyme Atg16L1NAtg5-Atg complex to form LC3-II, as LC3-Atg is exchanged with PE [4]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
