Hello,
I am treating my cells with a compound to study whether the autophagy can be affected or not. Using Western blot, I found that LC3 II form remains unchanged but p62 protein is increasing . Any explanations ?
Thank you.
P62 represents a negative correlation protein to autophagy. Because p62 is marking autophasosomes and the increment in p62 level suggests the accumulation interruption of lysosomal fusion and degradation. LC3 II is widely used as a marker for autophagy induction and if we consider an increased p62 we should expect a decreased in the conversion of LC3 I into LC3 II. So you may find the ratio of lower band that represents LC3 I over the upper band that represents LC3 II. I would expect increase in that ratio as similar as p62 if you are aiming to inhibit autophagy, hope it helps
While the new edition is still in the oven, I suggest you study the article below:
Klionsky DJ, Abdelmohsen K, Abe A, et al. Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition) [published correction appears in Autophagy. 2016;12(2):443. Autophagy. 2016;12(1):1-222.
I agree with the previous comments, but the literature has shown that the LC3-II / LC3-I ratio has not been considered a good parameter for the interpretation of the autophagy flow. In my previous experience, I obtained results (without the use of drugs that inhibit or activate autophagy) in which there was an increase in LC3-II concomitant with an increase in p62. My choice to better interpret my data was to evaluate other proteins from the different stages of the autophagy flow, such as beclin1, LAMP2 and CHMP2B. As well as immunofluorescence analyzes and transmission electron microscopy.
I reinforce the suggestion of reading made by Prof. Dr. Rafael Linden, authored by Klionsky et. al.
Good luck with your analysis.
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