If LC3-I and LC3-II expression both increase in cochlea of aged animal compared to young animal, can we say that autophagy is activated or enhanced? Thanks.
You should have an in-depth reading of the attached articles.
Best regards
Robert
Dear Bo,
My domain of expertise is cancer. So I do not want to speculate on your subject because it is completely out of my expertise.
Best regards
Robert
In general, an increase in LC3-II pool in comparison to LC3-I is a sign of autophagy induction but it is always advisable to normalize the results with additional control such as actin in WB assay. The mentioned control is more important in your experimental design since you are comparing the samples from animals of two different age groups. The publications suggested by Dr Robert Kiss, especially the one by Klionsky et al. 2016, are a great resource of information on autophagy research.
In the cancer area, nobody can today clearly state whether activating autophagy in cancer cells is detrimental or beneficial for these cells. You can find dozens of articles in favor of the first hypothesis and dozens in favor of the second one.
In contrast, it becomes more and more evident that inhibiting autophagy is detrimental for cancer cells.
I do not know what is the situation in an "extra cancer context".
Best regards
Robert
Well said Dr Kiss - I totally agree with you and yes, the role of Autophagy is contextual in various experimental or therapeutic situations. I remember reading a nice editorial piece in this regard and I thought you will also like it:
Gewirtz DA. Cytoprotective and nonprotective autophagy in cancer therapy. Autophagy. 2013 Sep;9(9):1263-5. http://www.tandfonline.com/toc/kaup20/9/9
How LC3-I and LC3-II simultaneously increase; the transcription of LC-3 itself increases, while the LC-3 conversion is promoted.
This is not a exceptional case. When MEFs are treated with HBSS starvation medium more than 16hr or treated with VP-16 (20uM) for more than 12hr, not only LC3-I but also LC3-II increase in amount.
However, in that case, you would have difficulty in determining whether autophagy is promoted or not. In fact, LC3-II/LC3-I ratio is the hallmark of the degree of autophagy activation.
That is why you should also check the expression of p62/SQSTM1.
When p62/SQSTM1 is down-regulated, you would think p62/SQSTM1 is consumed and LC3 conversion is promoted.
In contrast, when p62/SQSTM1 also increases (accumulates), the autophagy flux is considered to be stopped at the process of the lysosome. The disturbed autophagy flux induces ER stress and finally apoptosis.
Article How to Interpret LC3 Immunoblotting
After the treatment with CQ or bafilomycinA1, not only LC3-II but LC3-I are accumulated in cancer cells.
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