Mitophagy, when defined as the selective removal of damaged
mitochondria, is one of the sexiest mitochondrial research areas. At the same
time, there is little genetic evidence to support the existence of such
phenomenon and most studies are carried out under artificial cell culture
conditions (Kauppila et
al. 2017). Also, I have already lost count of the number of different
mitophagy pathways suggested to exist (Williams et al. 2018). For
these reasons I have stopped reading the mitophagy literature.
Nevertheless, two papers in my publication feed caught my eye both
describing how a new long form of phosphatase and tensin homolog (PTEN-L or
PTENα) regulates mitophagy (Wang et al. 2018, Yin et al. 2018). The
funny thing is that while Yin et al describe PTEN-L to be necessary for
mitophagy, Wang et al. describe the same protein to be a negative regulator of
mitophagy. I think this is a fair reflection of the quality of the contemporary
mitophagy research.
How these papers come to different conclusions? Well, both papers have
many of the common pitfalls in mitophagy research, which are
- Making a subcellular fractionation experiment using only a cytosolic and mitochondrial fractions. Most likely that mitochondrial fractions will have nuclear and microsome contaminations.
- Using the mRNA levels of Tfam, Nrf1 and Ppargc1α as markers for mitochondrial biogenesis. One should test this either using citrate synthase assay, by western or ideally both.
- Using CCCP, a strong protonophore uncoupling all cellular membranes, as a good reflection of mitochondrial damage.
- Using MitoTracker Red to visualize mitochondria under CCCP treatment because CCCP causes the dye to diffuse out from mitochondria into lysosomes (Padman et al. 2013).
I think I will continue ignoring the mitophagy research also in the future.
PS: Being pedantic here, but there is a difference between cytoplasm and cytosol. Cytoplasm is everything between the plasma membrane and nucleus, so it includes mitochondria. Cytosol is everything outside of cellular membranes/vesicles.
References:
Kauppila TES, Kauppila JHK, Larsson NG. Mammalian
Mitochondria and Aging: An Update. Cell Metab. 2017. PMID: 28094012
Padman BS, Bach M, Lucarelli G, Prescott M, Ramm G. The protonophore
CCCP interferes with lysosomal degradation of autophagic cargo in yeast and
mammalian cells. Autophagy.
2013. PMID: 24150213
Wang L, Cho YL, Tang Y, Wang J, Park JE, Wu Y, Wang C, Tong Y, Chawla R,
Zhang J, Shi Y, Deng S, Lu G, Wu Y, Tan HW, Pawijit P, Lim GG, Chan HY, Zhang J,
Fang L, Yu H, Liou YC, Karthik M, Bay BH, Lim KL, Sze SK, Yap CT, Shen HM. PTEN-L
is a novel protein phosphatase for ubiquitin dephosphorylation to inhibit
PINK1-Parkin-mediated mitophagy. Cell Res. 2018. PMID: 29934616
Williams JA, Ding WX. Mechanisms, pathophysiological roles and methods
for analyzing mitophagy - recent insights. Biol Chem. 2018. PMID: 28976892
Yin Y, Li G, Yang J, Yang C, Zhu M, Jin Y, McNutt MA. PTENα Regulates
Mitophagy and Maintains Mitochondrial Quality Control. Autophagy. 2018. PMID: 29969932
You raised some really important points here. They highlight some of the fundamental issues that have (and continue to) plague mitophagy research. I feel that I can speak from a position of authority on this because I'm the author of one of the articles you use for supporting evidence. That publication became the first chapter of my doctoral thesis. In fact, one of my thesis assessors even remarked that my data "convincingly demonstrated that damage-induced mitophagy does not exist." (Sentence has been reworded with redactions to avoid self incrimination).
ReplyDeleteI'm writing this response to you, because I spent years 4 banging my head against a wall in frustration, attempting to replicate the irreproducible mitophagy that continues to plague autophagy research like minefield.
I'm writing this response to you, because I continued to study mitophagy after completing my PhD... even though the final chapter of my thesis includes the sentence "my findings collectively indicate that damage-induced mitophagy does not exist."
I implore you to keep an open mind... an open, but very skeptical mind.
Mitophagy was the victim of being a "hot topic".
People wanted to stamp their name on it without caring if their name was associated with any kind of meaningful contribution.
There was a lot of noise, and it is STILL hard to separate the truth from it.
I won't name-and-shame here, but I gained a deep-seated distrust of almost every other researcher in my field.
I'm glad I stayed in the field though; It's a powerful thing to realize that the established literature is still up for re-evaluation.
There's very little competition when everyone else has accepted something as dogma.
(P.S. let me know if you want any more arguments against the existence of damage-induced mitophagy. I have plenty, but I also had the opportunity to conduct experiments that convinced me otherwise.)
(P.P.S. Mitophagy was the first type of autophagy that was discovered, and that wasnt even what won the nobel prize)