MITOtalks and FocalPlane features…. recording

Our recent FocalPlane features… webinar was a collaboration with the popular MITOtalks series and showcased research published in Journal of Cell Science’s Special Issue: Cell Biology of Mitochondria. MITOtalks organisers Sjoerd Wanrooij and Nuno Raimundo hosted the webinar with talks from Yuli Buckley on ‘Fis1 regulates mitochondrial morphology, bioenergetics and removal of mitochondrial DNA damage in irradiated glioblastoma cells’, Mireia Nager on ‘Mitophagy is induced in human engineered heart tissue after simulated ischemia and reperfusion’ and Dikaia Tsagkari on ‘NHR-85 modulates mitochondrial and lipid homeostasis to protect against α-synuclein aggregation’

Thanks for all the speakers for fantastic talks, and to Sjoerd and Nuno for hosting the event along with Heidi McBride, who coordinated JCS’s Cell Biology of Mitochondria Special Issue with Ana Garcia-Saez.

We had a few questions that were not addressed live, and you can find the answers below.

Question for Yuli Buckley:
Why didn’t mitophagy reduce upon Fis1 knockdown? Does it mean that there are other drivers of mitophagy upon IR radiation?

YB: Great question! The canonical mitophagy pathway involves PINK1 and Parkin, whereas Fis1 is involved in the non-canonical pathway. This is most likely why we didn’t see changes in mitophagy, as well as the fact that there wasn’t much damage introduced simply by silencing Fis1. When we combined the treatments, the increase in mitophagy may be due to an increase in the canonical mitophagic pathway, but Fis1 may be critical for the segregation of damage. Also, Fis1 interacts with a protein on the lysosome, leading to untethering of the mito-lysosome connection, so we hypothesize that the tether remains, which could affect mitophagic flux.

Very interesting work, I was wondering if you also checked if there was any alteration in the mitochondrial mass after silencing Fis-1. Also, can you please elaborate more on the signaling that’s involved in Fis-1 mediated fission in pathological conditions?

YB: Thank you! We looked at mito mass using citrate synthase assays and by quantifying mtDNA copy number and didn’t see changes with either approach. With respect to signaling, we are still trying to figure that out. Fis1 PTMs are not well-established. It can be ubiquitinated by MARCH5 or phosphorylated at Serine10, but we are not sure what the phosphorylation does. There are a few other suggested phos sites that may lead to dimerization of Fis1 before it recruits Drp1. Regarding its regulation of bioenergetics, we did see changes to iron-sulfur proteins in the ETC. These ISPs are nuclear-encoded and must be transported into the mito, so there may be some interaction with Fis1 at the outer mito membrane that regulates their import.

Really interesting, have you thought about other strategies for detecting mitochondrial DNA damage? What made you chose this strategy?

YB: Great question, we chose this strategy because another mitochondrial lab had really developed the protocol and was able to help us with troubleshooting. We could also look at nucleoid clustering or Polg expression.

Question for Mireia Nager:
Have you checked how long the changed morphology lasts for after reperfusion?

MN: We start to see a decrease in the number of red only dots after 4 to 6 hours

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